https://proteopedia.org/api.php?action=feedcontributions&feedformat=atom&user=Samer+KawakProteopedia - User contributions [en]2026-09-25T06:27:26ZUser contributionsMediaWiki 1.43.8https://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231072Samer Kawak sandbox 22011-04-18T03:40:20Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis with FBPA circled]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate (FBP) into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi. Due to the function of catalyzing the formation and cleavage of a carbon-carbon bond between various substrates, aldolases are considered a possible biocatalysts. <ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While FBPA can exist as a monomer, eukaryotic FBPA I, particularly, exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>, in which the subunits assume the fold of a parallel (βα)8-(TIM)-barrel. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. FBPA is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main chain nitrogens of <scene name='Samer_Kawak_sandbox_2/Ser271/2'>Ser271</scene> and <scene name='Samer_Kawak_sandbox_2/Gly272/1'>Gly272</scene> hold the 1-phosphate group while the <scene name='Samer_Kawak_sandbox_2/Lys41/1'>Lys41</scene>, <scene name='Samer_Kawak_sandbox_2/Arg42/1'>Arg42</scene> and <scene name='Samer_Kawak_sandbox_2/Arg303/1'>Arg303</scene> residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (<scene name='Samer_Kawak_sandbox_2/Tyr363/1'>Tyr363</scene>) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldol condensation. Catalysis occurs first when the nucleophilic <scene name='Samer_Kawak_sandbox_2/Lys229/2'>ε-amine group of Lys229</scene> attacks the carbonyl carbon of the substrate FBP in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
Kinetic studies have help to offer a great understanding of the intricacies of the active site. Generally in solution, the substrate FBP exists as a furanose ring, but in order for the aldolase reaction to take place, a ring-opened ketose is needed. The ring-opened ketose can be polarized through the carbonyl via a metal ion (FBPA II) or react to establish a Schiff-base intermediate (FBPA II). Overall, the rate of the Schiff-base formation is faster than the sugar ring-opening in solution.<ref name="kinetics2"> Choi, K., and Tolan, D. “Presteady-State Kinetic Evidence for a Ring-Opening Activity in Fructose-1,6-(bis)phosphate Aldolase.” J. Am. Chem. Soc. 126. (2004): 3402-3403</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231066Samer Kawak sandbox 22011-04-18T03:25:21Z<p>Samer Kawak: /* Kinetics */</p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate (FBP) into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi. Due to the function of catalyzing the formation and cleavage of a carbon-carbon bond between various substrates, aldolases are considered a possible biocatalysts. <ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While FBPA can exist as a monomer, eukaryotic FBPA I, particularly, exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>, in which the subunits assume the fold of a parallel (βα)8-(TIM)-barrel. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. FBPA is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main chain nitrogens of <scene name='Samer_Kawak_sandbox_2/Ser271/2'>Ser271</scene> and <scene name='Samer_Kawak_sandbox_2/Gly272/1'>Gly272</scene> hold the 1-phosphate group while the <scene name='Samer_Kawak_sandbox_2/Lys41/1'>Lys41</scene>, <scene name='Samer_Kawak_sandbox_2/Arg42/1'>Arg42</scene> and <scene name='Samer_Kawak_sandbox_2/Arg303/1'>Arg303</scene> residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (<scene name='Samer_Kawak_sandbox_2/Tyr363/1'>Tyr363</scene>) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldol condensation. Catalysis occurs first when the nucleophilic <scene name='Samer_Kawak_sandbox_2/Lys229/2'>ε-amine group of Lys229</scene> attacks the carbonyl carbon of the substrate FBP in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
Kinetic studies have help to offer a great understanding of the intricacies of the active site. Generally in solution, the substrate FBP exists as a furanose ring, but in order for the aldolase reaction to take place, a ring-opened ketose is needed. The ring-opened ketose can be polarized through the carbonyl via a metal ion (FBPA II) or react to establish a Schiff-base intermediate (FBPA II). Overall, the rate of the Schiff-base formation is faster than the sugar ring-opening in solution.<ref name="kinetics2"> Choi, K., and Tolan, D. “Presteady-State Kinetic Evidence for a Ring-Opening Activity in Fructose-1,6-(bis)phosphate Aldolase.” J. Am. Chem. Soc. 126. (2004): 3402-3403</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231065Samer Kawak sandbox 22011-04-18T03:24:55Z<p>Samer Kawak: /* Kinetics */</p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate (FBP) into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi. Due to the function of catalyzing the formation and cleavage of a carbon-carbon bond between various substrates, aldolases are considered a possible biocatalysts. <ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While FBPA can exist as a monomer, eukaryotic FBPA I, particularly, exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>, in which the subunits assume the fold of a parallel (βα)8-(TIM)-barrel. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. FBPA is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main chain nitrogens of <scene name='Samer_Kawak_sandbox_2/Ser271/2'>Ser271</scene> and <scene name='Samer_Kawak_sandbox_2/Gly272/1'>Gly272</scene> hold the 1-phosphate group while the <scene name='Samer_Kawak_sandbox_2/Lys41/1'>Lys41</scene>, <scene name='Samer_Kawak_sandbox_2/Arg42/1'>Arg42</scene> and <scene name='Samer_Kawak_sandbox_2/Arg303/1'>Arg303</scene> residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (<scene name='Samer_Kawak_sandbox_2/Tyr363/1'>Tyr363</scene>) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldol condensation. Catalysis occurs first when the nucleophilic <scene name='Samer_Kawak_sandbox_2/Lys229/2'>ε-amine group of Lys229</scene> attacks the carbonyl carbon of the substrate FBP in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
<br />
Kinetic studies have help to offer a great understanding of the intricacies of the active site. Generally in solution, the substrate FBP exists as a furanose ring, but in order for the aldolase reaction to take place, a ring-opened ketose is needed. The ring-opened ketose can be polarized through the carbonyl via a metal ion (FBPA II) or react to establish a Schiff-base intermediate (FBPA II). Overall, the rate of the Schiff-base formation is faster than the sugar ring-opening in solution.<ref name="kinetics2"> Choi, K., and Tolan, D. “Presteady-State Kinetic Evidence for a Ring-Opening Activity in Fructose-1,6-(bis)phosphate Aldolase.” J. Am. Chem. Soc. 126. (2004): 3402-3403</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231056Samer Kawak sandbox 22011-04-18T02:52:01Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate (FBP) into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi. Due to the function of catalyzing the formation and cleavage of a carbon-carbon bond between various substrates, aldolases are considered a possible biocatalysts. <ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While FBPA can exist as a monomer, eukaryotic FBPA I, particularly, exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>, in which the subunits assume the fold of a parallel (βα)8-(TIM)-barrel. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. FBPA is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main chain nitrogens of <scene name='Samer_Kawak_sandbox_2/Ser271/2'>Ser271</scene> and <scene name='Samer_Kawak_sandbox_2/Gly272/1'>Gly272</scene> hold the 1-phosphate group while the <scene name='Samer_Kawak_sandbox_2/Lys41/1'>Lys41</scene>, <scene name='Samer_Kawak_sandbox_2/Arg42/1'>Arg42</scene> and <scene name='Samer_Kawak_sandbox_2/Arg303/1'>Arg303</scene> residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (<scene name='Samer_Kawak_sandbox_2/Tyr363/1'>Tyr363</scene>) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldol condensation. Catalysis occurs first when the nucleophilic <scene name='Samer_Kawak_sandbox_2/Lys229/2'>ε-amine group of Lys229</scene> attacks the carbonyl carbon of the substrate FBP in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231055Samer Kawak sandbox 22011-04-18T02:49:43Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate (FBP) into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi. Due to the function of catalyzing the formation and cleavage of a carbon-carbon bond between various substrates, aldolases are considered a possible biocatalysts. <ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While FBPA can exist as a monomer, eukaryotic FBPA I, particularly, exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>, in which the subunits assume the fold of a parallel (βα)8-(TIM)-barrel. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. FBPA is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main chain nitrogens of <scene name='Samer_Kawak_sandbox_2/Ser271/2'>Ser271</scene> and <scene name='Samer_Kawak_sandbox_2/Gly272/1'>Gly272</scene> hold the 1-phosphate group while the <scene name='Samer_Kawak_sandbox_2/Lys41/1'>Lys41</scene>, <scene name='Samer_Kawak_sandbox_2/Arg42/1'>Arg42</scene> and <scene name='Samer_Kawak_sandbox_2/Arg303/1'>Arg303</scene> residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (<scene name='Samer_Kawak_sandbox_2/Tyr363/1'>Tyr363</scene>) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis. <br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldol condensation. Catalysis occurs first when the nucleophilic <scene name='Samer_Kawak_sandbox_2/Lys229/1'>ε-amine group of Lys229</scene> attacks the carbonyl carbon of the substrate FBP in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231053Samer Kawak sandbox 22011-04-18T02:29:50Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate (FBP) into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi. Due to the function of catalyzing the formation and cleavage of a carbon-carbon bond between various substrates, aldolases are considered a possible biocatalysts. <ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While FBPA can exist as a monomer, eukaryotic FBPA I, particularly, exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>, in which the subunits assume the fold of a parallel (βα)8-(TIM)-barrel. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. FBPA is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main chain nitrogens of <scene name='Samer_Kawak_sandbox_2/Ser271/2'>Ser271</scene> and <scene name='Samer_Kawak_sandbox_2/Gly272/1'>Gly272</scene> hold the 1-phosphate group while the <scene name='Samer_Kawak_sandbox_2/Lys41/1'>Lys41</scene>, <scene name='Samer_Kawak_sandbox_2/Arg42/1'>Arg42</scene> and <scene name='Samer_Kawak_sandbox_2/Arg303/1'>Arg303</scene> residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (<scene name='Samer_Kawak_sandbox_2/Tyr363/1'>Tyr363</scene>) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldol condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate FBP in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231051Samer Kawak sandbox 22011-04-18T02:10:29Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate (FBP) into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi. Due to the function of catalyzing the formation and cleavage of a carbon-carbon bond between various substrates, aldolases are considered a possible biocatalysts. <ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While FBPA can exist as a monomer, eukaryotic FBPA I, particularly, exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>, in which the subunits assume the fold of a parallel (βα)8-(TIM)-barrel. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. FBPA is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main chain nitrogens of <scene name='Samer_Kawak_sandbox_2/Ser271/2'>Ser271</scene> and <scene name='Samer_Kawak_sandbox_2/Gly272/1'>Gly272</scene> hold the 1-phosphate group while the <scene name='Samer_Kawak_sandbox_2/Lys41/1'>Lys41</scene>, <scene name='Samer_Kawak_sandbox_2/Arg42/1'>Arg42</scene> and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldol condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate FBP in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231050Samer Kawak sandbox 22011-04-18T01:56:04Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate (FBP) into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi. Due to the function of catalyzing the formation and cleavage of a carbon-carbon bond between various substrates, aldolases are considered a possible biocatalysts. <ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While FBPA can exist as a monomer, eukaryotic FBPA I, particularly, exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>, in which the subunits assume the fold of a parallel (βα)8-(TIM)-barrel. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. FBPA is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main chain nitrogens of <scene name='Samer_Kawak_sandbox_2/Ser271/2'>Ser271</scene> and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldol condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate FBP in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231044Samer Kawak sandbox 22011-04-18T01:26:58Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate (FBP) into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi. Due to the function of catalyzing the formation and cleavage of a carbon-carbon bond between various substrates, aldolases are considered a possible biocatalysts. <ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While FBPA can exist as a monomer, eukaryotic FBPA I, particularly, exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>, in which the subunits assume the fold of a parallel (βα)8-(TIM)-barrel. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. FBPA is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldol condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate FBP in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231043Samer Kawak sandbox 22011-04-18T01:15:27Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi. Due to the function of catalyzing the formation and cleavage of a carbon-carbon bond between various substrates, aldolases are considered a possible biocatalysts. <ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an α- or β- protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231040Samer Kawak sandbox 22011-04-18T00:58:33Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPAs I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPAs II utilize divalent metal ions and are only found in bacteria and fungi.<ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an α- or β- protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231039Samer Kawak sandbox 22011-04-18T00:49:18Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPA I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPA II utilize divalent metal ions and are only found in bacteria and fungi.<ref name="Schiff">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an α- or β- protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1231037Samer Kawak sandbox 22011-04-18T00:44:38Z<p>Samer Kawak: /* Introduction */</p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase (FBPA) is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref><br />
<br />
There are two classes of FBPAs including Class 1 FBPAs and Class 2 FBPAs. Class 1 FBPAs, or FBPA I, employ a Schiff base reaction mechanism found in most organisms. On the other hand, FBPA II utilize divalent metal ions and are only found in bacteria and fungi.<ref name="book">Lorentzen, E., Siebers, B., Hensel, R., and Pohl, E. “Mechanism of the Schiff base forming fructose-1,6-bisphosphate aldolase: structural analysis of reaction intermediates.” Biochem. J. 44. (2005): 4222-4229.</ref><br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an α- or β- protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230861Samer Kawak sandbox 22011-04-17T01:37:47Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process also generates two net ATP. <br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase in respect to gluconeogenesis or glycolysis.<ref name="book">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref> <br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an α- or β- protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labeling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230856Samer Kawak sandbox 22011-04-17T01:08:02Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
[[Image:Aldolase_reaction.png|400px|left|thumb| The reaction mechanism of aldolase.]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230855Samer Kawak sandbox 22011-04-17T01:06:49Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
<br />
[[Image:Aldolase_reaction.png|400px|left|thumb| Overview of Each Step of Glycolysis]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=File:Aldolase_reaction.png&diff=1230853File:Aldolase reaction.png2011-04-17T01:05:41Z<p>Samer Kawak: </p>
<hr />
<div></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230852Samer Kawak sandbox 22011-04-17T01:04:31Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
<br />
{|<br />
|[[Image:ALDO reaction.png|thumb|400px|<div style="border-width: 0px; border-bottom: 1px solid black; text-align: left;">'''The reaction mechanism of aldolase'''.<br />The enzyme's reactive site amino acid's side-chains are shown in <span style="color: blue; font-weight: bold;">blue</span>.</div>Abbreviations: DHAP - dihydroxyacetone phosphate; Fru1,6bP - Fructose-1,6-bisphosphate; GAD - glyceraldehyde 3-phosphate;]]<br />
|}<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230850Samer Kawak sandbox 22011-04-17T01:03:30Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
[[Image:ALDO reaction.png|thumb|400px|<div style="border-width: 0px; border-bottom: 1px solid black; text-align: left;">'''The reaction mechanism of aldolase'''.<br />The enzyme's reactive site amino acid's side-chains are shown in <span style="color: blue; font-weight: bold;">blue</span>.</div>Abbreviations: DHAP - dihydroxyacetone phosphate; Fru1,6bP - Fructose-1,6-bisphosphate; GAD - glyceraldehyde 3-phosphate;]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230848Samer Kawak sandbox 22011-04-17T01:02:42Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
[[Image:ALDO reaction.png|400px|left|thumb| The reaction mechanism of aldolase]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230847Samer Kawak sandbox 22011-04-17T01:00:49Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
<br />
{|<br />
|[[Image:ALDO reaction.png|thumb|400px|<div style="border-width: 0px; border-bottom: 1px solid black; text-align: left;">'''The reaction mechanism of aldolase'''.<br />The enzyme's reactive site amino acid's side-chains are shown in <span style="color: blue; font-weight: bold;">blue</span>.</div>Abbreviations: DHAP - dihydroxyacetone phosphate; Fru1,6bP - Fructose-1,6-bisphosphate; GAD - glyceraldehyde 3-phosphate;]]<br />
|}<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230846Samer Kawak sandbox 22011-04-17T00:56:00Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|left|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
<br />
[[Image:Aldolase1.jpg]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230845Samer Kawak sandbox 22011-04-17T00:55:34Z<p>Samer Kawak: </p>
<hr />
<div>[[Image:Glycolysis_overview.png|350px|right|thumb| Overview of Each Step of Glycolysis]]<br />
{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
<br />
[[Image:Aldolase1.jpg]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230843Samer Kawak sandbox 22011-04-17T00:54:32Z<p>Samer Kawak: </p>
<hr />
<div>{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction==<br />
[[Image:Glycolysis_overview.png|350px|right|thumb| Overview of Each Step of Glycolysis]]<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
==Structure==<br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
<br />
[[Image:Aldolase1.jpg]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230842Samer Kawak sandbox 22011-04-17T00:52:51Z<p>Samer Kawak: </p>
<hr />
<div>{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction and Structure==<br />
<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
[[Image:Glycolysis_overview.png|350px|right|thumb| Overview of Each Step of Glycolysis]]<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
<br />
[[Image:Aldolase1.jpg]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1230841Samer Kawak sandbox 22011-04-17T00:51:50Z<p>Samer Kawak: </p>
<hr />
<div>{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction and Structure==<br />
<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
[[Image:2000px-Glycolysis_overview.png|350px|right|thumb| Overview of Each Step of Glycolysis]]<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
<br />
[[Image:Aldolase1.jpg]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=File:Glycolysis_overview.png&diff=1230840File:Glycolysis overview.png2011-04-17T00:49:18Z<p>Samer Kawak: </p>
<hr />
<div></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_Sandbox_3&diff=1224770Samer Kawak Sandbox 32011-04-04T03:40:26Z<p>Samer Kawak: New page: {{STRUCTURE_3gpd | PDB=3gpd | SCENE= }} Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a very important enzyme in the production of energy. This enzyme catalyzes the sixth step i...</p>
<hr />
<div>{{STRUCTURE_3gpd | PDB=3gpd | SCENE= }}<br />
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a very important enzyme in the production of energy. This enzyme catalyzes the sixth step in the process of breaking down glucose into energy, also known as glycolysis. Though this is its main function, GAPDH has been shown to perform other functions including transcription activation, initiation of apoptosis, and ER to Golgi apparatus vesicle transportation. However, this page will focus on GAPDH’s role in glycolysis.<br />
GAPDH most commonly exists as what looks to be a dimer. Interesting though, the two monomers of the enzyme are not exactly the same. While one side consists only of parallel and antiparallel beta-sheets, the other monomer is made up of both <scene name='Nathan_Line_sandbox_3/Secondary_structure/1'>beta-sheets and alpha helixes</scene>. Though each monomer does not have to exact same sequence, each does contain replicate active sites and function. This is consistent with the following SCOP information:<br />
<br />
Class: Alpha and beta proteins (a/b)<br />
Fold: NAD(P)-binding Rossmann-fold domains<br />
Superfamily: NAD(P)-binding Rossmann-fold domains<br />
Family: Glyceraldehyde-3-phosphate dehydrogenase-like, N-terminus domain<br />
Protein: Glyceraldehyde-3-phosphate dehydrogenase<br />
Species: Human<br />
<br />
The specific reaction that GAPDH catalyzes is shown below:<br />
<br />
GAP + NAD+ + Pi +GAPDH <==> 1,3-bisphosphoglycerate + NADH <br />
<br />
<br />
The mechanism of the glycolysis reaction is fairly straight forward. After the aldehyde enters the <scene name='Nathan_Line_sandbox_3/Active_site/3'>active site</scene> (highlighted in green), the sulfhydryl group from <scene name='Nathan_Line_sandbox_3/Cystine/7'>Cystine 151</scene> attacks the nucleophilic carbon to form a thiohemiacetal. This intermediate undergoes oxidation due to a hydride transfer to a nearby NAD+ forming a thioester. From here, a phosphate group enters and attacks the same carbonyl while at the same time it is separated from the cystine by the protonated <scene name='Nathan_Line_sandbox_3/Histidine/5'>Histidine 178</scene> group. This produces the desired 1,3-bisphosphoglycerate. Though cystine-151 and histidine-178 are direct contributers to the catalytic process, other residues also influence the activity of this enzyme indirectly. <scene name='Nathan_Line_sandbox_3/Other/2'>Thr-210 and Arg-233</scene> are two such residues that contribute to the binding of the reactants rather than the catalytic mechanism. Regulation of GAPDH occurs through its coupling with the PGK reaction. This coupling is needed due to the slightly positive delta G of the glycolysis. The larger negative delta G of the PGK reaction results in the following overall net reaction with a delta G of -12.1 kJ/mol:<br />
<br />
GAP + Pi + NAD+ + ADP ==> 3PG + NADH + ATP<br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<br />
1) Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.<br />
<br />
2)Family: Glyceraldehyde-3-phosphate dehydrogenase-like, N-terminal domain. Retrived from: http://scop.mrc-lmb.cam.ac.uk/scop/data/scop.b.d.c.b.d.html<br />
<br />
3) <ref group="xtra">PMID:17676935</ref><references group="xtra"/><br />
<br />
4) <ref group="xtra">PMID:7340828</ref><references group="xtra"/><br />
<br />
5) <ref group="xtra">PMID:20164570</ref><references group="xtra"/></div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox_2&diff=1220467Samer Kawak sandbox 22011-03-30T00:18:47Z<p>Samer Kawak: New page: {{STRUCTURE_4ald | PDB=4ald | SCENE= }} ==Introduction and Structure== Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for th...</p>
<hr />
<div>{{STRUCTURE_4ald | PDB=4ald | SCENE= }}<br />
==Introduction and Structure==<br />
<br />
<br />
Fructose bisphosphate aldolase is an enzyme in glycolysis and gluconeogenesis. Glycolyis is responsible for the conversion of glucose into two three-carbon pyruvate molecules without the need for oxygen. The process generates two net ATP. The overall reaction is:<br />
<br />
Glucose + 2 NAD+ + 2 ADP + 2 Pi --> 2 pyruvate (3-carbon product) + 2 NADH + 2 ATP + 2 H20 + 4 H+<br />
<br />
Gluconeogenesis is responsible for maintaining the appropriate levels of blood glucose in animals by generating glucose from non-carbohydrate precursors. Gluconeogenesis can make glucose from lactate, pyruvate, citric acid cycle intermediates and from most amino acids (the exceptions being leucine and lysine). The common intermediate for all of the precursors on their way to becoming glucose must be oxaloacetate.<br />
<br />
The aldolase catalyzes the reversible cleavage of fructose-1,6-bisphosphate into dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (GAP). Different isozymes of aldolase can also catalyze the cleavage of fructose 1-phosphate to diydroxyacetone and glyceraldehyde (GA). Different isozymes exhibit preferences for either or both of the substrates, depending on the role of the aldolase (i.e. gluconeogenesis versus glycolysis).<ref name="book">Voet, D, Voet, J, & Pratt, C. (2008). Fundamentals of biochemistry, third edition. Hoboken, NJ: Wiley & Sons, Inc.</ref> <br />
<br />
While it can exist as a monomer, it normally exists as a <scene name='Austin_Drake_Sandbox/Tetramer/3'>homotetramer</scene>. The enzyme is an a/B protein with a TIM beta/alpha beta fold. The fold designation is based upon the nine alpha helices and eight parallel beta sheets in a closed barrel of each monomeric subunit. It is part of the aldolase superfamily and the class I aldolases.<ref>Protein: fructose-1,6-bisphosphate aldolase from human (homo sapiens), muscle isozyme. (2009). Retrieved from http://scop.mrc-lmb.cam.ac.uk</ref> <scene name='Austin_Drake_Sandbox/Different_colors/3'>α helices and β sheets</scene> can be seen in their specific regions mostly concentric to the active site, represented by the blue and red residues.<br />
<br />
Although some form of fructose bisphosphate aldolase is present in nearly all living things, certain isoforms carry a large degree of conservation. The enzyme from rabbit muscle has nearly the tertiary and primary structure as the enzyme in human muscle. As a result, implications from rabbit muscle aldolase also reveal a great deal about the human forms of the enzyme. <ref name="review">Gefflaut, T., B. Casimir, J. Perie, and M. Willson. "Class I Aldolases: Substrate Specificity, Mechanism, Inhibitors and Structural Aspects." Prog. Biophys. molec. Biol.. 63. (1995): 301-340.</ref><br />
<br />
==Binding and Catalysis==<br />
<br />
As an enzyme, the aldolase must not only encourage and favor the hydrolysis of fructose 1,6-bisphosphate, but also bind the substrate so as to hold it in the active site. The main-chain nitrogens of Ser271 and Gly272 hold the 1-phosphate group while the Lys41, Arg42 and Arg303 residues hold the 6-phosphate group. The five proposed binding residues are in close proximity to the catalytic Lys229, implicating them as participants in the binding process.<ref>PMID:10048322</ref> The <scene name='Austin_Drake_Sandbox/Tyr363/1'>C-terminus</scene>, which sits just outside of the barrel and catalytic site, of the enzyme also appears to contribute to the catalytic process of the aldolase. Mutations or suppression of the final tyrosine residue (Tyr363) causes a notable drop in the activity of the enzyme. Two cysteine residues have also been implicated in the catalytic process. Though they do not appear to be necessary for catalysis, modification of them does result in a decrease in catalytic activity. The two Cys residues are far from the active site, but do impact the movement of the C-terminus of the enzyme, which further implicates the terminus as participatory in the catalysis.<br />
<br />
The reaction is an aldol cleavage, or otherwise termed, retro aldo condensation. Catalysis occurs first when the nucleophilic ε-amine group of Lys229 attacks the carbonyl carbon of the substrate (FBP) in its open-ring state, pushing an electron pair to the oxygen of the carbonyl. The oxygen is protonated and leaves as water as a protonated <scene name='Austin_Drake_Sandbox/Schiff_base/2'>Schiff base</scene> is produced (an imine resulting from a ketone and amine) with the open-ring form of FBP, accompanied by electrostatic stabilization from <scene name='Austin_Drake_Sandbox/Catalytic_site_w_water/5'>Asp33</scene> Aldol cleavage between C3 and C4 produces GAP and an enamine precursor to DHAP.<ref name="book" /> The cleavage is facilitated by the positive charge from the Schiff base. The subsequent electron movement, which alleviates the positive charge, also breaks the C3-C4 bond.<ref name="review" /> Tautomerization, protonation and the hydrolysis of the Schiff base produce the final product of DHAP and regenerate the enzyme. The catalysis is driven by the more favorable stability of the protonated Schiff base compared to the enolate that would appear in basic catalysis pathways.<ref name="book" /> <br />
<br />
<br />
[[Image:Aldolase1.jpg]]<br />
<br />
==Kinetics==<br />
<br />
Isotopic labelling has revealed the rate-determining step for the reaction. Either the carbon-carbon bond cleavage or the release of glyceraldehyde-3-phosphate comprise the slow step of the catalysis reaction; however, studies do indicate that the GAP release is likely the slowest step.<ref name="review" /><br />
<br />
It has been shown that aldolase is inhibited allosterically by oxidized glutathione, which is an oxidizing species biologically present. The glutathione oxidizes a thiol 25 angstroms from the catalytic site, which subsequently causes a drop in catalytic activity. In addition, the enzyme shows no positive cooperativity, despite being an oligomer. In fact, kinetics data actually show that the enzyme exhibits negative cooperativity. Thus the catalysis is highly compartmentalized within each subunit and binding causes little distal change of the enzymes structure.<ref name="kinetics">Sygusch, J., and Beaudry, D. "Allosteric communication in mammalian muscle aldolase." Biochem. J.. 327. (1997): 717-720.</ref><br />
<br />
==Regulation==<br />
<br />
The regulation of fructose 1,6-bisphosphate aldolase is not well understood, but the understanding is every-increasing. As it is currently observed, aldolase C appears to be regulated mainly by the gene expression--the concentration of mRNA in the cytoplasm.<ref>Paolella, G, Buono, P, Mancini, F P, Izzo, P, and Salvatore, F. "Structure and expression of mouse aldolase genes." Eur. J. Biochem.. 156. (1986): 229-235.</ref> It is also known that adenosine 3',5'-cyclicmonophosphate (cAMP) affects the expression of the gene. cAMP concentration has been positively correlated with aldolase C expression. It is believed that cAMP acts upon a section of the promotor region, distal element D, causing the transcriptional promoter, NGFI-B, to bind. Once bound, the promoter activates the transcription of the gene coding for fructose bisphosphate aldolase.<ref>Buono, P, Cassano, S, Alfieri, A, Mancini, A, and Salvatore, F. "Human aldolase C gene expression is regulated by adenosine 30,50-cyclic monophosphate (cAMP) in PC12 cells." Gene. 291. (2002): 115-121.</ref> Given the inhibitory effects of an oxidant in the presence of aldolase, it is possible that this could be a mechanism of regulation of the enzyme. The deactivation that accompanies the oxidation of the surface thiol of Cys72 could be used intracellularly to slow the catalysis of the enzyme and regulate glycolysis.<ref name="kinetics" /><br />
<br />
==Additional Resources==<br />
For additional information, see: [[Carbohydrate Metabolism]]<br />
<br /><br />
<br />
==References==<br />
<references/></div>Samer Kawakhttps://proteopedia.org/index.php?title=User:Samer_Kawak&diff=1220465User:Samer Kawak2011-03-30T00:13:08Z<p>Samer Kawak: </p>
<hr />
<div>As a member of Proteopedia since the Fall 2009, I have contributed in a handful of areas including creating a new page and editing existing ones. I am currently a senior Chemistry major at Wabash College. First, when I was part of Chem 461,Chemistry of Cancer, last year, I produced a page on Bax Apoptosis. This year as a student in Chem 361, Biochemistry, I have worked on various editing projects.</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012638Samer Kawak sandbox2009-11-03T02:28:32Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. [4]<br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [4] <br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. [3]<br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [4]<br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]<br />
<br />
==Chipuk et al. (2004) Study==<br />
In the Chipuk et al. study from 2004, the authors determined that the Bax protein is directly activated by the tumor suppressor gene p53, which in turn mediates mitochondrial membrane permeabilization and apoptosis. Furthermore, the researchers discovered that cytosolic localization of endogenous wild-type or trans-activation deficient p53 was essential and adequate for apoptosis. Lastly, the transcription-independent activation of Bax by p53 happened with comparable kinetics and concentrations to that of activated Bid. [1] <br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein. [4]<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). [4]<br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. [4]<br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [4]<br />
<br />
==Reference==<br />
[1] Chipuk, J.E., Kuwana, T., Bouchier-Hayes, L., Droin, N.M., Newmeyer, D.D., Schuler, M., and Green, D.R. Direct activation of Bax by p53 mediates mitochondrial membrane permeabilization and apoptosis. Science 303: p. 1010-1014. <br />
<br />
[2] Gavathiotis, E., Suzuki M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D. BAX activation is initiated at a novel interaction site. Nature 455: p.1076-1081.<br />
<br />
[3] "OMIM - BCL2-ASSOCIATED X PROTEIN; BAX." NCBI HomePage. Web. 01 Nov. 2009. <http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=600040>.<br />
<br />
[4] Zhang, L., Jian, Y., Park, B.H., Kinzler, K.W., and Vogelstein, B. Role of Bax in the apoptotic response to anticancer agents. (2000). Science 290 (5493): 989-992.</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012446Samer Kawak sandbox2009-11-01T08:13:11Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. [2]<br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [2] <br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. [3]<br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [4]<br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein. [2]<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). [2]<br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. [2]<br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [2]<br />
<br />
==Reference==<br />
[1] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081.<br />
<br />
[2] Zhang, L., Jian, Y., Park, B.H., Kinzler, K.W., and Vogelstein, B. Role of Bax in the apoptotic response to anticancer agents. (2000). Science 290 (5493): 989-992. <br />
<br />
[3] "OMIM - BCL2-ASSOCIATED X PROTEIN; BAX." NCBI HomePage. Web. 01 Nov. 2009. <http://www.ncbi.nlm.nih.gov/entrez /dispomim.cgi?id=600040>.<br />
<br />
[4] Gavathiotis, E., Suzuki M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D. BAX activation is initiated at a novel interaction site. Nature 455: p.1076-1081.</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012445Samer Kawak sandbox2009-11-01T08:11:35Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. [2]<br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [2] <br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. [3]<br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [4]<br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein. [2]<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). [2]<br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. [2]<br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [2]<br />
<br />
==Reference==<br />
[1] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., <br />
Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081.<br />
<br />
[2] Zhang, L., Jian, Y., Park, B.H., Kinzler, K.W., and Vogelstein, B. Role of Bax in the apoptotic response to anticancer agents. <br />
(2000). Science 290 (5493): 989-992. <br />
<br />
[3] "OMIM - BCL2-ASSOCIATED X PROTEIN; BAX." NCBI HomePage. Web. 01 Nov. 2009. <http://www.ncbi.nlm.nih.gov/entrez <br />
/dispomim.cgi?id=600040>.<br />
<br />
[4] Gavathiotis, E., Suzuki M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., <br />
Walensky, L.D. BAX activation is initiated at a novel interaction site. Nature 455: p.1076-1081.</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012444Samer Kawak sandbox2009-11-01T07:41:08Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012443Samer Kawak sandbox2009-11-01T07:33:51Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
[4] <ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012442Samer Kawak sandbox2009-11-01T07:32:24Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
[[Image:300px-Signal transduction v1.png]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
[4] <ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012441Samer Kawak sandbox2009-11-01T07:30:47Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
[[Image:800px-Signal transduction v1.png|Apoptosis signal pathway|800 px|thumb]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
[4] <ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012440Samer Kawak sandbox2009-11-01T07:29:41Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
[[Image:800px-Signal transduction v1.png|Apoptosis signal pathway|200 px|thumb]]<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
[4] <ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012439Samer Kawak sandbox2009-11-01T07:27:12Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
<br />
[[Image:800px-Signal transduction v1.png|Apoptosis signal pathway|200 px|thumb]]<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
[4] <ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012438Samer Kawak sandbox2009-11-01T07:23:06Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
<br />
[[Image:800px-Signal transduction v1.png|Apoptosis signal pathway|200 px|thumb]]]]<br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
[4] <ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012437Samer Kawak sandbox2009-11-01T07:21:52Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
==Structure==<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
[[Image:800px-Signal transduction v1.png|Apoptosis signal pathway|700 px|thumb]]]] [4]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
[4] <ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012436Samer Kawak sandbox2009-11-01T07:21:03Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
==Structure==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
[[Image:800px-Signal transduction v1.png|Apoptosis signal pathway|600 px|thumb]]]] [4]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
[4] <ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012435Samer Kawak sandbox2009-11-01T07:20:28Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
==Structure==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
[[Image:800px-Signal transduction v1.png|Apoptosis signal pathway|500 px|thumb]]]] [4]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} <br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
[4] <ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012434Samer Kawak sandbox2009-11-01T07:18:05Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Overview of BAX and Apoptosis==<br />
<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. [1] <br />
<br />
==Structure==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} [2]<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131). [3]<br />
<br />
[[Image:800px-Signal transduction v1.png]] [4]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }} [2]<br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. [1]<br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
[2] structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
[4] <ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012433Samer Kawak sandbox2009-11-01T07:03:03Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==About Structure==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
Bax proteins contains 9 alpha helices while alpha-1 through alpha-8 are similar to that of Bcl-xL. The C-terminal alpha-9 helix occupies the hydrophobic pocket, which arbitrates the heterodimer formation and bioactivity of differing members of the Bcl-2 family. Researchers from the 2000 article by Suzuki et al. determined that the Bax structure indicates that the orientation of the alpha-9 helix offers concurrent control over its mitochondrial targeting and dimer formation. <br />
<br />
In another article by Gavathiotis et al. (2008), the authors discovered through NMR analysis that the BIM stabilized alpha-helix of Bcl-2 (SAHB) domain binds Bax at an interaction site different from the antiapoptotic proteins. The Bax binding site was also characterized by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131).<br />
<br />
==Overview of BAX and Apoptosis==<br />
<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. <br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. <br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
<ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012430Samer Kawak sandbox2009-11-01T05:47:07Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==About Structure==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
Apoptosis is stimulated by the insertion of BAX from the cytosol into mitochondrial membranes. Suzuki et al. (2000) determined the solution structure of BAX, including the putative transmembrane domain at the C terminus, in order to understand the regulation of its subcellular location. BAX consists of 9 alpha helices, and the assembly of helices alpha-1 through -8 resembles that of BCLXL. The C-terminal alpha-9 helix occupies the hydrophobic pocket proposed to mediate heterodimer formation and bioactivity of opposing members of the BCL2 family. The authors concluded that the BAX structure shows that the orientation of helix alpha-9 provides simultaneous control over its mitochondrial targeting and dimer formation. <br />
<br />
Gavathiotis et al. (2008) demonstrated by nuclear magnetic resonance (NMR) analysis that the BIM stabilized alpha-helix of BCL2 (SAHB) domain binds BAX at an interaction site that is distinct from the canonic binding groove characterized for antiapoptotic proteins. The specificity of the human BIM-SAHB-BAX interaction was highlighted by point mutagenesis that disrupts functional activity, confirming that BAX activation is initiated at this novel structural location. The BAX binding site is defined by lysine at <scene name='Samer_Kawak_sandbox/Lys_21/2'>position 21</scene> (K21), glutamine at <scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>positions 28 and 32</scene> (Q28, Q32), arginine at <scene name='Samer_Kawak_sandbox/Arg_134/1'>position 134</scene> (R134), and glutamic acid at <scene name='Samer_Kawak_sandbox/Glu_131/1'>position 131</scene> (E131).<br />
<br />
==Overview of BAX and Apoptosis==<br />
<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. <br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or <scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>codons 38 to 41</scene>) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. <br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
<ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012429Samer Kawak sandbox2009-11-01T05:40:07Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Abstract==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
BAX is a pro-apoptotic protein of the BCL-2 family that is stationed in the cytosol until activated by a diversity of stress stimuli to induce cell death. Anti-apoptotic proteins such as BCL-2 counteract BAX-mediated cell death. Although an interaction site that confers survival functionality has been defined for anti-apoptotic proteins, an activation site has not been identified for BAX, rendering its explicit trigger mechanism unknown. We previously developed stabilized alpha-helix of BCL-2 domains (SAHBs) that directly initiate BAX-mediated mitochondrial apoptosis. Here we demonstrate by NMR analysis that BIM SAHB binds BAX at an interaction site that is distinct from the canonical binding groove characterized for anti-apoptotic proteins. The specificity of the human BIM-SAHB-BAX interaction is highlighted by point mutagenesis that disrupts functional activity, confirming that BAX activation is initiated at this novel structural location. Thus, we have now defined a BAX interaction site for direct activation, establishing a new target for therapeutic modulation of apoptosis.<br />
<br />
<scene name='Samer_Kawak_sandbox/Lys_21/2'>LYS 21</scene><br />
<scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>GLN 28 32</scene><br />
<scene name='Samer_Kawak_sandbox/Arg_134/1'>TextToBeDisplayed</scene><br />
<scene name='Samer_Kawak_sandbox/Glu_131/1'>TextToBeDisplayed</scene><br />
<br />
==Overview of BAX and Apoptosis==<br />
<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. <br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or codons 38 to 41) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. <br />
<br />
<scene name='Samer_Kawak_sandbox/Codons_38_to_41/1'>Codons 38 to 41</scene><br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
<ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012427Samer Kawak sandbox2009-11-01T05:36:11Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Abstract==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
BAX is a pro-apoptotic protein of the BCL-2 family that is stationed in the cytosol until activated by a diversity of stress stimuli to induce cell death. Anti-apoptotic proteins such as BCL-2 counteract BAX-mediated cell death. Although an interaction site that confers survival functionality has been defined for anti-apoptotic proteins, an activation site has not been identified for BAX, rendering its explicit trigger mechanism unknown. We previously developed stabilized alpha-helix of BCL-2 domains (SAHBs) that directly initiate BAX-mediated mitochondrial apoptosis. Here we demonstrate by NMR analysis that BIM SAHB binds BAX at an interaction site that is distinct from the canonical binding groove characterized for anti-apoptotic proteins. The specificity of the human BIM-SAHB-BAX interaction is highlighted by point mutagenesis that disrupts functional activity, confirming that BAX activation is initiated at this novel structural location. Thus, we have now defined a BAX interaction site for direct activation, establishing a new target for therapeutic modulation of apoptosis.<br />
<br />
<scene name='Samer_Kawak_sandbox/Lys_21/2'>LYS 21</scene><br />
<scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>GLN 28 32</scene><br />
<scene name='Samer_Kawak_sandbox/Arg_134/1'>TextToBeDisplayed</scene><br />
<scene name='Samer_Kawak_sandbox/Glu_131/1'>TextToBeDisplayed</scene><br />
<br />
==Overview of BAX and Apoptosis==<br />
<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. <br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or codons 38 to 41) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. <br />
<br />
<scene name='Samer_Kawak_sandbox/Met_38/1'>MET 38</scene><br />
<br />
<scene name='Samer_Kawak_sandbox/Val_121/1'>VAL 121</scene><br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
<ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012425Samer Kawak sandbox2009-11-01T05:27:23Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Abstract==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
BAX is a pro-apoptotic protein of the BCL-2 family that is stationed in the cytosol until activated by a diversity of stress stimuli to induce cell death. Anti-apoptotic proteins such as BCL-2 counteract BAX-mediated cell death. Although an interaction site that confers survival functionality has been defined for anti-apoptotic proteins, an activation site has not been identified for BAX, rendering its explicit trigger mechanism unknown. We previously developed stabilized alpha-helix of BCL-2 domains (SAHBs) that directly initiate BAX-mediated mitochondrial apoptosis. Here we demonstrate by NMR analysis that BIM SAHB binds BAX at an interaction site that is distinct from the canonical binding groove characterized for anti-apoptotic proteins. The specificity of the human BIM-SAHB-BAX interaction is highlighted by point mutagenesis that disrupts functional activity, confirming that BAX activation is initiated at this novel structural location. Thus, we have now defined a BAX interaction site for direct activation, establishing a new target for therapeutic modulation of apoptosis.<br />
<br />
<scene name='Samer_Kawak_sandbox/Lys_21/2'>LYS 21</scene><br />
<scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>GLN 28 32</scene><br />
<scene name='Samer_Kawak_sandbox/Arg_134/1'>TextToBeDisplayed</scene><br />
<scene name='Samer_Kawak_sandbox/Glu_131/1'>TextToBeDisplayed</scene><br />
<br />
==Overview of BAX and Apoptosis==<br />
<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. <br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or codons 38 to 41) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. <br />
<br />
<scene name='Samer_Kawak_sandbox/Phe_114/1'>PHE 114</scene><br />
<br />
<scene name='Samer_Kawak_sandbox/Val_121/1'>VAL 121</scene><br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
<ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawakhttps://proteopedia.org/index.php?title=Samer_Kawak_sandbox&diff=1012424Samer Kawak sandbox2009-11-01T05:23:54Z<p>Samer Kawak: </p>
<hr />
<div>===BAX Protein===<br />
<br />
==Abstract==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
BAX is a pro-apoptotic protein of the BCL-2 family that is stationed in the cytosol until activated by a diversity of stress stimuli to induce cell death. Anti-apoptotic proteins such as BCL-2 counteract BAX-mediated cell death. Although an interaction site that confers survival functionality has been defined for anti-apoptotic proteins, an activation site has not been identified for BAX, rendering its explicit trigger mechanism unknown. We previously developed stabilized alpha-helix of BCL-2 domains (SAHBs) that directly initiate BAX-mediated mitochondrial apoptosis. Here we demonstrate by NMR analysis that BIM SAHB binds BAX at an interaction site that is distinct from the canonical binding groove characterized for anti-apoptotic proteins. The specificity of the human BIM-SAHB-BAX interaction is highlighted by point mutagenesis that disrupts functional activity, confirming that BAX activation is initiated at this novel structural location. Thus, we have now defined a BAX interaction site for direct activation, establishing a new target for therapeutic modulation of apoptosis.<br />
<br />
<scene name='Samer_Kawak_sandbox/Lys_21/2'>LYS 21</scene><br />
<scene name='Samer_Kawak_sandbox/Gln_28_and_32/1'>GLN 28 32</scene><br />
<scene name='Samer_Kawak_sandbox/Arg_134/1'>TextToBeDisplayed</scene><br />
<br />
==Overview of BAX and Apoptosis==<br />
<br />
By definition, apoptosis is programmed cell death, or suicide. To induce cell death among cancer cells is a critical component to cancer treatment. Specifically, the Bcl-2 protein family members have been found to have a prominent role in apoptosis, yet researchers have not been able to deduce more information on their mechanisms. Bcl-2 proteins occupy the mitochondrial outer membrane permeabilization, and are recognized either as pro-apoptotic (Bax, BAD, Bak, and Bok) or anti-apoptotic (Bcl-2 proper, Bcl-xL, and Bcl-w). Pro-apoptotic Bcl-2 proteins are death-promoting members while anti-apoptotic members are death-inhibiting structures. Currently, researchers have identified 25 genes in the Bcl-2 family. <br />
<br />
One particular Bcl-2 protein is the Bax protein, one of the death-promoting members. To date, researchers have found great trouble deciphering the nature of the Bax protein in the cell. Past studies have overexpressed the Bax protein in in vitro and in vivo studies to discover more information on its function. However, some of the criticism from such methods is that these conditions are not indicative of the normal activity of Bax. Either way, there have been mixed results through these types of studies. <br />
<br />
[[Image:800px-Signal transduction v1.png]]<br />
<br />
==Zhang et al. (2000) Study==<br />
{{STRUCTURE_1f16 | PDB=1f16 | SCENE= }}<br />
In the 2000 study by Lin Zhang, Jian Yu, Ben Ho Park, Kenneth W. Kinzler, and Bert Vogelstein titled “Role of Bax in the Apoptotic Response to Anticancer Agents,” the experimenters evaluated the role of the Bax protein in drug-induced apoptosis in human colorectal cancer cells. The review described several experiments that all attempted to elucidate the mystery behind the Bax protein.<br />
<br />
For the first experiment, the researchers used the fact that chemotherapeutic agents usually target epithelial cells to clarify the role of Bax in drug-induced apoptosis in epithelial cells through the creation and analysis of isogenic derivatives that vary only in the presence or absence of the Bax gene. The results from this experiment showed that 2% of HCT116 (or epithelial parental cells) have two intact Bax alleles (+/+), 94% had one intact allele (+/-), and 4% had two mutant alleles (-/-). <br />
<br />
In another experiment, the experiments presented that hypothesis that if Bax deficiency greatly affects the sensitivity to non-steroidal anti-inflammatory drugs (NSAIDs), which were previously test in a different experiment, then the parental cell populations treated with NSAIDs result in a mutated Bax cell population. After recovering grown Bax cells from an in vitro study with the NSAID indomethacin. The results showed that 70% of the Bax cells had insertions or deletions in the G8 tracts of both Bax alleles while 4% had mutations in the parental population. <br />
<br />
Ultimately, in regards to the clinical implications from this study, the results suggest that colorectal tumors may easily develop resistance to NSAIDs through an inherent instability in the mononucleotide tract (at nucleotides 114 to 121 or codons 38 to 41) in BAX, but more importantly, a combination of chemopreventive drugs must be considered as a leading cancer treatment. <br />
<br />
<scene name='Samer_Kawak_sandbox/Phe_114/1'>PHE 114</scene><br />
<br />
<scene name='Samer_Kawak_sandbox/Val_121/1'>VAL 121</scene><br />
<br />
==Reference==<br />
Gavathiotis E, Suzuki M, Davis ML, Pitter K, Bird GH, Katz SG, Tu HC, Kim H, Cheng EH, Tjandra N, Walensky LDBAX activation is initiated at a novel interaction siteNature v455, p.1076-1081<br />
<br />
structure obtained by:<br />
BAX activation is initiated at a novel interaction site.<br />
Gavathiotis, E., Suzuki, M., Davis, M.L., Pitter, K., Bird, G.H., Katz, S.G., Tu, H.C., Kim, H., Cheng, E.H., Tjandra, N., Walensky, L.D.(2008) Nature 455: 1076-1081<br />
<br />
<ref group="xtra">PMID:8875929</ref><references group="xtra"/><br />
[[Category: Xenopus laevis]]<br />
[[Category: P53 Tumor Suppressor]]<br />
[[Category: Kussie, P H.]]<br />
[[Category: Pavletich, N P.]]<br />
[[Category: Activator]]<br />
[[Category: Anti-oncogene]]<br />
[[Category: Dna-binding]]<br />
[[Category: Nuclear protein]]<br />
[[Category: Phosphorylation]]<br />
[[Category: Transcription regulation]]</div>Samer Kawak