Sandbox Reserved 1475: Difference between revisions

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== Function ==

The reaction of this enzyme is [(retinal) + (NAD+) + (H2O) ↔ (retinoic acid) + (NADH) + (H+)].

The main function of this enzyme is to produce Retinoic acid. RalDH2 requires (NAD+) as a cofactor.<ref name="Lamb AL, Newcomber ME" /> In the oxidoreductase reaction, NAD+ acts as an electron acceptor. Once the NAD+ is bound, hydrogen bonds form with non-polar residues and one basic Lysine residue. Chloride ions participate in hydrophobic interactions with Arginine residues.<ref name="Lamb AL, Newcomber ME" /> ~~Theres~~ interactions cause a structural change to occur in the RalDH2 enzyme which causes it to form a more favorable folded confirmation. In the enzyme a large binding cavity is formed.

The main function of this enzyme is to produce Retinoic acid. RalDH2 requires (NAD+) as a cofactor.<ref name="Lamb AL, Newcomber ME" /> In the oxidoreductase reaction, NAD+ acts as an electron acceptor. Once the NAD+ is bound, hydrogen bonds form with non-polar residues and one basic Lysine residue. Chloride ions participate in hydrophobic interactions with Arginine residues.<ref name="Lamb AL, Newcomber ME" /> These interactions cause a structural change to occur in the RalDH2 enzyme which causes it to form a more favorable folded confirmation. In the enzyme a large binding cavity is formed. Structural changes occur to stabilize the tertiary structure of RalDH2

~~tructural~~ changes occur to stabilize the tertiary structure of RalDH2

== Structural highlights ==

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===Cofactor NAD and Cl ions===

The crystal structure was cocrystallized with <scene name='80/800654/Nad/1'>NAD</scene>, and was determined at a 2.7 Angstrom resolution ([[Figure 2]] Chain D present with NAD represented in pink).<ref name="Lamb AL, Newcomber ME" /> NAD+ acts as a cofactor and is the electron acceptor in RalDH2 oxidoructase reaction as seen in the reaction presented above. RalDH2 ~~has to~~ be folded in a proper manner for its enzymatic function to occur. The folding of the enzyme is partially due to the interactions of NAD+ and Chloride ions. When NAD+ is present hydrogen bonds with Glu and Ser form, van der Waals interactions with non-polar residues and one polar residue (Lys) forms. The interaction with Lys-192 provides the transition state stability, making for a favorable confirmation.<ref name="Lamb AL, Newcomber ME" /> This conformational change is important since in the absence of NAD+, no crystal structures grew in any of the screened conditions.<ref name="Lamb AL, Newcomber ME" /> The Chloride ions participate in hydrophobic interactions with Arg which also help maintain the folded structure.<ref name="Lamb AL, Newcomber ME" /> With the cofactor NAD+ present the catalytic domain of RalDH2 is highly mobile and needs the selective substrate present to immobilize the catalytic domain. The binding of hydrophobic substrate and the NAD+ cofactor are needed to stabilize the catalytic domain.<ref name="Lamb AL, Newcomber ME" /> Short chain aldehydes do not have a large enough hydrophobic surface to bury inside the channel ~~and~~ therefor cannot work as suitable substrates. The long tails of the long chained aldehydes are needed to interact with the catalytic Cys-302 through hydrogen bonds. Short chained aldehydes can have hydrogen bond interactions with Cys-302 however are not large enough to fully bury the whole access channel, which is needed for the catalytic domain to be immobilized.

The crystal structure was cocrystallized with <scene name='80/800654/Nad/1'>NAD</scene>, and was determined at a 2.7 Angstrom resolution ([[Figure 2]] Chain D present with NAD represented in pink).<ref name="Lamb AL, Newcomber ME" /> NAD+ acts as a cofactor and is the electron acceptor in RalDH2 oxidoructase reaction as seen in the reaction presented above. RalDH2 must be folded in a proper manner for its enzymatic function to occur. The folding of the enzyme is partially due to the interactions of NAD+ and Chloride ions. When NAD+ is present hydrogen bonds with Glu and Ser form, van der Waals interactions with non-polar residues and one polar residue (Lys) forms. The interaction with Lys-192 provides the transition state stability, making for a favorable confirmation.<ref name="Lamb AL, Newcomber ME" /> This conformational change is important since in the absence of NAD+, no crystal structures grew in any of the screened conditions.<ref name="Lamb AL, Newcomber ME" /> The Chloride ions participate in hydrophobic interactions with Arg which also help maintain the folded structure.<ref name="Lamb AL, Newcomber ME" /> With the cofactor NAD+ present the catalytic domain of RalDH2 is highly mobile and needs the selective substrate present to immobilize the catalytic domain. The binding of hydrophobic substrate and the NAD+ cofactor are needed to stabilize the catalytic domain.<ref name="Lamb AL, Newcomber ME" /> Short chain aldehydes do not have a large enough hydrophobic surface to bury inside the channel, therefor cannot work as suitable substrates. The long tails of the long chained aldehydes are needed to interact with the catalytic Cys-302 through hydrogen bonds. Short chained aldehydes can have hydrogen bond interactions with Cys-302 however are not large enough to fully bury the whole access channel, which is needed for the catalytic domain to be immobilized.

===Cys-302===

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===Energetics===

[[Image:Relative efficiencies of RalDH2 for substrates.png|thumb|upright=1.5| [[Figure 5]] Relative efficiencies of RalDH2 for aldehyde substrates. Image reference.<ref name="Km value chart" />]]

With the cofactor NAD+ present the catalytic domain of RalDH2 is highly mobile and needs the selective substrate present to immobilize the catalytic domain. The binding of hydrophobic substrate and the NAD+ cofactor are needed to stabilize the catalytic domain.<ref name="Lamb AL, Newcomber ME" /> Short chain aldehydes do not have a large enough hydrophobic surface to bury inside the channel and therefor cannot work as suitable substrates. The long tails of the long chained aldehydes are needed to interact with the catalytic Cys-302 through hydrogen bonds. Short chained aldehydes can have hydrogen bond interactions with Cys-302 however are not large enough to fully bury the whole access channel, which is needed for the catalytic domain to be immobilized.

In [[Figure 5]] it is visible to see that acetaldehyde and benzaldehyde both have really high Km values, 645uM and 305uM respectfully, and relatively low Vmax values, 139nmol/min/mg and 200nmol/min/mg respectfully.<ref name="Km value chart">Wang, Xianshu. Penzes, Peter., Napoli, Joseph L., Cloning of a cDNA Encoding an Aldehyde Dehydrogenase and Its Expression in ''Escherichia coli'' RECOGNITION OF RETINAL AS SUBSTRATE. J. Biol. Chem. (1996) 271:16288-16293. doi:10.1074/jbc.271.27.16288 </ref> Octantal and decanal both have really low Km values, 5uM and 3uM respectfully, and relatively high Vmax values, 152nmol/min/mg and 214nmol/min/mg respectfully.<ref name="Km value chart" /> Acetaldehyde and benzaldehyde are both short chained aldehydes compared to octantal and decanal aldehydes. It is easier to compare the ratio of Vmax/Km. An energetically favorable substrate would display a ratio of Vmax/Km that has a large magnitude. As seen in [[Figure 5]], both the long chained octantal and decanal aldehydes had large Vmax/Km values, 152 AND 214 respectfully.<ref name="Km value chart" /> The substrate that RalDH2 uses to actually convert Vitamin A (Retinol) to retinoic acid is retinal in its "all-trans" form. As seen in [[Figure 5]] the Km value for the this substrate is the smallest out all that were tested, and the Vmax values was comparatively high. The Vmax/Km was also pretty large at a value of 49± 6.<ref name="Km value chart" />

== Relevance - Disease ==

[[Image:Embryo of RalDH2.png|thumb|upright=1.5| [[Figure 6]] "Morphological abnormalities of RalDH2 knock out embryos".<ref name="Embryonic retinoic acid synthesis" />]]

Multiple complications can occur if there is a deficiency of RalDH2 in mammals. If there were to be a RalDH2 deficiency during the embryonic development, possible congenital malformations can occur. The complications include defects such as lack of axial rotation, incomplete neural tube closure, and lack of heart looping and chamber morphogenesis.<ref name="Embryonic retinoic acid synthesis">PMID:10192400</ref> With the study of mouse with their RalDH2 emzyne knocked out, hearts consisted of single, medial, dilated cavities.<ref name="Embryonic retinoic acid synthesis" /> The mice displayed their frontonasal region to be truncated, and their otocysts to be reduced.<ref name="Embryonic retinoic acid synthesis" /> In [[Figure 6]], the comparison of wild-type embryos compared to ones that are RalDH2 negative. It is visible to see that there is lack of embryonic turning, associated with a truncation of the posterior region.<ref name="Embryonic retinoic acid synthesis" /> The WT 8.5 doc embryo is the wild-type embryo before turning has occurred. Section b displays a wild-type embryo compared to defective embryos, section c-d. The e and f show the embryos of a wild-type embryo compared to one that was low in the RalDH2 enzyme, respectfully.

~~== Disease ==~~

~~== Relevance ==~~

~~</StructureSection>~~

== References ==

Sandbox Reserved 1475: Difference between revisions

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@@ Line 13: / Line 13: @@
 == Function ==
 The reaction of this enzyme is [(retinal) + (NAD+) + (H2O) ↔ (retinoic acid) + (NADH) + (H+)].
-The main function of this enzyme is to produce Retinoic acid. RalDH2 requires (NAD+) as a cofactor.<ref name="Lamb AL, Newcomber ME" /> In the oxidoreductase reaction, NAD+ acts as an electron acceptor. Once the NAD+ is bound, hydrogen bonds form with non-polar residues and one basic Lysine residue. Chloride ions participate in hydrophobic interactions with Arginine residues.<ref name="Lamb AL, Newcomber ME" /> Theres interactions cause a structural change to occur in the RalDH2 enzyme which causes it to form a more favorable folded confirmation. In the enzyme a large binding cavity is formed.
+The main function of this enzyme is to produce Retinoic acid. RalDH2 requires (NAD+) as a cofactor.<ref name="Lamb AL, Newcomber ME" /> In the oxidoreductase reaction, NAD+ acts as an electron acceptor. Once the NAD+ is bound, hydrogen bonds form with non-polar residues and one basic Lysine residue. Chloride ions participate in hydrophobic interactions with Arginine residues.<ref name="Lamb AL, Newcomber ME" /> These interactions cause a structural change to occur in the RalDH2 enzyme which causes it to form a more favorable folded confirmation. In the enzyme a large binding cavity is formed. Structural changes occur to stabilize the tertiary structure of RalDH2
-tructural changes occur to stabilize the tertiary structure of RalDH2
 == Structural highlights ==
@@ Line 29: / Line 28: @@
 ===Cofactor NAD and Cl ions===
-The crystal structure was cocrystallized with <scene name='80/800654/Nad/1'>NAD</scene>, and was determined at a 2.7 Angstrom resolution ([[Figure 2]] Chain D present with NAD represented in pink).<ref name="Lamb AL, Newcomber ME" /> NAD+ acts as a cofactor and is the electron acceptor in RalDH2 oxidoructase reaction as seen in the reaction presented above. RalDH2 has to be folded in a proper manner for its enzymatic function to occur. The folding of the enzyme is partially due to the interactions of NAD+ and Chloride ions. When NAD+ is present hydrogen bonds with Glu and Ser form, van der Waals interactions with non-polar residues and one polar residue (Lys) forms. The interaction with Lys-192 provides the transition state stability, making for a favorable confirmation.<ref name="Lamb AL, Newcomber ME" /> This conformational change is important since in the absence of NAD+, no crystal structures grew in any of the screened conditions.<ref name="Lamb AL, Newcomber ME" />  The Chloride ions participate in hydrophobic interactions with Arg which also help maintain the folded structure.<ref name="Lamb AL, Newcomber ME" /> With the cofactor NAD+ present the catalytic domain of RalDH2 is highly mobile and needs the selective substrate present to immobilize the catalytic domain. The binding of hydrophobic substrate and the NAD+ cofactor are needed to stabilize the catalytic domain.<ref name="Lamb AL, Newcomber ME" /> Short chain aldehydes do not have a large enough hydrophobic surface to bury inside the channel and therefor cannot work as suitable substrates. The long tails of the long chained aldehydes are needed to interact with the catalytic Cys-302 through hydrogen bonds. Short chained aldehydes can have hydrogen bond interactions with Cys-302 however are not large enough to fully bury the whole access channel, which is needed for the catalytic domain to be immobilized.
+The crystal structure was cocrystallized with <scene name='80/800654/Nad/1'>NAD</scene>, and was determined at a 2.7 Angstrom resolution ([[Figure 2]] Chain D present with NAD represented in pink).<ref name="Lamb AL, Newcomber ME" /> NAD+ acts as a cofactor and is the electron acceptor in RalDH2 oxidoructase reaction as seen in the reaction presented above. RalDH2 must be folded in a proper manner for its enzymatic function to occur. The folding of the enzyme is partially due to the interactions of NAD+ and Chloride ions. When NAD+ is present hydrogen bonds with Glu and Ser form, van der Waals interactions with non-polar residues and one polar residue (Lys) forms. The interaction with Lys-192 provides the transition state stability, making for a favorable confirmation.<ref name="Lamb AL, Newcomber ME" /> This conformational change is important since in the absence of NAD+, no crystal structures grew in any of the screened conditions.<ref name="Lamb AL, Newcomber ME" />  The Chloride ions participate in hydrophobic interactions with Arg which also help maintain the folded structure.<ref name="Lamb AL, Newcomber ME" /> With the cofactor NAD+ present the catalytic domain of RalDH2 is highly mobile and needs the selective substrate present to immobilize the catalytic domain. The binding of hydrophobic substrate and the NAD+ cofactor are needed to stabilize the catalytic domain.<ref name="Lamb AL, Newcomber ME" /> Short chain aldehydes do not have a large enough hydrophobic surface to bury inside the channel, therefor cannot work as suitable substrates. The long tails of the long chained aldehydes are needed to interact with the catalytic Cys-302 through hydrogen bonds. Short chained aldehydes can have hydrogen bond interactions with Cys-302 however are not large enough to fully bury the whole access channel, which is needed for the catalytic domain to be immobilized.
 ===Cys-302===
@@ Line 36: / Line 35: @@
 ===Energetics===
 [[Image:Relative efficiencies of RalDH2 for substrates.png|thumb|upright=1.5| [[Figure 5]] Relative efficiencies of RalDH2 for aldehyde substrates. Image reference.<ref name="Km value chart" />]]
-With the cofactor NAD+ present the catalytic domain of RalDH2 is highly mobile and needs the selective substrate present to immobilize the catalytic domain. The binding of hydrophobic substrate and the NAD+ cofactor are needed to stabilize the catalytic domain.<ref name="Lamb AL, Newcomber ME" /> Short chain aldehydes do not have a large enough hydrophobic surface to bury inside the channel and therefor cannot work as suitable substrates. The long tails of the long chained aldehydes are needed to interact with the catalytic Cys-302 through hydrogen bonds. Short chained aldehydes can have hydrogen bond interactions with Cys-302 however are not large enough to fully bury the whole access channel, which is needed for the catalytic domain to be immobilized.
 In [[Figure 5]] it is visible to see that acetaldehyde and benzaldehyde both have really high Km values, 645uM and 305uM respectfully, and relatively low Vmax values, 139nmol/min/mg and 200nmol/min/mg respectfully.<ref name="Km value chart">Wang, Xianshu. Penzes, Peter., Napoli, Joseph L., Cloning of a cDNA Encoding an Aldehyde Dehydrogenase and Its Expression in ''Escherichia coli'' RECOGNITION OF RETINAL AS SUBSTRATE. J. Biol. Chem. (1996) 271:16288-16293. doi:10.1074/jbc.271.27.16288 </ref> Octantal and decanal both have really low Km values, 5uM and 3uM respectfully, and relatively high Vmax values, 152nmol/min/mg and 214nmol/min/mg respectfully.<ref name="Km value chart" /> Acetaldehyde and benzaldehyde are both short chained aldehydes compared to octantal and decanal aldehydes. It is easier to compare the ratio of Vmax/Km. An energetically favorable substrate would display a ratio of Vmax/Km that has a large magnitude. As seen in [[Figure 5]], both the long chained octantal and decanal aldehydes had large Vmax/Km values, 152 AND 214 respectfully.<ref name="Km value chart" /> The substrate that RalDH2 uses to actually convert Vitamin A (Retinol) to retinoic acid is retinal in its "all-trans" form. As seen in [[Figure 5]] the Km value for the this substrate is the smallest out all that were tested, and the Vmax values was comparatively high. The Vmax/Km was also pretty large at a value of 49± 6.<ref name="Km value chart" />
+== Relevance - Disease ==
+[[Image:Embryo of RalDH2.png|thumb|upright=1.5| [[Figure 6]] "Morphological abnormalities of RalDH2 knock out embryos".<ref name="Embryonic retinoic acid synthesis" />]]
+Multiple complications can occur if there is a deficiency of RalDH2 in mammals. If there were to be a RalDH2 deficiency during the embryonic development, possible congenital malformations can occur. The complications include defects such as lack of axial rotation, incomplete neural tube closure, and lack of heart looping and chamber morphogenesis.<ref name="Embryonic retinoic acid synthesis">PMID:10192400</ref> With the study of mouse with their RalDH2 emzyne knocked out, hearts consisted of single, medial, dilated cavities.<ref name="Embryonic retinoic acid synthesis" /> The mice displayed their frontonasal region to be truncated, and their otocysts to be reduced.<ref name="Embryonic retinoic acid synthesis" /> In [[Figure 6]], the comparison of wild-type embryos compared to ones that are RalDH2 negative. It is visible to see that there is lack of embryonic turning, associated with a truncation of the posterior region.<ref name="Embryonic retinoic acid synthesis" /> The WT 8.5 doc embryo is the wild-type embryo before turning has occurred. Section b displays a wild-type embryo compared to defective embryos, section c-d. The e and f show the embryos of a wild-type embryo compared to one that was  low in the RalDH2 enzyme, respectfully.
-== Disease ==
-== Relevance ==
-</StructureSection>
 == References ==
 <references/>