Fumarase 2: Difference between revisions

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===Overview===

Fumarase, also known as fumarate hydratase, functions as an enzyme in the metabolic pathway known as the ~~Kreb’s~~ cycle, or citric acid cycle. As the seventh step in the pathway, fumarase catalyzes the reversible reaction ~~converting~~ fumarate to S-malate. ~~It metabolizes Fumarate~~ in the ~~cytosol, which becomes a byproduct of the urea cycle and amino acid catabolism. It catalyzes the addition of water~~ to ~~make S-Malate~~; therefore, the mechanism of ~~fumarase in the~~ reaction involves hydration of fumarate in order to form malate.

Fumarase, also known as fumarate hydratase, is a protein that functions as an enzyme in the metabolic pathway known as the Krebs cycle, or citric acid cycle. Fumarase includes only α-helices for its secondary structure, and it has four identical subunits in its biological state. In the seventh step of the reaction pathway, fumarase catalyzes the reversible reaction that converts fumarate to malate. A water molecule is used in the reaction to form malate; therefore, the mechanism of reaction involves hydration of fumarate in order to form malate. The kinetics demonstrated by fumarase catalyzing the reaction in both directions indicate that the forward pathway is energetically favored. However, fumarase activity can be regulated by allosteric effects or inhibition, and the allosteric effects mainly result from conformational changes when substrate binds to one of its sites.

===Stucture ~~and Classification~~===

===Stucture===

Fumarase is classified as an all alpha protein which belongs to the L-aspartase/fumarase family, and the enzyme specifically consists of four identical subunits which form a tetramer (see image on right). From the four subunits, fumarase has three domains which comprise two binding sites: the active site and B site. Although the active site has a mostly solid structure and shifts very little when it binds, the B site shifts substantially more upon binding, and this shift helps regulate affinity for molecule binding at the active site <ref name="Weaver, et al."> Weaver,T. Structure of free fumarase C from ''Escherichia coli''. ''Acta Crystallographica'' (2005), '''D61''', 1395-1401. ['''http://dx.doi.org/10.1107/S0907444905024194''' doi:10.1107/S0907444905024194]</ref>.

Fumarase is classified as an all alpha protein which belongs to the L-aspartase/fumarase family, and the enzyme specifically consists of four identical subunits which form a tetramer that appears symmetric(see image on right). From the four subunits, fumarase has three domains which comprise two binding sites: the active site and B site. Although the active site has a mostly solid structure and shifts very little when it binds to the substrate, the B site shifts substantially more upon binding, and this shift helps regulate affinity for molecule binding at the active site <ref name="Weaver, et al."> Weaver,T. Structure of free fumarase C from ''Escherichia coli''. ''Acta Crystallographica'' (2005), '''D61''', 1395-1401. ['''http://dx.doi.org/10.1107/S0907444905024194''' doi:10.1107/S0907444905024194]</ref>. This has several implications for regulation of fumarase's activity and affinity to bind at the active site, but water molecules also play an important role in its function as an enzyme.

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===~~Enzyme~~ Kinetics===

===Kinetics===

Fumarase catalyzes the reversible reaction between S-malate and fumarate in the citric acid cycle for cellular metabolism. When it catalyzes the addition of water to S-malate in order to form fumarate, the Km and Vmax values are 0.30 mM and 129 s^-1, respectively. The reverse reaction (dehydration of fumarate to form S-malate) has a Km of 0.10 mM and Vmax of 60 s^-1 <ref name="Rose & Weaver"> Rose, I. & Weaver, T. The role of the allosteric B site in the fumarase reaction. ''Proc. Natl. Acad. Sci. USA'' (2004), '''101'''(10), 3393-3397. ['''http://www.pnas.org/cgi/doi/10.1073/pnas.0307524101''' doi:10.1073/pnas.0307524101]</ref>. Thus, the forward pathway (fumarate to S-malate) is favored because of its higher Km and Vmax values. Fumarase kinetics normally follows Michaelis-Menten kinetic plots at low concentrations of substrate, but high substrate concentrations influence the enzyme’s activity due to allosteric effects. Enzyme kinetics studies with fumarase mutants differ from wild type fumarase kinetics when the mutations alter amino acid residues involved in binding at the active site and B site. Also, the Michaelis-Menten kinetics plots for fumarase mutants exhibit a sigmoidal curve which suggests the presence of cooperativity in the enzymes activity.

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@@ Line 4: / Line 4: @@
 ===Overview===
-Fumarase, also known as fumarate hydratase, functions as an enzyme in the metabolic pathway known as the Kreb’s cycle, or citric acid cycle.  As the seventh step in the pathway, fumarase catalyzes the reversible reaction converting fumarate to S-malate.  It metabolizes Fumarate in the cytosol, which becomes a byproduct of the urea cycle and amino acid catabolism. It catalyzes the addition of water to make S-Malate; therefore, the mechanism of fumarase in the reaction involves hydration of fumarate in order to form malate.
+Fumarase, also known as fumarate hydratase, is a protein that functions as an enzyme in the metabolic pathway known as the Krebs cycle, or citric acid cycle.  Fumarase includes only α-helices for its secondary structure, and it has four identical subunits in its biological state.  In the seventh step of the reaction pathway, fumarase catalyzes the reversible reaction that converts fumarate to malate.  A water molecule is used in the reaction to form malate; therefore, the mechanism of reaction involves hydration of fumarate in order to form malate.  The kinetics demonstrated by fumarase catalyzing the reaction in both directions indicate that the forward pathway is energetically favored.  However, fumarase activity can be regulated by allosteric effects or inhibition, and the allosteric effects mainly result from conformational changes when substrate binds to one of its sites.
-===Stucture and Classification===
+===Stucture===
-Fumarase is classified as an all alpha protein which belongs to the L-aspartase/fumarase family, and the enzyme specifically consists of four identical subunits which form a tetramer (see image on right).  From the four subunits, fumarase has three domains which comprise two binding sites: the active site and B site.  Although the active site has a mostly solid structure and shifts very little when it binds, the B site shifts substantially more upon binding, and this shift helps regulate affinity for molecule binding at the active site <ref name="Weaver, et al."> Weaver,T.  Structure of free fumarase C from ''Escherichia coli''. ''Acta Crystallographica'' (2005), '''D61''', 1395-1401. ['''http://dx.doi.org/10.1107/S0907444905024194''' doi:10.1107/S0907444905024194]</ref>.
+Fumarase is classified as an all alpha protein which belongs to the L-aspartase/fumarase family, and the enzyme specifically consists of four identical subunits which form a tetramer that appears symmetric(see image on right).  From the four subunits, fumarase has three domains which comprise two binding sites: the active site and B site.  Although the active site has a mostly solid structure and shifts very little when it binds to the substrate, the B site shifts substantially more upon binding, and this shift helps regulate affinity for molecule binding at the active site <ref name="Weaver, et al."> Weaver,T.  Structure of free fumarase C from ''Escherichia coli''. ''Acta Crystallographica'' (2005), '''D61''', 1395-1401. ['''http://dx.doi.org/10.1107/S0907444905024194''' doi:10.1107/S0907444905024194]</ref>.  This has several implications for regulation of fumarase's activity and affinity to bind at the active site, but water molecules also play an important role in its function as an enzyme.
@@ Line 16: / Line 16: @@
-===Enzyme Kinetics===
+===Kinetics===
 Fumarase catalyzes the reversible reaction between S-malate and fumarate in the citric acid cycle for cellular metabolism.  When it catalyzes the addition of water to S-malate in order to form fumarate, the Km and Vmax values are 0.30 mM and 129 s^-1, respectively.  The reverse reaction (dehydration of fumarate to form S-malate) has a Km of 0.10 mM and Vmax of 60 s^-1 <ref name="Rose & Weaver"> Rose, I. & Weaver, T.  The role of the allosteric B site in the fumarase reaction.  ''Proc. Natl. Acad. Sci. USA'' (2004), '''101'''(10), 3393-3397.  ['''http://www.pnas.org/cgi/doi/10.1073/pnas.0307524101''' doi:10.1073/pnas.0307524101]</ref>.  Thus, the forward pathway (fumarate to S-malate) is favored because of its higher Km and Vmax values.  Fumarase kinetics normally follows Michaelis-Menten kinetic plots at low concentrations of substrate, but high substrate concentrations influence the enzyme’s activity due to allosteric effects.  Enzyme kinetics studies with fumarase mutants differ from wild type fumarase kinetics when the mutations alter amino acid residues involved in binding at the active site and B site.  Also, the Michaelis-Menten kinetics plots for fumarase mutants exhibit a sigmoidal curve which suggests the presence of cooperativity in the enzymes activity.