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[[Image:2cts polyview.jpg|450px|left|thumb| Citrate synthase 'closed form', [[2cts]]]] ==The Structure and Mechanism of Citrate Synthase== <Structure load='1cts' size='480' frame='true' align='right' scene='Daniel_Eddelman_Sandbox_2/Cts_open_monomer/2' caption='Citrate Synthase' /> Citrate synthase is an enzyme active in the mitochondria, where it is responsible for catalyzing the first reaction of the citric acid cycle (Krebs Cycle): the condensation of acetyl-CoA and oxaloacetate to form citrate. Although it is a mitochondrial enzyme (in fact, it is often used as a quantitative enzyme marker for intact mitochondria), it is encoded by nuclear DNA, not mitochondrial <ref>[http://en.wikipedia.org/wiki/Citrate_synthase "Citrate Synthase -." Wikipedia, the Free Encyclopedia. Web. 22 Mar. 2010].</ref>. The standard free energy change (ΔG°’) for the citrate synthase reaction is -31.5kJ/mol <ref name="voet">Voet, Donald, Judith G. Voet, and Charlotte W. Pratt. Fundamentals of Biochemistry: Life at the Molecular Level. Hoboken, NJ: Wiley, 2008.</ref>. This negative free energy value means that citrate synthase is likely to function far from equilibrium under physiological conditions, and is thus a rate-determining enzyme in the citric acid cycle. '''Structure:''' Biologically, citrate synthase exists as a <scene name='Daniel_Eddelman_Sandbox_2/Cts_open_monomer/2'>homodimer</scene> of a single amino acid chain <scene name='Daniel_Eddelman_Sandbox_2/Cts_open_monomer/1'>monomer</scene>. Each identical subunit consists of a large and a small domain, and is comprised almost entirely of α helices (making it an all α protein). In its free enzyme state, citrate synthase exists in <scene name='Daniel_Eddelman_Sandbox_2/Cts_open_monomer/2'>an “open” form of the homodimer</scene>, with its two domains forming a cleft containing the substrate (oxaloacetate) binding site (PDB: [[1cts]]) <ref>PMID:7120407</ref>. When oxaloacetate binds, the smaller domain undergoes an 18° rotation, sealing the oxaloacetate binding site and resulting in the <scene name='Daniel_Eddelman_Sandbox_2/Closed_homodimer/1'>closed conformation of the homodimer</scene> (PDB: [[2cts]])<ref>PMID:7308213</ref>. The dramatic conformational change is best illustrated via <scene name='User:Wayne_Decatur/1cts_to_2cts_(citrate_synthase)_morph_methods/1ctsto2ctsmorph/5'>a morph between the "open" and "closed" states</scene>. <nowiki>[</nowiki>Be patient; it may take longer than half a minute for the morph to load. <nowiki>]</nowiki> The conformational change not only prevents solvent from reaching the bound substrate, but also generates the acetyl-CoA binding site. This presence of “open” and “closed” forms results in citrate synthase having Ordered Sequential kinetic behavior <ref name="voet" />.<br> {{Button Toggle AnimationOnPause}}<br> {{Link Toggle FancyCartoonHighQualityView}} '''Mechanism:''' The reaction mechanism for citrate synthase was proposed by James Remington. In this mechanism, three ionizable side chains in the <scene name='Daniel_Eddelman_Sandbox_2/Cts_active_site/4' >active site</scene> of citrate synthase participate in acid-base catalysis: His 274, His 320, and Asp 375. First, <scene name='Daniel_Eddelman_Sandbox_2/Asp_375/1'>Asp 375</scene> (a base) removes a proton from the methyl group of acetyl-CoA to form its enol. <scene name='Daniel_Eddelman_Sandbox_2/His_274/1'>His 274</scene> stabilizes the acetyl-CoA enolate by forming a hydrogen bond with the enolate oxygen. The enolate then nucleophilically attacks oxaloacetate’s carbonyl carbon, and <scene name='Daniel_Eddelman_Sandbox_2/His_320/1'>His 320</scene> donates a proton to oxaloacetate’s carbonyl group in a concerted step, forming citryl-CoA (which remains bound to the enzyme). Finally, citryl-CoA is hydrolyzed to citrate and CoA. '''Regulation:''' Perhaps the most crucial regulators of the citrate synthase reaction are its substrates, acetyl-CoA and oxaloacetate. Both are present in the mitochondria at concentrations below saturation of citrate synthase. The metabolic flux is controlled by substrate availability, so controlling the levels of acetyl-CoA and oxaloacetate in the mitochondria controls the rate of reaction. Furthermore, citrate synthase is inhibited by NADH, <scene name='Daniel_Eddelman_Sandbox_2/Citrate/2'>citrate</scene> (which competes with oxaloacetate), and succinyl-CoA (an example of competitive feedback inhibition) <ref>PMID:3013232</ref>. <applet load='2cts' size='200' color='white' frame='true' align='right' caption='Citrate Synthase Closed Form of (Monomer) [[2cts]]' name='closed'/> <applet load='1cts' size='200' color='white' frame='true' align='right' caption='Citrate Synthase Open Form (Monomer) [[1cts]]' name='open'/> ==Literature and Notes== <references/> ==Additional Resources== For additional information, see: [[Carbohydrate Metabolism]] <br /> ==External Resources== [http://www.rcsb.org/pdb/static.do?p=education_discussion/molecule_of_the_month/pdb93_1.html Citrate Synthase: September 2007 Molecule of the Month] as part of the series of tutorials that are at [http://www.pdb.org/pdb/home/home.do the RCSB Protein Data Bank] and written by [[User:David_S._Goodsell|David Goodsell]]

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