Glyoxylate+Cycle

=The Glyoxylate Cycle = **Authors: Nancy Garcia, Megan Lawson, and Charles Lewis**

The Glyoxylate Cycle, so named for its unique two-carbon intermediate, is a metabolic cycle that functions as a modification of the Tri-Carboxylic Acid Cycle providing for a net fixation of carbon.



This cycle uses six intermediates and one coenzyme found in the TCA cycle to convert Acetyl-CoA to Oxaloacetate through a series of additions and transformations. Abbreviation of the TCA cycle is accomplished by eliminating all steps involving evolution of CO 2. In this manner, the glyoxylate cycle is able to provide the important cellular intermediate Succinate, without loss of carbon to the atmosphere.

Table of Contents:  1. Overview  2. Sequence of Reactions  - 2.A Claisen Condensation of Oxaloacetate with Acetyl-CoA  - 2.B Isomerization of Citrate  - 2.C Cleavage of Isocitrate  - 2.D Synthesis of Malate  - 2.E Oxidation of Malate to Oxaloacetate  3. Regulation  - 3.A Citrate Synthase  - 3.B Aconitase  - 3.C Isocitrate Lyase <span style="font-family: 'Courier New',Courier,monospace;"> - 3.D Malate Synthase <span style="font-family: 'Courier New',Courier,monospace;"> - 3.E Malate Dehydrogenase <span style="font-family: 'Courier New',Courier,monospace;"> 4. Investigation History <span style="font-family: 'Courier New',Courier,monospace;"> 5. References



<span style="font-family: 'Courier New',Courier,monospace;">1. Overview:
<span style="font-family: 'Courier New',Courier,monospace;">In plants, the Glyoxylate Cycle, located in organelles known as Glyoxysomes, provides a method for plants to utilize the oxidation of Fatty Acids or Acetate from the soil as the carbon source required for growth. Certain yeast and algae are also capable of performing the Glyoxylate Cycle, but having no organelles, their Glyoxylate Cycles operates in the cytoplasm.

<span style="font-family: 'Courier New',Courier,monospace;"> The overall reaction for this cycle is: 2 CH 3 -CO-S-CoA + 2 H 2 O + NAD + = - OOC-CH 2 -CH 2 -COO - + 2 HS-CoA + NADH + 3 H +



<span style="font-family: 'Courier New',Courier,monospace;"> One primary function of the Glyoxylate Cycle is to replenish the Tricarboxylic and Dicarboxylic Acid intermediates that are normally provided by the Krebs Cycle. The Glyoxylate Cycle is primarily an oxidative pathway in which Acetyl-CoA is generated from the oxidation of Acetate, which usually is derived from the oxidation of fatty acids. Pyruvate oxidation is not directly involved in the Glyoxylate train, yet this yields sufficient Succinate and Malate, which are required for energy production. The cycle also generates other precursor compounds needed for biosynthesis. (NCBI Database)



<span style="font-family: 'Courier New',Courier,monospace;">**2.A Claisen Condensation of Oxaloacetate with Acetyl-CoA**
<span style="font-family: 'Courier New',Courier,monospace;">The first step of the Glyoxylate Cycle, which is identical to that of the TCA Cycle, is accomplished by general acid-base catalysis using an Aspartic acid and a Histidine residue of the enzyme Citrate Synthase as the acid and base sources, respectively. This process condenses Acetyl-CoA and Oxaloacetate to form (S)-Citryl-CoA. As soon as this is completed, the Citryl-CoA is hydrolyzed by the same enzyme, splitting off HS-CoA and adding the other OH - to form a Citrate Anion, because the proton is immediately abstracted by the basic environment. (McMurry, p.187)





<span style="font-family: 'Courier New',Courier,monospace;">**2.B Isomerization of Citrate**
<span style="font-family: 'Courier New',Courier,monospace;">The second transformation in the Glyoxylate Cycle is catalyzed by Aconitase, which dehydrates the symmetric Citrate molecule at its 3 carbon to form the asymmetric //cis//-Aconitase molecule by using an iron-sulfur cluster to draw away the hydroxyl group from the 3 carbon, while a generic base abstracts the proton on the opposite face of the new double bond. The Aconitase enzyme then rehydrates the //cis//-Aconitase molecule at its 2 carbon with the hydroxyl group taken off the 3 carbon, and places the hydrogen from the 2 carbon onto the 3 carbon, forming the chiral molecule (2R, 3S)-Isocitrate. What is most noteworthy about this reaction is the fact that the Isocitrate molecule formed will always be 2R, 3S, due to the conformation of the enzyme and the difference in shape between the hydroxyl group and the lone hydrogen that share carbon 3. (McMurry, pp.188-9)





<span style="font-family: 'Courier New',Courier,monospace;">**2.C Cleavage of Isocitrate**
<span style="font-family: 'Courier New',Courier,monospace;">At this point, the Glyoxylate Cycle first differs from the TCA Cycle by producing Succinate in a single step, rather than the three steps required in TCA. In doing so, the Glyoxylate Cycle bypasses the evolution of two CO 2 molecules, saving carbon for use elsewhere in the cell. The enzyme Isocitrate Lyase accomplishes this by a general acid-base catalysis, in which a base abstracts the proton of the terminal alcohol, while the bond between the newly formed carbonyl and its tertiary α-carbon is broken to relieve the extraneous fifth bond of the new carbonyl. As that bond is broken, the former α-carbon takes a proton from a generic acid within the enzyme. In breaking this bond, the two-carbon Glyoxylate and the four-carbon Succinate are released, the first to continue the cycle, the second to return to the TCA or ETC Cycles. (Garrett, p.588)





<span style="font-family: 'Courier New',Courier,monospace;">**2.D Synthesis of Malate**
<span style="font-family: 'Courier New',Courier,monospace;">The fourth step of the Glyoxylate Cycle is where it rejoins the TCA Cycle with the synthesis of Malate from Glyoxylate and Acetyl-CoA. Malate Synthase, like Citrate Synthase, catalyzes a Claisen Condensation between the carbonyl carbon of glyoxylate and the α-carbon of Acetyl-CoA, forming Malate and ejecting an HS-CoA. (Garret, p.588)





<span style="font-family: 'Courier New',Courier,monospace;">**2.E Oxidation of Malate to Oxaloacetate**
<span style="font-family: 'Courier New',Courier,monospace;">The closing step of the cycle is the transformation of Malate to Oxaloacetate by dehydrogenation. This oxidation, being highly endergonic, can only be accomplished by the concomitant reduction of NAD + to NADH + H + under catalysis by Malate Dehydrogenase.





<span style="font-family: 'Courier New',Courier,monospace;">3. Regulation:
<span style="font-family: 'Courier New',Courier,monospace;">Regulation of the cycle inhibits unwanted reactions and activates the desired transformations using specific enzymes. The Glyoxylate Cycle is regulated by ATP, NADH, Acetyl-CoA, Citrate, Succinyl-CoA, Iron Ions, Nitro Analogs, Glutathione, Glutaredoxin, α-<span style="font-family: 'Courier New',Courier,monospace;">Ketoglutarate Dehydrogenase, Mitochondrial Aspartate Aminotransferase, α-<span style="font-family: 'Courier New',Courier,monospace;">Ketoglutarate, and Citrate. The regulation uses some enzymes that are not found in the glyoxysome and must go through the mitochondria then back to the glyoxysome to control the desired enzyme. Succinate must travel to the mitochondria to be converted to Oxaloacetate and is then converted to Aspartate to return to the glyoxysomes. The Aspartate is then converted to α-<span style="font-family: 'Courier New',Courier,monospace;">Ketoglutarate which finally converts to the desired Oxaloacetate.  <span style="font-family: 'Courier New',Courier,monospace;">**3.A Citrate Synthase** <span style="font-family: 'Courier New',Courier,monospace;">  <span style="font-family: 'Courier New',Courier,monospace;">**3.B Aconitase** <span style="font-family: 'Courier New',Courier,monospace;">  <span style="font-family: 'Courier New',Courier,monospace;">**3.C Isocitrate Lyase** <span style="font-family: 'Courier New',Courier,monospace;">  <span style="font-family: 'Courier New',Courier,monospace;">**3.D Malate Synthase** <span style="font-family: 'Courier New',Courier,monospace;">  <span style="font-family: 'Courier New',Courier,monospace;">**3.E Malate Dehydrogenase** <span style="font-family: 'Courier New',Courier,monospace;">
 * ~ Regulator ||~ Action ||~ Location ||
 * ATP || Inhibitor || Allosteric ||
 * NADH || Inhibitor || Allosteric ||
 * Acetyl-CoA || Inhibitor || Allosteric ||
 * Citrate || Inhibitor || Allosteric ||
 * Succinyl-CoA || Inhibitor || Allosteric ||
 * ~ Regulator ||~ Action ||~ Location ||
 * Iron Ions || Activator || Allosteric ||
 * Nitro Analogs || Inhibitor || Allosteric ||
 * ~ Regulator ||~ Action ||~ Location ||~ Type ||
 * Glutathione || Inhibitor || Allosteric || Post-Translational Modification ||
 * Glutaredoxin || Activator || Allosteric || Post-Translational Modification ||
 * ~ Regulator ||~ Action ||~ Location ||
 * None Found ||
 * ~ Regulator ||~ Action ||~ Location ||
 * α-Ketoglutarate Dehydrogenase || Activator || Allosteric ||
 * Mitochondrial Aspartate Aminotransferase || Activator || Allosteric ||
 * α-Ketoglutarate || Inhibitor || Allosteric ||
 * Citrate || Inhibitor || Allosteric ||



<span style="font-family: 'Courier New',Courier,monospace;">4. Investigation History:
<span style="font-family: 'Courier New',Courier,monospace;"> The Glyoxylate Cycle was discovered by Dr. Hans Krebs, FRS and Dr. Hans Kornberg, FRS in 1957; their findings were published in the journal //Nature// under the title "Synthesis of Cell Constituents from C2-Units by a Modified Tricarboxylic Acid Cycle" (//Nature// **179**, 988-991 (18 May 1957) | doi:10.1038/179988a0). This was the third metabolic cycle Krebs had studied, identified, and named, the most noted, of course, being the Krebs Cycle. (Medical News Website, "Metabolism History")The Glyoxylate Cycle was discovered as an unusual metabolic pathway during an attempt to learn how Lipid (or Acetate) oxidation in bacteria and plant seeds could lead to the direct biosynthesis of carbohydrates. The Glyoxylate Cycle is found in many bacteria, including Azotobacter //vinelandii// and particularly often in organisms which thrive in media where acetate and other Krebs Cycle Dicarboxylic Acid intermediates are the sole carbon growth source. (NCBI Database)



<span style="font-family: 'Courier New',Courier,monospace;">5. References
<span style="font-family: 'Courier New',Courier,monospace;">1. McMurry, John; Begley, Tadhg. //The Organic Chemistry of Biological Pathways//. 2005, Roberts and Company Publishers, Colorado.

<span style="font-family: 'Courier New',Courier,monospace;"> 2. Garrett, Reginald H.; Grisham, Charles M. //Biochemistry, Fourth Edition//. 2010. Brooks/Cole, Massachusetts.

<span style="font-family: 'Courier New',Courier,monospace;"> 3. KEGG Database; http://www.genome.jp Accessed December 12, 2010

<span style="font-family: 'Courier New',Courier,monospace;"> 4. 3DMET Database; http://www.3dmet.dna.affrc.go.jp Accessed December 12, 2010

<span style="font-family: 'Courier New',Courier,monospace;"> 5. "Metabolism History", The Medical News website, http://www.news-medical.net/health/Metabolism-History.aspx Accessed December 12, 2010

<span style="font-family: 'Courier New',Courier,monospace;">6. NCBI Database; http://www.ncbi.nlm.nih.gov Accessed December 12, 2010