Acetyl-CoA with Oxaloacetate to Form Citrate

Condensation of Acetyl-CoA with Oxaloacetate to Form Citrate 

The first reaction of the TCA cycle is catalyzed by citrate synthase and involves a carbanion formed at the methyl group of acetyl-CoA that undergoes aldol condensation with the carbonyl carbon atom of the oxaloacetate  

Citrate provides the precursors (acetyl-CoA, NADPH) for fatty acid synthesis and is a positive allosteric modulator of acetyl-CoA carboxylase, which is involved in the initiation of long-chain fatty acid synthesis (Chapter 18). It regulates glycolysis by negative modulation of 6-phosphofructokinase activity (see above). All of the above reactions occur in the cytoplasm, and citrate exits from mitochondria via the tricarboxylate carrier. 

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Step 1 is the condensation of acetyl-CoA with oxaloacetate to form citrate, catalyzed by citrate synthase. 

The citric acid cycle begins with the condensation of a molecule of acetyl-CoA with oxaloacetate to form citrate, which is eventually reconverted to oxaloacetate. During this process, two molecules of C02, three molecules of NADH, one molecule of FADH2, and one molecule of GTP are produced. 

The glyoxylate cycle (Figure 17.21), like the citric acid cycle, begins with the condensation of acetyl CoA and oxaloacetate to form citrate, which is then isomerized to isocitrate. Instead of being decarboxylated, isocitrate is cleaved by isocitrate lyase into succinate and glyoxylate. The subsequent steps regenerate oxaloacetate from glyoxylate. Acetyl CoA condenses with... 

The acetyl-CoA derived from amino acid degradation is normally insufficient for fatty acid biosynthesis, and the acetyl-CoA produced by pyruvate dehydrogenase and by fatty acid oxidation cannot cross the mitochondrial membrane to participate directly in fatty acid synthesis. Instead, acetyl-CoA is linked with oxaloacetate to form citrate, which is transported from the mitochondrial matrix to the cytosol (Figure 25.1). Here it can be converted back into acetyl-CoA and oxaloacetate by ATP-citrate lyase. In this manner, mitochondrial acetyl-CoA becomes the substrate for cytosolic fatty acid synthesis. (Oxaloacetate returns to the mitochondria in the form of either pyruvate or malate, which is then reconverted to acetyl-CoA and oxaloacetate, respectively.)... 

The citrate cycle is the final common pathway for the oxidation of acetyl-CoA derived from the metabolism of pyruvate, fatty acids, ketone bodies, and amino acids (Krebs, 1943 Greville, 1968). This is sometimes known as the Krebs or tricarboxylic acid cycle. Acetyl-CoA combines with oxaloacetate to form citrate which then undergoes a series of reactions involving the loss of two molecules of CO2 and four dehydrogenation steps. These reactions complete the cycle by regenerating oxaloacetate which can react with another molecule of acetyl-CoA (Figure 4). 

The citric acid cycle is the final pathway for the oxidation of carbohydrate, Upid, and protein whose common end-metabolite, acetyl-CoA, reacts with oxaloacetate to form citrate. By a series of dehydrogenations and decarboxylations, citrate is degraded, releasing reduced coenzymes and 2CO2 and regenerating oxaloacetate. 

Figure 11.3 Mechanism of transfer of acetyl-CoA out of the mitochondrion. In the mitochondrion, acetyl-CoA reacts with oxaloacetate to form citrate, which is transported across the mitochondrial inner membrane. In the cytosol, citrate is split to re-form citrate and oxaloacetate, catalysed by citrate lyase. It has been shown that inhibition of citrate lyase inhibits fatty acid synthesis.

A. Acetyl CoA enters the TCA cycle by condensing with oxaloacetate to form citrate (Figure 7-2). 

Acetyl-CoA enters the TCA cycle by reacting with oxaloacetate to form citrate. At this point, there is an important flux of citrate leaving the TCA cycle and being exported to the cytosol, where it contributes to lipid... 

Answer In the citric acid cycle, the entering acetyl-CoA combines with oxaloacetate to form citrate. One turn of the cycle regenerates oxaloacetate and produces two C02 molecules. There is no net synthesis of oxaloacetate in the cycle. If any cycle intermediates are channeled into biosynthetic reactions, replenishment of oxaloacetate is essential. Four enzymes can... 

Fatty acids are predominantly formed in the liver and adipose tissne, as well as the mammary glands during lactation. Fatty acid synthesis occurs in the cytosol (fatty acid oxidation occurs in the mitochondria compartmentalisation of the two pathways allows for distinct regulation of each). Oxidation or synthesis of fats utilises an activated two-carbon intermediate, acetyl-CoA, but the acetyl-CoA in fat synthesis exists temporarily bound to the enzyme complex as malonyl-CoA. Acetyl-CoA is mostly produced from pyruvate (pyruvate dehydrogenase) in the mitochondria it is condensed with oxaloacetate to form citrate, which is then transported into the cytosol and broken down to yield acetyl-CoA and oxaloacetate (ATP citrate lyase). 

Gluconeogenesis is the production of glucose from non-carbohydrate substrates, e.g. amino acids. NB 6 ATP equivalents (from P-oxidation, see (4)) are needed to produce 1 glucose molecule. Ketogenesis mitochondrial oxaloacetate is depleted. Therefore, acetyl CoA cannot react with oxaloacetate to form citrate for oxidation in Krebs cycle. Instead, acetyl CoA reacts with itself to form acetoacetate and 3-hydroxybntyrate (the ketone bodies). 

That is, it furnishes pyruvate which enters the tricarboxylic (TCA) cycle, and it serves as an anaplerotic source of oxaloacetic acid (OAA) which couples with acetyl-CoA (from pyruvate) to form citrate, etc. It would be of interest to understand the mechanism by which pyruvate is channeled away from the TCA cycle system to provide acetyl-CoA for fatty acid biosynthesis. 

In the first reaction of the citric acid cycle, acetyl-CoA reacts with oxaloacetate to form citrate. The mechanism for the reaction shows that an aspartate side chain of the enzyme removes a proton from the a-carbon of acetyl-CoA, creating an enolate ion. This enolate ion adds to the keto carbonyl carbon of oxaloacetate and the carbonyl oxygen picks up a proton from a histidine side chain. This is similar to an aldol addition where the a-carbanion (enolate ion) of one molecule is the nucleophile and the carbonyl carbon of another is the electrophile (Section 18.10). The intermediate (a thioester) that results is hydrolyzed to citrate in a nucleophilic addition-elimination reaction (Section 16.9). 

We have seen that the citric acid cycle begins when a two-carbon acetyl group from acetyl-CoA combines with oxaloacetate to form citrate. Through oxidation and reduction, two carbon atoms are removed from citrate to yield two CO2 and a four-carbon compound that undergoes reactions to regenerate oxaloacetate. 

Acetyl-CoA is therefore the substrate for two competing reactions with oxaloacetate to form citrate or with acetoacetyl-CoA to form ketone bodies (ketogenesis). Which reaction predominates depends partly on the rate of -oxidation itself and partly on the redox state of the mitochondrial matrix which controls the oxidation of malate to oxaloacetate, hence, the amount of oxaloacetate available to react with acetyl-CoA. The proportion of acetyl groups going into the TCA cycle relative to ketogenesis is often referred to as the acetyl ratio . The overall rate of -oxidation may be controlled by a number of well-known mechanisms ... 

Citrate is isomerized to isocitrate by the enzyme aconitase (aconitate hydratase) the reaction occurs in two steps dehydration to r-aconitate, some of which remains bound to the enzyme and rehydration to isocitrate. Although citrate is a symmetric molecule, aconitase reacts with citrate asymmetrically, so that the two carbon atoms that are lost in subsequent reactions of the cycle are not those that were added from acetyl-CoA. This asymmetric behavior is due to channeling— transfer of the product of citrate synthase directly onto the active site of aconitase without entering free solution. This provides integration of citric acid cycle activity and the provision of citrate in the cytosol as a source of acetyl-CoA for fatty acid synthesis. The poison fluo-roacetate is toxic because fluoroacetyl-CoA condenses with oxaloacetate to form fluorocitrate, which inhibits aconitase, causing citrate to accumulate. 

The synthesis of palmitate requires the input of 8 molecules of acetyl CoA, 14 molecules of NADPH, and 7 molecules of ATP. Fatty acids are synthesized in the cytosol, whereas acetyl CoA is formed from pyruvate in mitochondria. Hence, acetyl CoA must be transferred from mitochondria to the cytosol. Mitochondria, however, are not readily permeable to acetyl CoA. Recall that carnitine carries only long-chain fatty acids. The barrier to acetyl CoA is bypassed by citrate, which carries acetyl groups across the inner mitochondrial membrane. Citrate is formed in the mitochondrial matrix by the condensation of acetyl CoA with oxaloacetate (Figyu-e 22.25). When present at high levels, citrate is transported to the cytosol, where it is cleaved hy ATP-citrate lyase. 

The citric acid cycle is a central metabolic pathway which generates NADH and FADH2 for use in electron transport. It also produces GTP via substrate-level phosphorylation. Many metabolic processes use intermediates of the citric acid cycle in their pathways. The cyclic process is generally considered to "begin" with addition of acetyl-CoA to oxaloacetate to form citrate. Remember, however, that the pathway is cyclic. 

Citrate synthase binds acetyl CoA, condenses it with oxaloacetate to form citryl CoA, and then hydrolyzes the thioester bond of this intermediate. Why doesn t citrate synthase hydrolyze acetyl CoA ... 

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