Oxaloacetic oxidation

Oxidation. Succinic acid reacts with hydrogen peroxide, giving different products that depend on the experimental conditions peroxysuccinic acid [2279-96-1] (CH2COOOH)2, oxosuccinic acid [328-42-7] (oxaloacetic acid) malonic acid [141-82-2] or a mixture of acetaldehyde, malonic acid, and make acid [6915-15-7]. Succinic anhydride in dimethylformamide (DMF) with H2O2 gives monoperoxysuccinic acid [3504-13-0], HOOCCH2CH2COOOH, mp 107°C (70). 

Potassium permanganate oxidizes succinic acid to a mixture of malic and tartaric acid [133-37-9]. 3-Hydroxypropionic acid [503-66-2] is obtained with sodium perchlorate. Cerium(IV) sulfate in sulfuric acid medium oxidizes succinic acid to oxaloacetic acid (71). 

In 1932 Krebs was studying the rates of oxidation of small organic acids by kidney and liver tissue. Only a few substances were active in these experiments —notably succinate, fumarate, acetate, malate, and citrate (Figure 20.2). Later it was found that oxaloacetate could be made from pyruvate in such tissues, and that it could be further oxidized like the other dicarboxylic acids. 

Another important piece of the puzzle came from the work of Carl Martius and Franz Knoop, who showed that citric acid could be converted to isocitrate and then to a-ketoglutarate. This finding was significant because it was already known that a-ketoglutarate could be enzymatically oxidized to succinate. At this juncture, the pathway from citrate to oxaloacetate seemed to be as shown in Figure 20.3. Whereas the pathway made sense, the catalytic effect of succinate and the other dicarboxylic acids from Szent-Gyorgyi s studies remained a puzzle. 

In 1937 Krebs found that citrate could be formed in muscle suspensions if oxaloacetate and either pyruvate or acetate were added. He saw that he now had a cycle, not a simple pathway, and that addition of any of the intermediates could generate all of the others. The existence of a cycle, together with the entry of pyruvate into the cycle in the synthesis of citrate, provided a clear explanation for the accelerating properties of succinate, fumarate, and malate. If all these intermediates led to oxaloacetate, which combined with pyruvate from glycolysis, they could stimulate the oxidation of many substances besides themselves. (Kreb s conceptual leap to a cycle was not his first. Together with medical student Kurt Henseleit, he had already elucidated the details of the urea cycle in 1932.) The complete tricarboxylic acid (Krebs) cycle, as it is now understood, is shown in Figure 20.4. 

The TCA cycle can now be completed by converting succinate to oxaloacetate. This latter process represents a net oxidation. The TCA cycle breaks it down into (consecutively) an oxidation step, a hydration reaction, and a second oxidation step. The oxidation steps are accompanied by the reduction of an [FAD] and an NAD. The reduced coenzymes, [FADHg] and NADH, subsequently provide reducing power in the electron transport chain. (We see in Chapter 24 that virtually the same chemical strategy is used in /3-oxidation of fatty acids.)... 

Until now we have viewed the TCA cycle as a catabolic process because it oxidizes acetate units to COg and converts the liberated energy to ATP and reduced coenzymes. The TCA cycle is, after all, the end point for breakdown of food materials, at least in terms of carbon turnover. However, as shown in Figure 20.22, four-, five-, and six-carbon species produced in the TCA cycle also fuel avariety of biosynthetic processes. a-Ketoglutarate, succinyl-CoA, fumarate, and oxaloacetate are all precursors of important cellular species. (In order to par-... 

Finally, citrate can be exported from the mitochondria and then broken down by ATP-citrate lyase to yield oxaloacetate and acetyl-CoA, a precursor of fatty acids (Figure 20.23). Oxaloacetate produced in this reaction is rapidly reduced to malate, which can then be processed in either of two ways it may be transported into mitochondria, where it is reoxidized to oxaloacetate, or it may be oxidatively decarboxylated to pyruvate by malic enzyme, with subse-... 

The catabolism of amino acids provides pyruvate, acetyl-CoA, oxaloacetate, fumarate, a-ketoglutarate, and succinate, ail of which may be oxidized by the TCA cycle. In this way, proteins may serve as excellent sources of nutrient energy, as seen in Chapter 26. 

The second electron shuttle system, called the malate-aspartate shuttle, is shown in Figure 21.34. Oxaloacetate is reduced in the cytosol, acquiring the electrons of NADH (which is oxidized to NAD ). Malate is transported across the inner membrane, where it is reoxidized by malate dehydrogenase, converting NAD to NADH in the matrix. This mitochondrial NADH readily enters the electron transport chain. The oxaloacetate produced in this reaction cannot cross the inner membrane and must be transaminated to form aspartate, which can be transported across the membrane to the cytosolic side. Transamination in the cytosol recycles aspartate back to oxaloacetate. In contrast to the glycerol phosphate shuttle, the malate-aspartate cycle is reversible, and it operates as shown in Figure 21.34 only if the NADH/NAD ratio in the cytosol is higher than the ratio in the matrix. Because this shuttle produces NADH in the matrix, the full 2.5 ATPs per NADH are recovered. 

FIGURE 23.5 Pyruvate carboxyl compartmentalized reaction. Pyruva verted to oxaloacetate in the mitoci Because oxaloacetate cannot be trai across the mitochondrial membrant reduced to malate, transported to tl and then oxidized back to oxaloace gluconeogenesis can continue. 

Succinyl-CoA derived from propionyl-CoA can enter the TCA cycle. Oxidation of succinate to oxaloacetate provides a substrate for glucose synthesis. Thus, although the acetate units produced in /3-oxidation cannot be utilized in glu-coneogenesis by animals, the occasional propionate produced from oxidation of odd-carbon fatty acids can be used for sugar synthesis. Alternatively, succinate introduced to the TCA cycle from odd-carbon fatty acid oxidation may be oxidized to COg. However, all of the 4-carbon intermediates in the TCA cycle are regenerated in the cycle and thus should be viewed as catalytic species. Net consumption of succinyl-CoA thus does not occur directly in the TCA cycle. Rather, the succinyl-CoA generated from /3-oxidation of odd-carbon fatty acids must be converted to pyruvate and then to acetyl-CoA (which is completely oxidized in the TCA cycle). To follow this latter route, succinyl-CoA entering the TCA cycle must be first converted to malate in the usual way, and then transported from the mitochondrial matrix to the cytosol, where it is oxida-... 

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.)... 

As its name implies, the citric acid cycle is a closed loop of reactions in which the product of the hnal step (oxaloacetate) is a reactant in the first step. The intermediates are constantly regenerated and flow continuously through the cycle, which operates as long as the oxidizing coenzymes NAD+ and FAD are available. To meet this condition, the reduced coenzymes NADH and FADH2 must be reoxidized via the electron-transport chain, which in turn relies on oxygen as the ultimate electron acceptor. Thus, the cycle is dependent on the availability of oxygen and on the operation of the electron-transport chain. 

Oxidation of (S)-malate gives oxaloacetate, completing the cycle. 

Steps 7-8 of Figure 29.12 Hydration and Oxidation The final two steps in the citric acid cycle are the conjugate nucleophilic addition of water to fumarate to yield (S)-malate (L-malate) and the oxidation of (S)-malate by NAD+ to give oxaloacetate. The addition is cataiyzed by fumarase and is mechanistically similar to the addition of water to ris-aconitate in step 2. The reaction occurs through an enolate-ion intermediate, which is protonated on the side opposite the OH, leading to a net anti addition. 

The final step is the oxidation of (S)-malate by NAD+ to give oxaloacetate, a reaction catalyzed by malate dehydrogenase. The citric acid cycle has now returned to its starting point, ready to revolve again. The overall result of the cycle is... 

Osmium tetroxide, reaction with alkenes, 235-236 toxicity of, 235 Oxalic add, structure of, 753 Oxaloacetic acid, structure of, 753 Oxetane, reaction with Grignard reagents, 680 Oxidation, 233, 348 alcohols, 623-626 aldehydes, 700-701 aldoses, 992-994 alkenes, 233-236 biological, 625-626 phenols, 631 sulfides, 670 thiols, 668... 

The metabolism of amino acids is complex and is described in standard text books. These are usually converted by aminotransferases to the corresponding 2-oxoacids which are partly oxidized in the matrix of muscle mitochondria and partly exported to the liver. Glutamate and aspartate yield 2-oxoglutarate and oxaloacetate, respectively, which enter the citrate cycle directly, and other 2-... 

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 cycle starts with reaction between the acetyl moiety of acetyl-CoA and the four-carbon dicarboxylic acid ox-aloacetate, forming a six-carbon tricarboxylic acid, citrate. In the subsequent reactions, two molecules of COj are released and oxaloacetate is regenerated (Figure 16-1). Only a small quantity of oxaloacetate is needed for the oxidation of a large quantity of acetyl-CoA oxaloacetate may be considered to play a catalytic role. 

The onward metabohsm of succinate, leading to the regeneration of oxaloacetate, is the same sequence of chemical reactions as occurs in the P-oxidation of fatty acids dehydrogenation to form a carbon-carbon double bond, addition of water to form a hydroxyl group, and a hirther dehydrogenation to yield the oxo- group of oxaloacetate. 

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. 

Theoretically, a fall in concentration of oxaloacetate, particularly within the mitochondria, could impair the ability of the citric acid cycle to metabolize acetyl-CoA and divert fatty acid oxidation toward ketogenesis. Such a fall may occur because of an increase in the [NADH]/[NAD+] ratio caused by increased P-oxida-tion affecting the equilibrium between oxaloacetate and malate and decreasing the concentration of oxaloacetate. However, pyruvate carboxylase, which catalyzes the conversion of pyruvate to oxaloacetate, is activated by acetyl-CoA. Consequently, when there are significant amounts of acetyl-CoA, there should be sufficient oxaloacetate to initiate the condensing reaction of the citric acid cycle. 

This cycle is a series of reactions starting from the reaction of oxaloacetic acid with acetylcoenzyme A and finally regenerates oxaloacetic acid again, forming two moles of carbon dioxide during one cycle. The first step of the decarboxylation is conjugated with the oxidation of isocitric acid (Fig. 3). 

The bioluminescent determinations of ethanol, sorbitol, L-lactate and oxaloacetate have been performed with coupled enzymatic systems involving the specific suitable enzymes (Figure 5). The ethanol, sorbitol and lactate assays involved the enzymatic oxidation of these substrates with the concomitant reduction of NAD+ in NADH, which is in turn reoxidized by the bioluminescence bacterial system. Thus, the assay of these compounds could be performed in a one-step procedure, in the presence of NAD+ in excess. Conversely, the oxaloacetate measurement involved the simultaneous consumption of NADH by malate dehydrogenase and bacterial oxidoreductase and was therefore conducted in two steps. 

In the absence of rotenone, the NADH that is made from the conversion of succinate to oxaloacetate can be oxidized by the electron transport chain. The metabolism of succinate then becomes... 

In this case, succinate metabolism to oxaloacetate produces 5 ATPs from the oxidation of the NADH and FADH2—2 from die FADH2 and 3 from the NADH. Two O s are consumed, 1 for each of the NADH and FADH2 molecules. The P/O is then 5/2 = 2.5. Thus the P/O can be a nonintegral number. 

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