Citrate into Succinate

Some LAB species cannot truly convert citrate into pyruvate. Instead, the CitT transporter genraates succinate via malate and fumarate. Furthermore, the complete tricarboxylic acids (TCA) pathway has recently been identified in the Lact. casei genome using in silica analysis (Dfaz-Muniz et al. 2006). In this LAB species, the dominant end-products of citrate metabolism wctb acetic acid and L-lactic acid at both excess and limiting amounts of carbohydrates. Trace amounts of D-lactic acid, acetoin, formic acid, ethanol, and diacetyl confirm OAD activity however, succinic acid, malic acid, and butanendiol were not observed (Dfaz-Muniz et al. 2006 Mortera el al. 2013). 

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First, all the individual stages which constitute the cycle have been demonstrated to occur in muscle tissue, and the rates at which the individual reactions can proceed are sufficient to account for the maximum rate of respiration. The occurrence of some of the reactions in muscle tissue, as already mentioned, has been known since 1911, when Batelli and Stern demonstrated the rapid oxidation of citrate, succinate, fuma-rate, and malate in frog muscle. In 1936 the work of Martiusand Knoop - revealed the mechanism of the conversion of citrate into succinate, and the last major step of the cycle was discovered in 1937, when the formation of citrate from oxalacetate and pyruvate was demonstrated. It was this reaction which made a series of reactions into a cyclic sequence and which linked the series of reactions leading from citrate to oxalacetate with carbohydrate metabolism. 

Succinic acid can also be generated from fumarate [100] or citrate [101] in the presence of a readily metabolizable carbon source to serve as the hydrogen donor. When citrate is the hydrogen acceptor, it is split into oxaloacetate and acetate by citrate lyase. Oxaloacetate is in turn converted into succinate [102]. The rate of conversion and yield of succinate from fumarate can be enhanced by the amplification of genes that synthesize fumarate reductase [103, 104]. Table 1 shows the fermentation results reported by Wang et al. [104]. In addition, succinic acid can be generated from glucose with mixed culture fermentation, in which fumarate produced by a Rhizopus culture is converted into succinate by a bacterial culture [105]. 

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... 

Pyruvate produced by glycolysis is transformed by oxidative decarboxylation into acetyl-GoA in the presence of coenzyme A. Acetyl-GoA then enters the citric acid cycle by reacting with oxaloacetate to produce citrate. The reactions of the citric acid cycle include two other oxidative decarboxylations, which transform the six-carbon compound citrate into the four-carbon compound succinate. The cycle is completed by... 

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. 

Retey et al. (72) used this same principle in their work on the stereochemistry of citrate formation from chiral acetyl-CoA with Si-citrate synthase (Scheme 15). The chiral methyl group was converted into one of the methylene groups of succinate and the distinction between sets (I, 2, 3) and (4, 5, 6) was then based on the known different isotope effects for the removal of pro-/ ( hMd = 5.3 kH/kT = 12) vs. pro-5 (kH/kD = 1.35 kH/kj = 1.5) hydrogens of succinate in the succinate dehydrogenase reaction (73). However, the malate synthase/fumarase procedure is clearly the most commonly used method to analyze the configuration and chiral purity of chiral methyl groups. 

Carboxylic acids such as pyruvate, succinate, and citrate are transported into the matrix by the pyruvate transporter, the dicarboxylic acid transporter, and the tricarboxylic acid transporter, respectively. Pymvate transport operates as an antiporter with hydroxide ion. The other transporters are driven by concentration gradients for their substrates. For example, high concentrations of citrate in the matrix lead to export of citrate to the cytoplasm, where it can inhibit phospho-fmctokinase (see Chapter 9). 

The entry of acetyl groups from acetyl CoA into the citric acid cycle does not contribute to the net synthesis of oxaloacetate, because two carbons are lost as CO2 in the pathway from citrate to oxaloacetate. Only the entry of compounds with three or more carbons, like succinate, can increase the relative number of carbon atoms in the pathway. Thus, while odd-numbered fatty acids contribute to the net synthesis of oxaloacetate, those compounds with an even number of fatty acids do not. 

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