Tri Carboxylic Acid With Anaplerotic Sequences

TCA CYCLE and its ANAPLEROTIC SEQUENCES •

•

•

•

•

•

•

•

•

•

TCAcycle (tricarboxylic acid cycle; citric acid cycle; Krebs cycle) A cyclic sequence of reactions which plays a central role in the metabolism of many microorganisms, both eukaryotic and prokaryotic, as well as of higher organisms. In eukaryotes the enzymes of the TCA cycle are located in the MITOCHONDRION; in both prokaryotes and eukaryotes most of the enzymes are soluble in the cytoplasm or matrix, respectively, but the succinate dehydrogenase is membrane-bound in both cases. The TCA cycle can have both catabolic and anabolic functions, the relative importance of which often depends largely on the physiological state of the cell. When the TCA cycle is functioning catabolically, acetyl- CoA – derived from carbohydrate metabolism or e.g. from fatty acid degradation – enters the cycle by condensation with oxaloacetate (OAA) to form citrate. With one complete turn of the cycle, 2 molecules of CO2 are eliminated, 3 molecules of NAD(P)+ and one of FAD are reduced, one molecule of ATP or GTP is synthesized by SUBSTRATE-LEVEL PHOSPHORYLATION, and one molecule of OAA is regenerated. Thus, the acetyl group is effectively completely oxidized. The NADH and FADH2 may be reoxidized via a respiratory chain to yield energy. The enzyme isocitrate dehydrogenase is specific for NADP in most bacteria and e.g. in yeasts, and this reaction – together with the HEXOSE MONOPHOSPHATEPATHWAY–is probably an important source of NADPH for biosynthesis in many organisms. When functioning anabolically, the TCA cycle provides precursors for various biosynthetic pathways. Since intermediates withdrawn from the cycle cannot be used to regenerate OAA, OAA must be generated in some other way if the cycle is to operate continuously. Various reactions for achieving this occur in microorganisms: e.g., OAA may be formed directly by the carboxylation of pyruvate or of phosphoenolpyruvate (PEP). Such replenishing reactions are known as ANAPLEROTIC SEQUENCES. Many bacteria can use TCA cycle intermediates (di- and tricarboxylic acids) as sole sources of carbon and energy. However, possession of the TCA cycle does not necessarily enable an organism to grow on such compounds; the presence of a TRANSPORT SYSTEM for the substrate is also necessary. Citrate, for example, can be converted to OAA via reactions of the TCA cycle; however, acetate must then be formed (from another molecule of citrate) if the cycle is to continue. This may be achieved by the decarboxylation of malate (by the ‘malic enzyme’) to pyruvate which, in turn, is converted to acetyl-CoA by the pyruvate dehydrogenase complex. If acetate (or an acetate-generating substrate such as a fatty acid) is used as the sole source of carbon and energy under aerobic conditions, acetate can be converted to acetyl-CoA by acetyl-CoA synthetase with simultaneous formation of AMP and PPi from ATP and can thus be metabolized via the TCA cycle. However, if intermediates are withdrawn from the cycle for biosynthesis, OAA cannot be generated by carboxylation of pyruvate or PEP since in aerobic organisms pyruvate cannot be synthesized directly from acetate. Under these conditions dicarboxylic acids are synthesized by the GLYOXYLATE CYCLE (glyoxylate shunt) involving two reactions, catalysed by isocitrate lyase (ICL) and malate synthase, which bypass the decarboxylation reactions of the TCA cycle. Thus, in effect, one molecule of a 4-C dicarboxylic acid is generated from two of acetate for each turn of the glyoxylate cycle; pyruvate – required e.g. for GLUCONEOGENESIS – can be formed by the decarboxylation of OAA generated in this way. The key enzyme in controlling whether isocitrate is metabolized via the TCA cycle or via the glyoxylate cycle is isocitrate dehydrogenase (ICDH). 1 | TCA 2008

Dr. Shiva C. Aithal, Dept. of Microbiology, Dnyanopasak College, PARBHANI

[email protected]

•

•

•

•

ICDH has a much higher affinity for isocitrate than does ICL. In Escherichia coli, growth on acetate induces the synthesis of a bifunctional enzyme, ICDH kinase–phosphatase, which catalyses the phosphorylation and dephosphorylation of ICDH; phosphorylation inactivates ICDH, allowing isocitrate to reach concentrations sufficient to permit its flow into the glyoxylate cycle. The phosphatase function of ICDH kinase–phosphatase is stimulated, and the kinase function inhibited, by various metabolites– particularly pyruvate. Some bacteria can grow on glyoxylate (formed e.g. by purine degradation), or on glyoxylate precursors such as glycolate, by using the glyoxylate cycle. In this case acetyl-CoA is formed via 3-phosphoglycerate produced by the tartronic semialdehyde pathway (= glycerate pathway) acetyl- CoA may then condense with another molecule of glyoxylate to form malate and hence OAA. TCA cycle enzymes are subject to various control mechanisms affecting their biosynthesis and/or activity. For example, in E. coli they are repressed by anaerobiosis. However, when conditions are such that some of the enzymes are necessary for biosynthesis, the levels of those enzymes are increased despite anaerobiosis; under such conditions 2oxoglutarate dehydrogenase (α-ketoglutarate dehydrogenase) remains repressed and the pathway ceases to be cyclic, functioning in biosynthesis as two linear pathways: an oxidative pathway from citrate to 2-oxoglutarate, and a reductive pathway from OAA to succinylCoA. 2-Oxoglutarate dehydrogenase is also repressed by anaerobiosis in many other facultatively anaerobic organisms. The TCA cycle is inherently incomplete in some bacteria: e.g. in cyanobacteria, in Gluconobacter, and in some members of the Methylococcaceae; in at least some cyanobacteria succinyl-CoA is formed via the glyoxylate cycle rather than via the reductive pathway from OAA. A complete cycle is apparently involved in the oxidation of acetate by sulphate in the obligately anaerobic sulphate-reducing bacterium Desulfobacter postgatei, although some of the enzymes involved are unusual in certain respects: e.g., malate and possibly succinate dehydrogenases appear to use quinones instead of pyridine nucleotides as coenzymes, and 2- oxoglutarate dehydrogenase is specific for a ferredoxin instead of NAD+ as electron acceptor. In D. postgatei, pyruvate (and hence OAA) can be synthesized from acetyl-CoA by direct reductive carboxylation catalysed by a ferredoxin-dependent pyruvate synthase.

2 | TCA 2008

Dr. Shiva C. Aithal, Dept. of Microbiology, Dnyanopasak College, PARBHANI

[email protected]

3 | TCA 2008

Dr. Shiva C. Aithal, Dept. of Microbiology, Dnyanopasak College, PARBHANI

[email protected]

4 | TCA 2008

Dr. Shiva C. Aithal, Dept. of Microbiology, Dnyanopasak College, PARBHANI

[email protected]