Enzymes glyceraldehyde 3-phosphate dehydrogenase

Thus far, we have considered enzyme-catalyzed reactions involving one or two substrates. How are the kinetics described in those cases in which more than two substrates participate in the reaction An example might be the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (Chapter 19) ... 

The most confusing aspect of the pathway proposed by Ochoa and his group now rests with the NAD requirement. In proceeding from L-malic acid to L-lactic acid, there is no net change in oxidation state. Yet in whole cells or cell-free extracts, the malo-lactic fermentation will not proceed in the absence of NAD. Therefore, by the proposed mechanism, one is unable to demonstrate the appearance of reduced cofactor, and the NAD specificity cannot be explained as a redox requirement. However, in the time since this mechanism was proposed, an NAD dependent enzyme (glyceraldehyde-3-phosphate dehydrogenase) has been described which requires NAD in a non-redox capacity (29), and it is possible that the same is true for the enzyme causing the malic acid-lactic acid transformation. 

One of the major integral proteins of the erythrocyte membrane is the anion channel, or band-3 protein, which moves Cl- and HC03 anions across the membrane. The anion transporter has two identical subunits with molecular weights of about 95,000, and each subunit probably has 10 or 11 transmembrane helices. The band-3 protein is attached to the spectrin cytoskeleton through a smaller protein, anky-rin. The cytosolic domain of the anion transporter also binds the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase. 

Gregus, Z. and Nemeti, B. (2005) The glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase works as an arsenate reductase in human red blood cells and rat liver cytosol. Toxicological Sciences, 85(2), 859-69. 

This metabolic scheme, which is called lactate fermentation, is shown in Fig. 11-7. The coreactant cycle between the two dehydrogenase enzymes, glyceraldehyde-3-phosphate dehydrogenase (Step 6) and lactate dehydrogenase, ensures that there is regeneration of NAD+ in this particular oxidation state so that glycolysis, lactate fermentation, and the production of ATP can continue. 

Afterwards, the enzyme glyceraldehyde-3-phosphate dehydrogenase transforms glyceraldehyde-3-phosphateinto 1,3-diphosphoglycerate. This reaction involves the oxidation of the molecule that is linked to reducing NAD+ to NADH in order to redress the redox balance. Simultaneously, a substrate level phosphorylation takes... 

Several different amino acid side chains can act as nucleophiles in enzyme catalysis. The most powerful nucleophile is the thiol side chain of cysteine, which can be deproto-nated to form the even more nucleophilic thiolate anion. One example in which cysteine is used as a nucleophile is the enzyme glyceraldehyde 3-phosphate dehydrogenase, which uses the redox coenzyme NAD+. As shown in Fig. 10, the aldehyde substrate is attacked by an active site cysteine, Cys-149, to form a hemi-thioketal intermediate, which transfers hydride to NAD+ to form an oxidized thioester intermediate (7). Attack of phosphate anion generates an energy-rich intermediate 3-phosphoglycerate. 

Glyceraldehyde 3-phosphate is oxidized by NAD+ and reacts with inorganic phosphate (Pi) to form 1,3-bisphosphoglycerate and NADH + H+. -Enzyme glyceraldehyde 3-phosphate dehydrogenase... 

A cartoon representation of the oxidation path used in Equation 9.5 is shown in Scheme 9.5. The process is catalyzed by the enzyme glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.12). A large body of evidence has been accumulated about the role of the enzyme and, in particular, that the catalytic site on the enzyme that is used to help the oxidation occur bears a thiol (-SH, Chapter 8) group (from the amino acid cysteine. Chapter 12). Further, it has been shown that hydride (H ) transfer to nicotinamide dinucleotide (NAD ") (see Chapter 12), a required cofactor that accounts for the transfer of the hydride anion and, thus, the oxidation itself, is also involved. 

Little, C., and O Brien, P. J., 1969, Mechanism of peroxide inactivation of the sulfhydryl enzyme glyceraldehyde-3-phosphate dehydrogenase, Eur. J. Biochem. 10 533-538. 

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