Lactate dehydrogenase coenzyme binding

The active center of an LDH subunit is shown schematically in Fig. 2. The peptide backbone is shown as a light blue tube. Also shown are the substrate lactate (red), the coenzyme NAD (yellow), and three amino acid side chains (Arg-109, Arg-171, and His-195 green), which are directly involved in the catalysis. A peptide loop (pink) formed by amino acid residues 98-111 is also shown. In the absence of substrate and coenzyme, this partial structure is open and allows access to the substrate binding site (not shown). In the enzyme lactate NAD"" complex shown, the peptide loop closes the active center. The catalytic cycle of lactate dehydrogenase is discussed on the next page. 

In contrast to the flavin oxidases, flavin dehydrogenases pass electrons to carriers within electron transport chains and the flavin does not react with 02. Examples include a bacterial trimethylamine dehydrogenase (Fig. 15-9) which contains an iron-sulfur duster that serves as the immediate electron acceptor167 169 and yeast flavocytochrome b2, a lactate dehydrogenase that passes electrons to a built-in heme group which can then pass the electrons to an external acceptor, another heme in cytochrome c.170-173 Like glycolate oxidase, these enzymes bind their flavin coenzyme at the ends of 8-stranded a(i barrels similar... 

N. Bernard, K. Johnson, J. J. Holbrook, and J. Delcour, D175 discriminates between NADH and NADPH in the coenzyme binding site of Lactobacillus delbrueckii subsp. bulgaricus D-lactate dehydrogenase, Biochem. Biophys. Res. Commun. 1995, 208, 895-900. 

Scheme 11. Idealized sketch showing the electroen matic oxidation of L-lactate at gold modified electrode surfaces, (a) Lactate dehydrogenase bound to CB-terminated alkylthiol SAMs prepared by covalent attachment of CB to 3-mercaptopropionic acid SAM derivatized with 1,4-diaminobutane. The electroenzymatic oxidation of lactate is observed only in the presence of soluble coenzyme (NAD" ") and a redox mediator (phenazine methosulfate) [215]. (b) Lactate deh3tdrogenase bound to NAD-terminated alkylthiol SAMs prepared by covalent attachment of Af -(2-aminoethyl)-NAD to a cystamine SAM derivatized with pjrrroloquinoline quinone. The reconstituted enzyme is electrically wired to the electrode surface via two NAD" -binding pockets involved in the affinity-binding surface reaction [242].

R. (1992) Raman Spectroscopic Studies of the Effects of Substrate Binding on Coenzymes Bound to Lactate Dehydrogenase, J. Am. Chem. Soc. 114, 7997-8003. 

The raison d etre for an ordered mechanism may be that binding of the coenzyme creates the binding site for the second substrate by inducing a conformational change of the enzyme 23). There is direct evidence for this from recent X-ray diffraction studies of lactate dehydrogenase 24). The conformational change will, however, be manifest kinetically only if it occurs as an elementary step separate from coenzyme binding, thus. 

The enzyme lactate dehydrogenase, which catalyzes the transfer of electrons from lactate to NAD, illustrates these principles (Fig. 8.14). The coenzyme nicotinamide adenine dinucleotide (NAD ) is synthesized from the vitamin niacin (which forms the nicotinamide ring), and from ATP (which contributes an AMP). The ADP portion of the molecule binds tightly to the enzyme and causes conformational... 

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