L-Lactate: NAD oxidoreductase

Lactate dehydrogenase (EC 1.1.1.27 L-lactate NAD" oxidoreductase LD) is a hydrogen transfer enzyme that catalyzes the oxidation of L-lactate to pyruvate with the mediation of NAD as a hydrogen acceptor. 

There were two clearly separated peaks, C and D, sedimenting at 1.2 M and 1.3 M-sucrose (Fig. 2). Examination by the electron microscope showed them both to have the morphological characteristics of synaptosomes. It is well established that a good enzyme marker for intact synaptosomes is occluded lactic dehydrogenase (L-lactate NAD oxidoreductase, EC 1.1.1.27) (Marchbanks, 1967), a component of the cell sap. As can be seen in Table I, 83 % of the occluded form of the enzyme of the original P2 fraction is shared between peaks C and D. Both also contained succinic dehydrogenase activity owing to the presence of intraterminal mitochondria. The membrane marker acetylcholinesterase (acetylcholine hydrolase, EC 3.1.1.7) was also present in these peaks and was notably absent from the mitochondrial and lysosomal fractions (Table I). 

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. 

Lactate dehydrogenase is a pyridine nucleotide oxidoreductase, a tetramer of 140 kD molecular weight, which has been extensively investigated (Bloxham et al., 1975 Eventoff et al., 1977). It catalyses the reversible oxidation of L-lactate to pyruvate using NAD+ as a coenzyme. The reaction scheme with a view of the active site with bound substrate and essential amino-acid side chains are depicted in Equation (3) and in Figure 17. The probable reaction mechanism, involving proton and hydride transfers,... 

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