Stereospecificity of hydrogen transfer by pyridine nucleotide-linked hydroxysteroid dehydrogenases.

The present studies are concerned with the determination of the steric course of the transfer of hydrogen catalyzed by hydroxysteroid dehydrogenases. The classic work of Vennesland et al. (l-3) has demonstrated that pyridine nucleotide-linked dehydrogenases catalyze a direct and stereospecific transfer of hydrogen between the substrate and the para position of the nicotinamide ring of the nucleotide. With the aid of deuterium labeling, it was possible to distinguish two classes of dehydrogenases which transferred the isotope respectively to give one or the other of the two diastereomeric forms of reduced diphosphopyridine nucleotide-nicotinamide-4-d. The first three enzymes to be studied (the dehydrogenases for alcohol, n-lactate, n-malate) all had the same stereospecificity of hydrogen transfer, which was designated as (Y (or side 1)’ in the absence of information on the absolute configuration (4-6). In 1955, two other pyridine nucleotide-linked enzymes were shown likewise to catalyze a direct transfer of hydrogen, but to utilize the other diastereomeric form of reduced DPN in which the reacting hydrogen was designated as fi (side II). These enzymes were the P-hydroxysteroid dehydrogenase of Pseudomonas testosteroni which catalyzed the DPN-dependent interconversion of testosterone and 4-androstene-3,17-dione (7)) and the pyridine nucleotide transhydrogenase of Pseudomonas Juorescens (8). Subsequently, other dehydrogenases which transfer hydrogen to one or the other side of the nicotinamide ring have been described. These studies have been recently extended to several TPNlinked enzymes where analogous stereospecificity with respect to the pyridine nucleotide was observed (9-11). The basic significance of the two types of stereospecificity among these hydrogen transferring reactions has remained obscure. It has been suggested that metabolic coupling of reactions between enzymes of opposite stereospecificity without dissociation of DPN might be facilitated (12). This argument is strengthened by the finding that, for instance, the bound DPN of triosephosphate dehydrogenase (side II stereospecificity) reacts more rapidly with lactic dehydrogenase (side I stereospecificity) than does free DPN (13). Recently, specific dehydrogenases which catalyze the oxidation of either member of an enantiomorphic pair of substrates