TRANSHYDROGENASE REACTIONS AND ESTROGENS 179 



dependent transfer of hydrogen from DPNH to acetylpyridineDPN. 

 The latter transh)'drogenation is inliibited by low concentrations of 

 TPN as is the case with the placental enzyme. Hydrogen transfer 

 from TPNH to DPN or to acetylpx ridineDPN by the hepatic soluble 

 3a-h\'droxysteroid dehydrogenase can be demonstrated only if the 

 concentration of TPNH is kept very low in comparison with the con- 

 centration of the acceptor nucleotides. This accounts for the fact 

 tliat Stein and Kaplan ( 1959 ) were unable to demonstrate more than 

 negligible rates of hydrogen transfer from high concentrations of 

 TPNH to acet\ lp\ridineDPN when soluble liver extracts were in- 

 cubated witli androsterone. 



The 3a- and the (3- and 17-)^-hydroxysteroid dehydrogenases of 

 Fseudomonas testosteroni (Marcus and Talalav, 1956) are two en- 

 zymes of this class which have been isolated in a high state of purity 

 (cf. Talalay, 1957a, 1957b). Neither of these enzymes reacts with 

 TPN, but Talalay and Adams have demonstrated that they reduce 

 a number of pyridine nucleotide analogues in the presence of their 

 steroid substrates (cf. Talalay and Williams-Ashman, 1960). They 

 also promote the transfer of hydrogen from DPNH to certain ana- 

 logues upon the addition of catalytic levels of appropriate steroids. 

 Thus, Talalay and Adams found that the bacterial 3a-hydroxysteroid 

 deliydrogenase catalyzes an androsterone-dependent transhydroge- 

 nation between DPNH and the acetylpyridine, pyridinealdehyde, 

 and thionicotinamide analogues of DPN. The bacterial (3- and 

 17-)yS-hydroxysteroid dehydrogenase catalyzes the reduction of pyri- 

 dinealdehydeDPN by DPNH in the presence of catalytic levels of 

 4-androstene-3, 17-dione. In these model transhydrogenations with 

 bacterial hydroxysteroid dehydrogenases, the rates of hydrogen 

 transfer were most sensitive to the relative concentrations of donor 

 and acceptor nucleotides in the reaction mixture. 



It would appear that the high affinity of hydroxysteroid dehydro- 

 genases for steroids and the favorable equilibrium constants for the 

 oxidoreduction of steroids by pyridine nucleotides are two factors 

 which determine the peculiar efficiency with which hydroxysteroid 

 dehydrogenases catalyze transhydrogenations between pyridine nu- 

 cleotides in the presence of very low levels of steroids (Talalay and 

 Wilhams- Ashman, 1958, I960).' 



