3a-Hydroxysteroid dehydrogenase

The reduction of this oxobicyclo[3.2.0]heptene using 3a,20/J-hydroxysteroid dehydrogenase from Streptomyces hydrogenans gives the (6S )-< n95% ee) and unreacted optically active ketone. 

The enzyme 3a-hydroxysteroid dehydrogenase (EC 1.1.1.50), which is isolated from Pseudomonas testosteroni, catalyzes the conversion of all 3a-hydroxycholanic acids to 3-ketochoIanic acids, with the concomitant reduction of NAD to NADH. The NADH formed in the reaction is then determined spectrophotometrically at 340 nm. To ensure complete reaction, hydrazine is usually added to bind the 3-keto products (P2). The optimum conditions for enzymatic assay include a pH of 9.0 to 9.5 and reaction temperature in the range 20 to 40 C (T13). Reaction rates for individual bile acids may not be identical, but the assay is normally carried out as an endpoint determination. Alternatively, the addition of bovine serum albumin appears to overcome the problem of variable aflinity of 3a-hy-droxysteroid dehydrogenase for different bile acids if reaction rates are to be measured (S13). 

Hydroxysteroid dehydrogenases /3-HSDH, 20/7-HSDH and 3a-HSDH free and immobilized on CNBr-activated Sepharose (comparative study)... 

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