Reference

Enzymic preparation of 12-ketochenodeoxycholic acid with NADP regeneration

Enzymic preparation of 12-ketochenodeoxycholic acid with NADP regeneration. Carrea, G.; Bovara, R.; Cremonesi, P.; Lodi, R. (Ist. Chim. Ormoni, Milan 20131, Italy). Biotechnol. Bioeng., 26(5), 560-3 (English) 1984. CODEN: BIBIAU. ISSN: 0006-3592. DOCUMENT TYPE: Journal CA Section: 63 (Pharmaceuticals) Section cross-reference(s): 7, 32 12-Ketochenodeoxycholic acid (I) [2458-08-4] was prepd. by oxidizing cholic acid (II) [81-25-4] with NADP [53-59-8] in the presence of 12a-hydroxysteroid dehydrogenase (12a-HSDH) [61642-40-8] and regenerating NADP+ through the glutamate dehydrogenase [9029-12-3]-catalyzed redn. of a-ketoglutarate [328-50-7].

3

3.alpha.-, 7.alpha.- and 12.alpha.-hydroxysteroid dehydrogenase activities from Clostridium perfringens. MacDonald, Ian A.; Meier, Elizabeth C.; Mahony, David E.; Costain, Gary A. (Dep. Med., Dalhousie Univ., Haifax, Nova Scotia, Can.). Biochim. Biophys. Acta, 450(2), 142-53 (English) 1976. CODEN: BBACAQ. DOCUMENT TYPE: Journal CA Section: 10 (Microbial Biochemistry) Section cross-reference(s): 7 Twenty-five strains of C. perfringens were screened for hydroxysteroid dehydrogenase activity; 19 contained NADP-dependent 3.alpha.-hydroxysteroid dehydrogenase and 8 contained NAD-dependent 12.alpha.-hydroxysteroid dehydrogenase active against conjugated and unconjugated bile salts. All strains contg. 12.alpha.-hydroxysteroid dehydrogenase also contained 3.alpha.-hydroxysteroid dehydrogenase although 12.alpha.-hydroxysteroid dehydrogenase was invariably in lesser quantity than the 3.alpha.-hydroxysteroid dehydrogenase. In addn., 7.alpha.-hydroxysteroid dehydrogenase activity was evident only when 3.alpha.,7.alpha.,12.alpha.-trihydroxy-5.beta.-cholanoate was substrate but notably absent when 3.Several reagents with their cas registry numbers 39361-64-3 and 61642-40-8 are used here.alpha.,7.alpha.-dihydroxy-5.beta.-cholanoate was substrate. The oxidn. product 12.alpha.-hydroxy-3,7-diketo-5.beta.-cholanoate is rapidly further degraded to an unknown compd. devoid of either 3.alpha.- or 7.alpha.-OH groups. Group specificity of these enzymes was confirmed by thin-layer chromatog. studies of the oxidn. products. These enzyme systems appear to be constitutive rather than inducible. In contrast to C. perfringens, C. paraputrificum (5 strains tested) contained no measurable hydroxysteroid dehydrogenase activity. The C. perfringens enzymes revealed a sharp pH optimum at pH 11.3 and 10.5 for the 3.alpha.-OH- and 12.alpha.-OH-oriented activities, resp. Kinetic studies gave Km ests. of .apprx.5 .times. 10-5 and 8 .times. 10-4 M with 3.alpha.,7.alpha.-dihydroxy-5.beta.-cholanoate and 3.alpha.,12.alpha.-dihydroxy-5.beta.-cholanoate as substrates for the 2 resp. enzymes. 3.alpha.-Hydroxysteroid dehydrogenase was active against 3.alpha.-OH-contg. steroids such as androsterone regardless of the stereochem. of the 5H (both A/B cis and A/B trans steroids were substrates). There was no activity against 3.BETA.-OH-contg. steroids. The 3.alpha.- and 12.alpha.-hydroxysteroid dehydrogenase activities, although differing in cofactor requirements cannot be distinguished by their appearance in the growth curve, their mobility on disc gel electrophoresis, elution vol. on passage through Sephadex G-200 or heat inactivation studies. .