Properties of a 4-ene-3-ketosteroid-5 alpha-reductase in cell extracts of the intestinal anaerobe, Eubacterium sp. strain 144.
When Eubacterium sp. 144 was grown in the presence of progesterone, extracts of these cells contained a 4-ene-3-ketosteroid-5 alpha-reductase (5 alpha-reductase). No evidence for the presence of a 5 beta-steroid-reductase or a 5 alpha to 5 beta-steroid-isomerase was found. 5 alpha-Reductase activity was dependent on reduced methyl viologen as the electron donor and this could be generated biologically by adding pyruvate or H2 to cell extracts or chemically by adding sodium dithionite. NADH or NADPH with or without flavin nucleotides were not electron donors for 5 alpha-reductase. Most of the 5 alpha-reductase activity (60-65%) of crude extracts was located in the membrane fraction and the enzyme was solubilized by treatment with 1% Triton X-100. Optimum 5 alpha-reductase activity occurred at pH 7.0-7.5 in potassium phosphate buffer but was stimulated by Tris-HCl buffer (pH 8.0-9.0). 5 alpha-Reductase activity was highest at 10% (v/v) methanol and was progressively inhibited by higher methanol concentrations. Sulfhydryl reagents strongly inhibited 5 alpha-reductase but the enzyme was not affected by other metabolic inhibitors. Extracts prepared from cells induced with 16-dehydroprogesterone and grown without hemin contained 5 alpha-reductase and 16-dehydroprogesterone-reductase activities equivalent to those found in extracts of induced cells grown with hemin. This indicates that hemin is not required for the synthesis of active steroid double bond-reductases in strain 144.
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Department of Veterinary Science and Microbiology, North Dakota State University, Fargo 58105.
Saxerud
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Casper
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D042944:Cholestenone 5 alpha-Reductase; D005051:Eubacterium / Q000201:enzymology*; D008401:Gas Chromatography-Mass Spectrometry; D006427:Hemin / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007422:Intestines / Q000382:microbiology; D000432:Methanol / Q000494:pharmacology; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D010088:Oxidoreductases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010269:Paraquat / Q000378:metabolism; D011374:Progesterone / Q000494:pharmacology
D009243:NAD; D011374:Progesterone; D009249:NADP; D006427:Hemin; D010088:Oxidoreductases; D042944:Cholestenone 5 alpha-Reductase; D010269:Paraquat; D000432:Methanol