Purification and characterization of 7 beta-hydroxysteroid dehydrogenase from Ruminococcus sp. of human intestine.

Abstract:

:7 beta-Hydroxysteroid dehydrogenase (7 beta-HSD) was produced by Ruminococcus sp. PO1-3 obtained from among human intestinal bacteria. The enzyme was purified from a crude extract by ammonium sulfate fractionation, and Butyl-Toyopearl 650M, Sephadex G-150, Matrex Red A and Octyl-Sepharose chromatographies. The purified enzyme was obtained as a single band on polyacrylamide gel electrophoresis with enzyme activity staining and as one band corresponding to a molecular weight of 30,000 on SDS-polyacrylamide gel electrophoresis. On gel filtration, its apparent molecular weight was estimated to be 60,000. The enzyme had a sulfhydryl group(s) in its active site. Substrate specificity studies revealed that the enzyme showed absolute specificity for the beta-configuration of a hydroxyl group at the 7 position of bile acids, and required NADP+ and NADPH as cosubstrates. The Km values for ursodeoxycholic acid, 7-k etolithocholic acid, NADP+, and NADPH were 5.0, 8.5, 7.7, and 24 microM, respectively.

journal_name

J Biochem

journal_title

Journal of biochemistry

authors

Akao T,Akao T,Kobashi K

doi

10.1093/oxfordjournals.jbchem.a122095

subject

Has Abstract

pub_date

1987-09-01 00:00:00

pages

613-9

issue

3

eissn

0021-924X

issn

1756-2651

journal_volume

102

pub_type

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