help button home button Endocrine Society Endocrinology
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
This Article
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Shen, C. C.
Right arrow Articles by Kochakian, C. D.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Shen, C. C.
Right arrow Articles by Kochakian, C. D.

Endocrinology, Vol 103, 2040-2052, Copyright © 1978 by Endocrine Society

ARTICLES

Purification and properties of 17 beta-hydroxy-C19-steroid dehydrogenases of adult male guinea pig kidney

CC Shen and CD Kochakian

The 17 beta-hydroxy-C19-steroid dehydrogenase activity of adult male guinea pig kidney was separated into one isolated and two contiguous enzymatically pure fractions by submitting the cytosol to (NH4)2SO4 (40- 80% saturation) precipitation, Sephadex G-75 filtration, and DEAE- Sephadex A-50 and CM-Sephadex C-50 chromatography. Further DEAE- Sephadex A-50 and CM-Sephadex C-50 chromatography separated eight isozymes, which were divided into four groups in accordance with their behavior on chromatography and mobility on gel electrophoresis. The molecular weights of the purified enzymes were identical by Sephadex filtration (33,000) and SDS gel electrophoresis (34,000). Two of the enzymes were separated by SDS gel electrophoresis into two subcomponents of 23,000 and 11,000 daltons. The amino acid composition, Km values, and coenzyme and substrate specificities of the enzymes (except one) were very similar. The pI values varied from 5.1-6.4 HgCl2 and PCMB inhibited enzyme activity, but prior addition of cysteine prevented the inhibition. The presence of phosphate or pyrophosphate greatly enhanced the trace of DPN+ -linked activity. The heterogeneity was due to at least two factors. Rapid (within 3 h) preparation of the cytosol in 7 mM 2-mercaptoethanol yielded one intense and one minor enzyme band on gel electrophoresis. Omission of the mercaptoethanol resulted in the appearance of three enzyme bands, which were reversible on the addition of 2-mercaptoethanol. Storage of the cytosol at 4 or - 20 C in the absence or presence of 7 mM 2-mercaptoethanol and of the kidney before extraction also resulted in the exhibition of enzyme bands which were not reversible on addition of 2-mercaptoethanol. The purified enzymes exhibited only a single form on storage at -20 C with 2-mercaptoethanol, but multiple forms appeared when the mercaptoethanol was removed by dialysis. The multiple forms were reverted to a single form on addition of mercaptoethanol.


This article has been cited by other articles:


Home page
 J. Biol. Chem.Home page
Y. Deyashiki, K. Ohshima, M. Nakanishi, K. Sato, K. Matsuura, and A. Hara
Molecular Cloning and Characterization of Mouse Estradiol 17[IMAGE]-Dehydrogenase (A-Specific), a Member of the Aldoketoreductase Family
J. Biol. Chem., May 5, 1995; 270(18): 10461 - 10467.
[Abstract] [Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Endocrinology Endocrine Reviews J. Clin. End. & Metab.
Molecular Endocrinology Recent Prog. Horm. Res. All Endocrine Journals
Copyright © 1978 by The Endocrine Society