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Purification and Characterization of the Corticosteroid 1lβ-Dehydrogenase Component of the Rat Liver 1lβ-Hydroxysteroid Dehydrogenase Complex

Purification and Characterization of the Corticosteroid 1lβ-Dehydrogenase Component of the Rat... We have proposed that 11β-hydroxysteroid dehydrogenase is composed of structurally independent units with 11β-dehydrogenase and 11-reductase activities. We now report the purification of rat liver 11β-dehydrogenase to apparent homogeneity. Starting with microsomes, 800-fold purification was achieved with agarose-NADP affinity chromatography. No 11- reductase accompanied the purification. Homogeneity of 11β- dehydrogenase was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and amino acid end-group analysis and immunoprecipitation. The terminal amino acid was methionine. Monomer mol wt was 34,000. The enzyme was found to be a glycoprotein. A sequence of 40 amino acid units was identified from the amino end. The amino-terminal region was found to be highly nonpolar. Unlike unpurified microsomal 110- dehydrogenase, which showed curvilinear Eadie plots, homogeneous enzyme gave rectilinear plots. Michaelis constants were 1.83 ± 0.06 μM for corticosterone and 17.3 ± 2.24 μM for cortisol. First order rate constants were 10 times greater for corticosterone than cortisol, and maximum velocities were similar. (Endocrinology123: 2390–2398, 1988) This content is only available as a PDF. Author notes * This work was supported by USPHS Grant AM-37094. Protein sequence analysis was performed by the Rockefeller University Protein Sequencing Facility, supported in part by funds provided by the U.S. Army Research Office for the purchase of equipment. Copyright © 1988 by The Endocrine Society http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Endocrinology Oxford University Press

Purification and Characterization of the Corticosteroid 1lβ-Dehydrogenase Component of the Rat Liver 1lβ-Hydroxysteroid Dehydrogenase Complex

LAKSHMI,, VIJAYA; MONDER,, CARL
Endocrinology , Volume 123 (5) – Nov 1, 1988
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References (22)

Publisher
Oxford University Press
Copyright
Copyright © 1988 by The Endocrine Society
ISSN
0013-7227
eISSN
1945-7170
DOI
10.1210/endo-123-5-2390
Publisher site
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Abstract

We have proposed that 11β-hydroxysteroid dehydrogenase is composed of structurally independent units with 11β-dehydrogenase and 11-reductase activities. We now report the purification of rat liver 11β-dehydrogenase to apparent homogeneity. Starting with microsomes, 800-fold purification was achieved with agarose-NADP affinity chromatography. No 11- reductase accompanied the purification. Homogeneity of 11β- dehydrogenase was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and amino acid end-group analysis and immunoprecipitation. The terminal amino acid was methionine. Monomer mol wt was 34,000. The enzyme was found to be a glycoprotein. A sequence of 40 amino acid units was identified from the amino end. The amino-terminal region was found to be highly nonpolar. Unlike unpurified microsomal 110- dehydrogenase, which showed curvilinear Eadie plots, homogeneous enzyme gave rectilinear plots. Michaelis constants were 1.83 ± 0.06 μM for corticosterone and 17.3 ± 2.24 μM for cortisol. First order rate constants were 10 times greater for corticosterone than cortisol, and maximum velocities were similar. (Endocrinology123: 2390–2398, 1988) This content is only available as a PDF. Author notes * This work was supported by USPHS Grant AM-37094. Protein sequence analysis was performed by the Rockefeller University Protein Sequencing Facility, supported in part by funds provided by the U.S. Army Research Office for the purchase of equipment. Copyright © 1988 by The Endocrine Society

Journal

Endocrinology – Oxford University Press

Published: Nov 1, 1988

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