Carboxyl-Terminal and Arg38 are Essential for Activity of the 7α-Hydroxysteroid Dehydrogenase from Clostridium absonum

ISSN: 1875-5305 (Online)
ISSN: 0929-8665 (Print)


Volume 21, 12 Issues, 2014


Download PDF Flyer




Protein & Peptide Letters

Aims & ScopeAbstracted/Indexed in


Submit Abstracts Online Submit Manuscripts Online

Editor-in-Chief:
Prof. Ben M. Dunn
Department of Biochemistry and Molecular Biology
University of Florida
College of Medicine
P.O. Box 100245
Gainesville, FL
USA
Email: bdunn@ufl.edu

View Full Editorial Board

Subscribe Purchase Articles Order Reprints

Current: 1.735
5 - Year: 1.493

Carboxyl-Terminal and Arg38 are Essential for Activity of the 7α-Hydroxysteroid Dehydrogenase from Clostridium absonum

Author(s): Deshuai Lou, Bochu Wang, Jun Tan and Liancai Zhu

Affiliation: College of Bioengineering, Chongqing University, No.174, Shapingba Main Street, Chongqing, P.R. China.

Abstract

7α-hydroxysteroid dehydrogenase (7α-HSDH, EC 1.1.1.159), one of the short-chain dehydrogenase/reductase superfamily, catalyzes the dehydrogenation of C7 hydroxyl group of the steroid skeleton of bile acids. Clostridium absonum 7α-HSDH (Ca 7α-HSDH) was cloned and heterologously expressed in Escherichia coli. The function of carboxyterminal (C-terminal) and Arg38 of Ca 7α-HSDH was investigated through truncations and site-directed mutagenesis, respectively. When 2 and 6 amino acids of C-terminal were removed, the catalytic efficiency (kcat/Km) of Ca 7α-HSDH remained 19.1% and 2.5%, respectively. Furthermore, the activity could not be detected after 8, 14 and 17 amino acids were deleted. No activity could be detected with coenzyme either NADP+ or NAD+ after replacement of arginine at position 38 by aspartic acid. The metal ions Mg2+ (50 mM), Na+ (200 mM) and K+ (500 mM) could maximally improve the activity of Ca 7α-HSDH by 61.4%, 64.7% and 105.7%, respectively. The activity had no significant change after incubation at 4 or 25 °C for 108 h, but decreased dramatically at 37 °C. Our study confirmed that C-terminal and Arg38 were essential for the catalytic function of Ca 7α-HSDH and the enzyme activity can be improved by metal ions.




Keywords: Bile acid, hydroxysteroid dehydrogenase, site-directed mutagenesis, taurochenodeoxycholic acid, tauroursodeoxycholic acid, truncation.

Purchase Online Order Reprints Order Eprints Rights and Permissions

  
  



Article Details

Volume: 21
Issue Number: 9
First Page: 894
Last Page: 900
Page Count: 7
DOI: 10.2174/0929866521666140507160050
Advertisement

Related Journals




Webmaster Contact: urooj@benthamscience.org Copyright © 2014 Bentham Science