Skip Navigation


Journal of Biochemistry Advance Access originally published online on March 27, 2007
Journal of Biochemistry 2007 141(6):791-797; doi:10.1093/jb/mvm082
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
141/6/791    most recent
mvm082v1
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 Add to My Personal Archive
Right arrow Download to citation manager
Right arrow Request Permissions
Google Scholar
Right arrow Articles by Ohkuri, T.
Right arrow Articles by Yamagishi, A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Ohkuri, T.
Right arrow Articles by Yamagishi, A.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

© 2007 The Japanese Biochemical Society.

The Effects of Mutations at Position 253 on the Thermostability of the Bacillus subtilis 3-Isopropylmalate Dehydrogenase Subunit Interface

Takatoshi Ohkuri1,2 and Akihiko Yamagishi1,*

1Department of Molecular Biology, Tokyo University of Pharmacy and Life Science, 1432-1 Horinouchi, Hachioji, Tokyo 192-0392; and 2Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka 812-8582, Japan

*To whom correspondence should be addressed. Tel: 81-426-76-7139, Fax: 84-426-76-7145, E-mail: yamagish{at}ls.toyaku.ac.jp

Received December 13, 2006; Accepted March 14, 2007


  Abstract

3-Isopropylmalate dehydrogenase (IPMDH) is a dimeric enzyme with a strongly hydrophobic core that is composed of residues from four {alpha}-helices. We replaced Glu253, which is found in the hydrophobic core and is part of the subunit interface of the Bacillus subtilis (Bs) IPMDH, with several other amino acids to probe. The thermostabilities of the mutants were assessed by measuring the residual enzymatic activities at 40°C after heat treatment and by monitoring changes in ellipticity at 222 nm as the environmental temperature increased incrementally. The results of these studies indicate that, for residues with non-polar side chains, when positioned at residue 253, the thermostabilities of their corresponding mutants correlate positively with the relative hydrophobicities of the side chains. Relative activities of all mutants are lower than that of the wild-type enzyme. For two of the mutants, we directly show that the substitution at position 253 negatively affects Mn2+ binding, which is required for catalysis. When a lysine is the position 253 residue, the protein dissociates. The results presented herein increase our understanding of the role played by the BsIPMDH dimer interface on the stability and activity of BsIPMDH.

Key Words: dimeric enzyme, hydrophobic core, isopropylmalate dehydrogenase, subunit interaction, thermostability

Abbreviations: BsIPMDH, Bacillus subtilis IPMDH; EcIPMDH, E. coli IPMDH; IPM, d-l-3-isopropylmalate; IPMDH, 3-isopropylmalate dehydrogenase; TtIPMDH, Thermus thermophilus IPMDH


Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?




Disclaimer:
Please note that abstracts for content published before 1996 were created through digital scanning and may therefore not exactly replicate the text of the original print issues. All efforts have been made to ensure accuracy, but the Publisher will not be held responsible for any remaining inaccuracies. If you require any further clarification, please contact our Customer Services Department.