Skip Navigation

This Article
Right arrow FREE Full Text (PDF) Freely available
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 ISI Web of Science
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 Search for citing articles in:
ISI Web of Science (7)
Right arrowRequest Permissions
Google Scholar
Right arrow Articles by Koga, Y.
Right arrow Articles by Kanaya, S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Koga, Y.
Right arrow Articles by Kanaya, S.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

Protein Engineering, Vol 11, 1219-1227, Copyright © 1998 by Oxford University Press

ARTICLES

Thermostable glycerol kinase from a hyperthermophilic archaeon: gene cloning and characterization of the recombinant enzyme

Y Koga, M Morikawa, M Haruki, H Nakamura, T Imanaka and S Kanaya
Department of Material and Life Science, Graduate School of Engineering, Osaka University, Suita, Japan.

The Pk-glpK gene, which encodes glycerol kinase (GK) from a hyperthermophilic archaeon Pyrococcus kodakaraensis KOD1, was cloned and expressed in Escherichia coli. The amino acid sequence of this enzyme (Pk-GK) deduced from the nucleotide sequence showed 57% identity with that of E. coli GK and 47% identity with that of human GK. Pk-GK, which has a molecular weight of 55902 (497 amino acid residues), was purified from E. coli and characterized. Despite the high sequence similarity, Pk-GK and E. coli GK are greatly divergent in structure and function from each other. Unlike E. coli GK, which exists as a tetramer, Pk-GK exists as a dimer. The preferred divalent cation for Pk- GK is Co2+, instead of Mg2+. The optimum pH and temperature for Pk-GK activity are 8.0 and 80 degrees C, respectively. Pk-GK can utilize other nucleoside triphosphates than ATP as a phosphoryl donor. It is fairly resistant to an allosteric inhibitor of E. coli GK, fructose-1,6- bisphosphate. Determination of the kinetic parameters indicates that the Km value of the enzyme is 15.4 microM for ATP and 111 microM for glycerol and its kcat value is 940 s(-1). The enzyme was shown to be fairly resistant to irreversible heat inactivation and still retained 50% of its enzymatic activity even after heating at 100 degrees C for 30 min. Construction of a model for the three-dimensional structure of the enzyme suggests that the formation of extensive ion-pair networks is responsible for the high stability of this enzyme.
Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?


This article has been cited by other articles:


Home page
 Protein Eng Des SelHome page
S.-i. Sakasegawa, H. Takehara, I. Yoshioka, M. Takahashi, Y. Kagimoto, H. Misaki, H. Sakuraba, and T. Ohshima
Increasing the thermostability of Flavobacterium meningosepticum glycerol kinase by changing Ser329 to Asp in the subunit interface region
Protein Eng. Des. Sel., September 1, 2001; 14(9): 663 - 667.
[Abstract] [Full Text] [PDF]


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.