Skip Navigation


PEDS Advance Access originally published online on March 15, 2005
Protein Engineering Design and Selection 2005 18(1):51-58; doi:10.1093/protein/gzi005
This Article
Right arrow Full Text Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow All Versions of this Article:
18/1/51    most recent
gzi005v1
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 (6)
Right arrowRequest Permissions
Google Scholar
Right arrow Articles by Roodveldt, C.
Right arrow Articles by Tawfik, D.S.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Roodveldt, C.
Right arrow Articles by Tawfik, D.S.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

© The Author 2005. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions{at}oupjournals.org

Directed evolution of phosphotriesterase from Pseudomonas diminuta for heterologous expression in Escherichia coli results in stabilization of the metal-free state

C. Roodveldt and D.S. Tawfik1

Department of Biological Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel

1 To whom correspondence should be addressed. E-mail: tawfik{at}weizmann.ac.il

Phosphotriesterase from Pseudomonas diminuta (PTE) is an extremely efficient metalloenzyme that hydrolyses a variety of compounds including organophosphorus nerve agents. Study of PTE has been hampered by difficulties with efficient expression of the recombinant form of this highly interesting and potentially useful enzyme. We identified a low-level esterolytic activity of PTE and then screened PTE gene libraries for improvements in 2-naphthyl acetate hydrolysis. However, the attempt to evolve this promiscuous esterase activity led to a variant (S5) containing three point mutations that resulted in a 20-fold increase in functional expression. Interestingly, the zinc holoenzyme form of S5 appears to be more sensitive than wild-type PTE to both thermal denaturation and addition of metal chelators. Higher functional expression of the S5 variant seems to lie in a higher stability of the metal-free apoenzyme. The results obtained in this work point out another—and often overlooked—possible determinant of protein expression and purification yields, i.e. the stability of intermediates during protein folding and processing.

Received January 27, 2005; accepted February 1, 2005.

Edited by Frances Arnold


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
T.E. Reeves, M.E. Wales, J.K. Grimsley, P. Li, D.M. Cerasoli, and J.R. Wild
Balancing the stability and the catalytic specificities of OP hydrolases with enhanced V-agent activities
Protein Eng. Des. Sel., June 1, 2008; 21(6): 405 - 412.
[Abstract] [Full Text] [PDF]

Home page
 Protein Eng Des SelHome page
C. Mee-Hie Cho, A. Mulchandani, and W. Chen
Functional analysis of organophosphorus hydrolase variants with high degradation activity towards organophosphate pesticides
Protein Eng. Des. Sel., March 1, 2006; 19(3): 99 - 105.
[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.