Skip Navigation

Nucleic Acids Research 2004 32(17):e136; doi:10.1093/nar/gnh135
This Article
Right arrow Full Text Freely available
Right arrow Print PDF (131K) Freely available
Right arrow Supplementary Material
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 arrowRequest Permissions
Right arrow Commercial Re-use Guidelines
for Open Access NAR Content
Google Scholar
Right arrow Articles by Osuna, J.
Right arrow Articles by Gaytán, P.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Osuna, J.
Right arrow Articles by Gaytán, P.
Related Collections
Right arrow Mutagenesis
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

Published online 30 September 2004

Nucleic Acids Research, Vol. 32 No. 17 © Oxford University Press 2004; all rights reserved

Protein evolution by codon-based random deletions

Joel Osuna, Jorge Yáñez, Xavier Soberón and Paul Gaytán*

Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología/UNAM, Ap. Postal 510-3 Cuernavaca, Morelos 62250, México

* To whom correspondence should be addressed. Tel: +52 777 329 1604; Fax: +52 777 317 2388; Email: paul{at}ibt.unam.mx

Received August 13, 2004; Revised September 10, 2004; Accepted September 15, 2004

A method to delete in-phase codons throughout a defined target region of a gene has been developed. This approach, named the codon-based random deletion (COBARDE) method, is able to delete complete codons in a random and combinatorial mode. Robustness, automation and fine-tuning of the mutagenesis rate are essential characteristics of the method, which is based on the assembly of oligonucleotides and on the use of two transient orthogonal protecting groups during the chemical synthesis. The performance of the method for protein function evolution was demonstrated by changing the substrate specificity of TEM-1 ß-lactamase. Functional ceftazidime-resistant ß-lactamase variants containing several deleted residues inside the catalytically important omega-loop region were found. The results show that the COBARDE method is a useful new molecular tool to access previously unexplorable sequence space.


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
 J. Biol. Chem.Home page
C. Urbach, J. Fastrez, and P. Soumillion
A New Family of Cyanobacterial Penicillin-binding Proteins: A MISSING LINK IN THE EVOLUTION OF CLASS A {beta}-LACTAMASES
J. Biol. Chem., November 21, 2008; 283(47): 32516 - 32526.
[Abstract] [Full Text] [PDF]

Home page
 Protein Eng Des SelHome page
S. Emond, P. Mondon, S. Pizzut-Serin, L. Douchy, F. Crozet, K. Bouayadi, H. Kharrat, G. Potocki-Veronese, P. Monsan, and M. Remaud-Simeon
A novel random mutagenesis approach using human mutagenic DNA polymerases to generate enzyme variant libraries
Protein Eng. Des. Sel., April 1, 2008; 21(4): 267 - 274.
[Abstract] [Full Text] [PDF]

Home page
 Proc. Natl. Acad. Sci. USAHome page
G. Guntas, T. J. Mansell, J. R. Kim, and M. Ostermeier
Directed evolution of protein switches and their application to the creation of ligand-binding proteins
PNAS, August 9, 2005; 102(32): 11224 - 11229.
[Abstract] [Full Text] [PDF]

Home page
 Nucleic Acids ResHome page
J. Yanez, M. Arguello, J. Osuna, X. Soberon, and P. Gaytan
Combinatorial codon-based amino acid substitutions
Nucleic Acids Res., November 10, 2004; 32(20): e158 - e158.
[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.