Directed evolution of efficient secretion in the SRP-dependent export of TolB

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Title Directed evolution of efficient secretion in the SRP-dependent export of TolB
Author Zalucki, Yaramah M.; Shafer, William M.; Jennings, Michael Paul
Journal Name Biochimica et Biophysica Acta-Biomembranes
Year Published 2011
Place of publication Netherlands
Publisher Elsevier BV
Abstract Signal sequence non-optimal codons have been shown to be important for the folding and efficient export of maltose binding protein (MBP), a SecB dependent protein. In this study, we analysed the importance of signal sequence non-optimal codons of TolB, a signal recognition particle (SRP) dependent exported protein. The protein production levels of wild type TolB (TolB-wt) and a mutant allele of TolB in which all signal sequence non-optimal codons were changed to a synonymous optimal codon (TolB-opt), revealed that TolB-opt production was 12-fold lower than TolB-wt. This difference could not be explained by changes in mRNA levels, or plasmid copy number, which was the same in both strains. A directed evolution genetic screen was used to select for mutants in the TolB-opt signal sequence that resulted in higher levels of TolB production. Analysis of the 46 independent TolB mutants that reverted to wild type levels of expression revealed that at least four signal sequence non-optimal codons were required. These results suggest that non-optimal codons may be required for the folding and efficient export of all proteins exported via the Sec system, regardless of whether they are dependent on SecB or SRP for delivery to the inner membrane.
Peer Reviewed Yes
Published Yes
Alternative URI
Volume 1808
Issue Number 10
Page from 2544
Page to 2550
ISSN 0005-2736
Date Accessioned 2012-03-05; 2012-03-07T05:38:20Z
Date Available 2012-03-07T05:38:20Z
Research Centre Institute for Glycomics
Faculty Faculty of Science, Environment, Engineering and Technology
Subject Medical Microbiology
Publication Type Journal Articles (Refereed Article)
Publication Type Code c1

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