Secretory signal sequence non-optimal codons are required for expression and export of beta-lactamase.
In this study we altered the codon usage in the signal sequence of the bla gene, encoding beta-lactamase in Escherichia coli. Changing all of the thirteen non-optimal codons to optimal lowered expression 4-fold as measured by minimum inhibitory concentration (MIC) to the beta-lactam antibiotic ampicillin. The difference in ampicillin resistance was reduced at 28 degrees C compared to expression at 37 degrees C, suggesting that the optimised bla allele is misfolded and degraded by heat-shock regulated proteases. A screen was carried out, designed specifically to identify revertants with changes in codon usage resulting in higher MIC to ampicillin. The nine revertants revealed by this method all had optimal to non-optimal codon changes in the signal sequence. These results, and those of our previous study with maltose binding protein model system, confirm that non-optimal codons are important for expression and export of secretory proteins via both the SecB-dependent and -independent pathways.
School of Molecular & Microbial Sciences, University of Queensland, St. Lucia, Qld 4072, Australia.
Yaramah M
YM
Zalucki
Karlee L
KL
Gittins
Michael P
MP
Jennings
D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D003062:Codon / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics*; D018832:Molecular Chaperones / Q000235:genetics*; D008969:Molecular Sequence Data; D021381:Protein Transport / Q000235:genetics*; D015398:Signal Transduction / Q000235:genetics*; D001618:beta-Lactamases / Q000235:genetics*
D001426:Bacterial Proteins; D003062:Codon; D018832:Molecular Chaperones; C057842:SecB protein, Bacteria; D001618:beta-Lactamases