PMID:12843673
| Citation |
He, M, Nie, YF and Xu, P (2003) A T42M substitution in bacterial 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) generates enzymes with increased resistance to glyphosate. Biosci. Biotechnol. Biochem. 67:1405-9 |
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| Abstract |
Mutants of class I enolpyruvylshikimate 3-phosphate synthase (EPSPS) with resistance to glyphosate were produced in a previous study using the staggered extension process with aroA genes from S. typhimurium and E. coli. Two of these mutants shared a common amino acid substitution, T42M, near the hinge region between the large globular domains of EPSPS. Using site-directed mutagenisis, we produced the T42M mutants without the other amino acid changes of the original mutants. The T42M substitution alone produced enzymes with a 9- to 25-fold decreased K(m)[PEP] and a 21- to 26-fold increased K(i)[glyphosate] compared to the wild-type enzymes. These results provide more testimony for the powerful approach for protein engineering by the combination of directed evolution and rational design. |
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| Keywords |
3-Phosphoshikimate 1-Carboxyvinyltransferase; Alkyl and Aryl Transferases/antagonists & inhibitors; Alkyl and Aryl Transferases/genetics; Alkyl and Aryl Transferases/metabolism; Amino Acid Substitution; Directed Molecular Evolution; Drug Resistance/genetics; Escherichia coli/enzymology; Glycine/analogs & derivatives; Glycine/pharmacology; Herbicides/pharmacology; Kinetics; Mutation; Plants, Genetically Modified/enzymology; Salmonella typhimurium/enzymology |
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