Abstract：
:The site-directed mutagenesis of a number of proposed active site residues of 5-enolpyruvyl shikimate-3-phosphate (EPSP) synthase is reported. Several of these mutations resulted in complete loss of enzyme activity indicating that these residues are probably involved with catalysis, notably K22R, K411R, D384A, R27A, R100A, and D242A. Of those, K22R, R27A, and D384A did not bind either the substrate shikimate-3-phosphate (S3P) or glyphosate (GLP). The K411R and D242A mutants bind S3P only in the presence of GLP. The kinetic characterization of mutants R100K, K340R, and E418A, which retain activity, is reported. Of those, R100K and K340R do not accumulate enzyme intermediate of enzyme-bound product under equilibrium conditions. These residues, while not essential for catalysis, are most likely important for substrate binding. All of the mutants are shown to be correctly folded by NMR spectroscopy.
journal_name
Biochemistryjournal_title
Biochemistryauthors
Shuttleworth WA,Pohl ME,Helms GL,Jakeman DL,Evans JNdoi
10.1021/bi9815142subject
Has Abstractpub_date
1999-01-05 00:00:00pages
296-302issue
1eissn
0006-2960issn
1520-4995pii
bi9815142journal_volume
38pub_type
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