Isolation and mapping of a mutant penicillin G acylase with altered substrate specificity from Escherichia coli |
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Authors: | Helmut Niersbach Walter Tischer Marijke Weber Frank Wedekind Roland Plapp |
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Affiliation: | (1) Department of Microbiology, University of Kaiserslautern, PO box 3049, D-67663 Kaiserslautern;(2) Boehringer Mannheim GmbH, Nonnenwald 2, D-82377 Penzberg;(3) Present address: Department of Microbiology, NYU Medical School, 550 First Avenue, 10016 New York, New York |
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Abstract: | Summary Penicillin G acylase of Escherichia coli ATCC 11105 catalyzes hydrolysis as wellas synthesis of penicillin G. In this work a recombinant penicillin G acylase genewas mutagenized in vivo. A mutant with altered penicillin G acylase was selectedby its ability to grow with phthalyl-L-leucine as sole source of leucine. Themutant enzyme obtained was deficient in hydrolyzing penicillin G. A mutation ofGly359 to aspartic acid was mapped first by construction of chimeric pac genescomposed of wild type and mutant DNA, followed by nucleotide sequencing. |
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