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Antimicrobial Agents and Chemotherapy, May 1999, p. 1215-1224, Vol. 43, No. 5
Department of Biological Sciences, University
of Alberta, Edmonton, Alberta, Canada T6G 2E9,1
and SmithKline Beecham Pharmaceuticals, Worthing, West
Sussex BN14 8QH, United Kingdom2
Received 4 October 1998/Returned for modification 11 January
1999/Accepted 20 February 1999
Portions of the Streptomyces clavuligerus chromosome
flanking cas1, which encodes the clavaminate synthase 1 isoenzyme (CAS1), have been cloned and sequenced. Mutants of S. clavuligerus disrupted in cvm1, the open reading
frame located immediately upstream of cas1, were
constructed by a gene replacement procedure. Similar techniques were
used to generate S. clavuligerus mutants carrying a
deletion that encompassed portions of the two open reading frames, cvm4 and cvm5, located directly downstream of
cas1. Both classes of mutants still produced clavulanic
acid and cephamycin C but lost the ability to synthesize the antipodal
clavam metabolites clavam-2-carboxylate, 2-hydroxymethyl-clavam, and
2-alanylclavam. These results suggested that cas1 is
clustered with genes essential and specific for clavam metabolite
biosynthesis. When a cas1 mutant of S. clavuligerus was constructed by gene replacement, it produced lower levels of both clavulanic acid and most of the antipodal clavams
except for 2-alanylclavam. However, a double mutant of S. clavuligerus disrupted in both cas1 and
cas2 produced neither clavulanic acid nor any of the
antipodal clavams, including 2-alanylclavam. This outcome was
consistent with the contribution of both CAS1 and CAS2 to a common pool
of clavaminic acid that is shunted toward clavulanic acid and clavam
metabolite biosynthesis.
0066-4804/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Genes Specific for the Biosynthesis of Clavam Metabolites
Antipodal to Clavulanic Acid Are Clustered with the Gene for
Clavaminate Synthase 1 in Streptomyces
clavuligerus


*
Corresponding author. Mailing address: Department of
Biological Sciences, CW405 Biological Sciences Building, University of Alberta, Edmonton, Alberta, Canada T6G 2E9. Phone: (780) 492-0672. Fax:
(780) 492-2216. E-mail: susan.jensen{at}ualberta.ca.
Present address: Biology Program, University of Illinois at
Springfield, Springfield, IL 62794-9243.
Present address: Diversa Corporation, San Diego, CA 92121.
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