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Journal of Bacteriology, July 2000, p. 4087-4095, Vol. 182, No. 14
Department of Chemistry, The Johns Hopkins
University, Baltimore, Maryland 21218
Received 11 January 2000/Accepted 28 April 2000
Clavulanic acid is a potent inhibitor of
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Expansion of the Clavulanic Acid Gene Cluster: Identification
and In Vivo Functional Analysis of Three New Genes Required for
Biosynthesis of Clavulanic Acid by Streptomyces
clavuligerus
-lactamase enzymes and
is of demonstrated value in the treatment of infections by
-lactam-resistant bacteria. Previously, it was thought that eight contiguous genes within the genome of the producing strain
Streptomyces clavuligerus were sufficient for clavulanic
acid biosynthesis, because they allowed production of the antibiotic in
a heterologous host (K. A. Aidoo, A. S. Paradkar, D. C. Alexander, and S. E. Jensen, p. 219-236, In V. P. Gullo et al., ed., Development in industrial microbiology
series, 1993). In contrast, we report the identification of three
new genes, orf10 (cyp), orf11
(fd), and orf12, that are required for
clavulanic acid biosynthesis as indicated by gene replacement and
trans-complementation analysis in S. clavuligerus. These genes are contained within a 3.4-kb DNA
fragment located directly downstream of orf9
(cad) in the clavulanic acid cluster. While the
orf10 (cyp) and orf11
(fd) proteins show homologies to other known
CYP-150 cytochrome P-450 and [3Fe-4S] ferredoxin enzymes
and may be responsible for an oxidative reaction late in the pathway,
the protein encoded by orf12 shows no significant
similarity to any known protein. The results of this study extend the
biosynthetic gene cluster for clavulanic acid and attest to the
importance of analyzing biosynthetic genes in the context of their
natural host. Potential functional roles for these proteins are proposed.
*
Corresponding author. Mailing address: Department of
Chemistry, The Johns Hopkins University, 3400 North Charles St.,
Baltimore, MD 21218. Phone: (410) 516-7444. Fax: (410) 261-1233. E-mail: Townsend{at}jhunix.hcf.jhu.edu.
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