Identification of a cyclohexylcarbonyl CoA biosynthetic gene cluster and application in the production of doramectin

T. A. Cropp, D. J. Wilson, K. A. Reynolds

Research output: Contribution to journalArticlepeer-review

75 Scopus citations

Abstract

The side chain of the antifungal antibiotic ansatrienin A from Streptomyces collinus contains a cyclohexanecarboxylic acid (CHC)-derived moiety. This moiety is also observed in trace amounts of ω-cyclohexyl fatty acids (typically less than 1% of total fatty acids) produced by S. collinus. Coenzyme A-activated CHC (CHC-CoA) is derived from shikimic acid through a reductive pathway involving a minimum of nine catalytic steps. Five putative CHC-CoA biosynthetic genes in the ansatrienin biosynthetic gene cluster of S. collinus have been identified. Plasmid-based heterologous expression of these five genes in Streptomyces avermitilis or Streptomyces lividans allows for production of significant amounts of ω-cyclohexyl fatty acids (as high as 49% of total fatty acids). In the absence of the plasmid these organisms are dependent on exogenously supplied CHC for ω-cyclohexyl fatty acid production. Doramectin is a commercial antiparasitic avermectin analog produced by fermenting a bkd mutant of S. avermitilis in the presence of CHC. Introduction of the S. collinus CHC-CoA biosynthetic gene cassette into this organism resulted in an engineered strain able to produce doramectin without CHC supplementation. The CHC-CoA biosynthetic gene cluster represents an important genetic tool for precursor-directed biosynthesis of doramectin and has potential for directed biosynthesis in other important polyketide-producing organisms.

Original languageEnglish (US)
Pages (from-to)980-983
Number of pages4
JournalNature biotechnology
Volume18
Issue number9
DOIs
StatePublished - 2000
Externally publishedYes

Bibliographical note

Funding Information:
We thank Ron Fedechcko and Hamish McArthur at Pfizer, for providing doramectin and for carrying out LC-MS analyses of the S. avermitilis/pAC12 fermentations. We also thank Claudio Denoya (Pfizer) for advice for working with the bkd mutant of S. avermitilis. We thank Stephanie Patton for carrying out 2-cyclohexenylcarbonyl CoA isomerase assays. This work was supported in part by a grant from the National Science Foundation (MCB 9418581).

Copyright:
Copyright 2007 Elsevier B.V., All rights reserved.

Keywords

  • Avermectin
  • Biosynthesis
  • Polyketide
  • Streptomyces

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