Identification of a cyclohexylcarbonyl CoA biosynthetic gene cluster and application in the production of doramectin.
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 omega-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 omega-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 omega-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.
Cropp
T A
TA
Department of Medicinal Chemistry and Institute for Structural Biology and Drug Discovery, Virginia Commonwealth University, 800 E. Leigh St. Suite 212, Richmond, VA 23219, USA.
Wilson
D J
DJ
Reynolds
K A
KA
D000871:Anthelmintics / Q000138:chemical synthesis*; D001709:Biotechnology / Q000379:methods; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D003065:Coenzyme A / Q000737:chemistry* / Q000235:genetics*; D003509:Cyclohexanecarboxylic Acids / Q000737:chemistry*; D005227:Fatty Acids / Q000737:chemistry; D008401:Gas Chromatography-Mass Spectrometry; D007559:Ivermectin / Q000031:analogs & derivatives* / Q000138:chemical synthesis; D008956:Models, Chemical; D008957:Models, Genetic; D005810:Multigene Family; D010957:Plasmids / Q000378:metabolism; D012765:Shikimic Acid / Q000031:analogs & derivatives; D013302:Streptomyces / Q000737:chemistry / Q000235:genetics
D000871:Anthelmintics; D003509:Cyclohexanecarboxylic Acids; D005227:Fatty Acids; D012765:Shikimic Acid; D007559:Ivermectin; C084101:doramectin; D003065:Coenzyme A