Open AccessChemobiosynthesis of Novel 6-Deoxyerythronolide B Analogues by Mutation of the Loading Module of 6-Deoxyerythronolide B Synthase 1
Author(s) -
Sumati Murli,
Karen S. MacMillan,
Zhihao Hu,
Gary W. Ashley,
Steven D. Dong,
James T. Kealey,
Christopher D. Reeves,
Jonathan Kennedy
Publication year - 2005
Publication title -
applied and environmental microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.552
H-Index - 324
eISSN - 1070-6291
pISSN - 0099-2240
DOI - 10.1128/aem.71.8.4503-4509.2005
Subject(s) - thioester , streptomyces coelicolor , stereochemistry , polyketide , polyketide synthase , escherichia coli , atp synthase , biosynthesis , chemistry , biochemistry , actinomycetales , depsipeptide , streptomyces , biology , mutant , bacteria , enzyme , gene , genetics
Chemobiosynthesis (J. R. Jacobsen, C. R. Hutchinson, D. E. Cane, and C. Khosla, Science 277:367-369, 1997) is an important route for the production of polyketide analogues and has been used extensively for the production of analogues of 6-deoxyerythronolide B (6-dEB). Here we describe a new route for chemobiosynthesis using a version of 6-deoxyerythronolide B synthase (DEBS) that lacks the loading module. When the engineered DEBS was expressed in both Escherichia coli and Streptomyces coelicolor and fed a variety of acyl-thioesters, several novel 15-R-6-dEB analogues were produced. The simpler "monoketide" acyl-thioester substrates required for this route of 15-R-6-dEB chemobiosynthesis allow greater flexibility and provide a cost-effective alternative to diketide-thioester feeding to DEBS KS1(o) for the production of 15-R-6-dEB analogues. Moreover, the facile synthesis of the monoketide acyl-thioesters allowed investigation of alternative thioester carriers. Several alternatives to N-acetyl cysteamine were found to work efficiently, and one of these, methyl thioglycolate, was verified as a productive thioester carrier for mono- and diketide feeding in both E. coli and S. coelicolor.