HETEROLOGOUS PRODUCTION OF DAPTOMYCIN IN STREPTOMYCES LIVIDANS

Authors: PENN, JULIA - CUBIST PHARMACEUTICALS; LI, XIAG - CUBIST PHARMACEUTICALS; WHITING, ANDREW - CUBIST PHARMACEUTICALS; LATIF, MOHAMMED - CUBIST PHARMACEUTICALS; Silva, Christopher; BRIAN, PAUL - CUBIST PHARMACEUTICALS; DAVIES, JULIAN - CUBIST PHARMACEUTICALS; MIAO, VIVIAN - CUBIST PHARMACEUTICALS; WRIGLEY, STEPHEN - CUBIST PHARMACEUTICALS; BALTZ, RICHARD - CUBIST PHARMACEUTICALS

Submitted to: Journal of Industrial Microbiology and Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/4/2005
Publication Date: 11/1/2005
Citation: Penn, J., Li, X., Whiting, A., Latif, M., Silva, C.J., Brian, P., Davies, J., Miao, V., Wrigley, S.K., Baltz, R.H. 2005. Heterologous production of daptomycin in streptomyces lividans [Epub ahead of print]. Journal of Industrial Microbiology and Biotechnology. Nov 1:1-8.

Interpretive Summary: Daptomycin and related compounds are lipopeptide antibiotics. They are naturally produced by the bacterium Streptomyces roseosporus. A related bacterium, Streptomyces lividans, can be made to produce daptomycin when a large DNA (128 kbp) fragment containing the daptomycin gene cluster is cloned into a specific site in its genome (phiC31 att site). The fermentation yields of daptomycin and related compounds were initially much lower in S. lividans than in S. roseosporus, and detection was complicated by the production of host metabolites. The production in S. lividans was improved by deletion of genes encoding the production of a major host metabolite and by medium optimization. Adding phosphate to a defined fermentation medium resulted in formation of host metabolites, which were well separated from daptomycin on chromatographic analysis. Adjusting the level of phosphate in the medium led to an improvement in daptomycin yield from 20 to 55 mg/l.

Technical Abstract: Daptomycin and the A21978C antibiotic complex are lipopeptides produced by Streptomyces roseosporus and also in recombinant Streptomyces lividans TK23 and TK64 strains, when a 128 kbp region of cloned S. roseosporus DNA containing the daptomycin gene cluster is inserted site-specifically in the varphiC31 attB site. A21978C fermentation yields were initially much lower in S. lividans than in S. roseosporus, and detection was complicated by the production of host metabolites. However A21978C production in S. lividans was improved by deletion of genes encoding the production of actinorhodin and by medium optimization to control the chemical form of the calcium dependent antibiotic (CDA). This latter compound has not previously been chemically characterized as a S. lividans product. Adding phosphate to a defined fermentation medium resulted in formation of only the phosphorylated forms of CDA, which were well separated from A21978C on chromatographic analysis. Adjusting the level of phosphate in the medium led to an improvement in A21978C yield from 20 to 55 mg/l.