The polyketide epothilone is a potential anticancer agent that stabilizes microtubules in a similar manner to Taxol. The gene cluster responsible for epothilone biosynthesis in the myxobacterium Sorangium cellulosum was cloned and completely sequenced. It encodes six multifunctional proteins composed of a loading module, one nonribosomal peptide synthetase module, eight polyketide synthase modules, and a P450 epoxidase that converts desoxyepothilone into epothilone. Concomitant expression of these genes in the actinomycete Streptomyces coelicolor produced epothilones A and B. Streptomyces coelicolor is more amenable to strain improvement and grows about 10-fold as rapidly as the natural producer, so this heterologous expression system portends a plentiful supply of this important agent.

HAL is a multi-disciplinary open access archive for the deposit and dissemination of scientific research documents, whether they are published or not. The documents may come from teaching and research institutions in France or abroad, or from public or private research centers. L'archive ouverte pluridisciplinaire HAL, est destinée au dépôt et à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

No abstract

Patients exposed to a surgical safety checklist experience better postoperative outcomes, but this could simply reflect wider quality of care in hospitals where checklist use is routine.

Epothilones are potential anticancer drugs that stabilize microtubules in a manner similar to paclitaxel (Taxol). Epothilones are produced from the myxobacterium Sorangium cellulosum, which has a 16-h doubling time and produces only milligram-per-liter amounts of epothilone A and epothilone B. Furthermore, genetic manipulation of S. cellulosum is difficult. To produce epothilones in a more genetically amenable and rapidly growing host, we chose the closely related and best-characterized myxobacteria Myxococcus xanthus. We inserted 65.4 kb of S. cellulosum DNA that encompassed the entire epothilone gene cluster into the chromosome of M. xanthus by a series of homologous recombination events. The resulting strain produced epothilones A and B. Construction of a strain that contained a mutation in epoK, the P450 epoxidase, resulted in production of epothilones C and D.Epothilones are potent cytotoxic macrocyclic lactones that show promise as anticancer drugs (21,22,26). The mechanism of action is similar to the anticancer drug paclitaxel (Taxol); both bind and stabilize microtubules, which leads to cell death (3). Epothilones have superior features relative to paclitaxel. First, epothilones are more water soluble. This may enable a formulation without the use of the solubilizing agent cremophor, currently used in formulations of paclitaxel. Cremophor, on its own, can affect cardiac function and cause severe hypersensitivity (25). Second, epothilones are effective against tumors resistant to paclitaxel (3, 31). These advantages make epothilones likely successors to paclitaxel.The need for sufficient material is a major obstacle to the development of epothilones as marketable drugs. The total synthesis of epothilone A and epothilone B has been accomplished (2, 32). However, the number of steps required for synthesis of these molecules precludes this as an economical method of production. Fermentation methods thus remain the favored route of production.Epothilones are produced from the gram-negative myxobacterium Sorangium cellulosum (8). The reported yields of epothilones from S. cellulosum strain So ce90 are approximately 20 mg of epothilone A per liter and 10 mg of epothilone B per liter (8). A disadvantage of S. cellulosum is the relatively long doubling time, which is approximately 16 h and is the longest of all myxobacteria. Furthermore, S. cellulosum is difficult to engineer, due to the low efficiency of introducing DNA into the bacterium (11) and the limited number of molecular tools and markers that have been developed.Recently, the epothilone biosynthetic gene cluster was sequenced, and the genes were introduced into Streptomyces coelicolor, a common host used for production of a variety of polyketides from actinomycetes (33). The heterologous strain produced small quantities of epothilones; production of epothilones in S. coelicolor may have a cytotoxic effect (L. Tang et al., unpublished data). Thus, an alternative heterologous host is desired.We speculated that a superior expression host for epothilones mi...