Renee Brost

Cubist Pharmaceuticals, Lexington, Massachusetts, United States

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Publications (2)5 Total impact

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    ABSTRACT: Ca(2+)-dependent cyclic lipodepsipeptides are an emerging class of antibiotics for the treatment of infections caused by Gram-positive pathogens. These compounds are synthesized by nonribosomal peptide synthetase (NRPS) complexes encoded by large gene clusters. The gene cluster encoding biosynthetic pathway enzymes for the Streptomyces fradiae A54145 NRP was cloned from a cosmid library and characterized. Four NRPS-encoding genes, responsible for subunits of the synthetase, as well as genes for accessory functions such as acylation, methylation and hydroxylation, were identified by sequence analysis in a 127 kb region of DNA that appears to be located subterminally in the bacterial chromosome. Deduced epimerase domain-encoding sequences within the NRPS genes indicated a D: -stereochemistry for Glu, Lys and Asn residues, as observed for positionally analogous residues in two related compounds, daptomycin, and the calcium-dependent antibiotic (CDA) produced by Streptomyces roseosporus and Streptomyces coelicolor, respectively. A comparison of the structure and the biosynthetic gene cluster of A54145 with those of the related peptides showed many similarities. This information may contribute to the design of experiments to address both fundamental and applied questions in lipopeptide biosynthesis, engineering and drug development.
    No preview · Article · Mar 2006 · Journal of Industrial Microbiology and Biotechnology
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    ABSTRACT: Daptomycin is a 13 amino acid, cyclic lipopeptide produced by a non-ribosomal peptide synthetase (NRPS) mechanism in Streptomyces roseosporus. A 128 kb region of S. roseosporus DNA was cloned and verified by heterologous expression in Streptomyces lividans to contain the daptomycin biosynthetic gene cluster (dpt). The cloned region was completely sequenced and three genes (dptA, dptBC, dptD) encoding the three subunits of an NRPS were identified. The catalytic domains in the subunits, predicted to couple five, six or two amino acids, respectively, included a novel activation domain and amino-acid-binding pocket for incorporating the unusual amino acid l-kynurenine (Kyn), three types of condensation domains and an extra epimerase domain (E-domain) in the second module. Novel genes (dptE, dptF) whose products likely work in conjunction with a unique condensation domain to acylate the first amino acid, as well as other genes (dptI, dptJ) probably involved in supply of the non-proteinogenic amino acids l-3-methylglutamic acid and Kyn, were located next to the NRPS genes. The unexpected E-domain suggested that daptomycin would have d-Asn, rather than l-Asn, as originally assigned, and this was confirmed by comparing stereospecific synthetic peptides and the natural product both chemically and microbiologically.
    Full-text · Article · Jun 2005 · Microbiology