RESUMO
Phenalinolactones are novel terpene glycoside antibiotics produced by Streptomyces sp. Tü6071. Inactivation of three oxygenase genes (plaO2, plaO3 and plaO5), two dehydrogenase genes (plaU, plaZ) and one putative acetyltransferase gene (plaV) led to the production of novel phenalinolactone derivatives (PL HS6, PL HS7, PL HS2 and PL X1). Furthermore, the exact biosynthetic functions of two enzymes were determined, and their in vitro activities were demonstrated. PlaO1, an Fe(II)/alpha-ketoglutarate-dependent dioxygenase, is responsible for the key step in gamma-butyrolactone formation, whereas PlaO5, a cytochrome P450-dependent monooxygenase, catalyses the 1-C-hydroxylation of phenalinolactone D. In addition, stable isotope feeding experiments with biosynthetic precursors shed light on the origin of the carbons in the gamma-butyrolactone moiety.
Assuntos
Antibacterianos/biossíntese , Glicosídeos/biossíntese , Streptomyces/enzimologia , Terpenos/metabolismo , 4-Butirolactona/biossíntese , Acetiltransferases/genética , Acetiltransferases/metabolismo , Antibacterianos/química , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biocatálise , Sistema Enzimático do Citocromo P-450/metabolismo , Dioxigenases/metabolismo , Glicosídeos/química , Família Multigênica , Oxirredutases/genética , Oxirredutases/metabolismo , Oxigenases/genética , Oxigenases/metabolismo , Streptomyces/genética , Terpenos/químicaRESUMO
Polyketomycin is a tetracyclic quinone glycoside produced by Streptomyces diastatochromogenes Tü6028. It shows cytotoxic and antibiotic activity, in particular against Gram-positive multi-drug-resistant strains (for example, MRSA). The polyketomycin biosynthetic gene cluster has been sequenced and characterised. Its identity was proven by inactivation of a alpha-ketoacyl synthase gene (pokP1) of the "minimal polyketide synthase II" system. In order to obtain valuable information about tailoring steps, we performed further gene-inactivation experiments. The generation of mutants with deletions in oxygenase genes (pokO1, pokO2, both in parallel and pokO4) and methyltransferase genes (pokMT1, pokMT2 and pokMT3) resulted in new polyketomycin derivatives, and provided information about the organisation of the biosynthetic pathway.
Assuntos
Antibacterianos/biossíntese , Glicosídeos/biossíntese , Glioxilatos/metabolismo , Família Multigênica , Streptomyces/enzimologia , Streptomyces/genética , Clonagem Molecular , Deleção de Genes , Inativação Gênica , Genes Bacterianos , Teste de Complementação Genética , Biblioteca Genômica , Glicosídeos/química , Metiltransferases/genética , Oxigenases/genética , Policetídeo Sintases/genética , Policetídeo Sintases/metabolismo , Análise de Sequência de DNA , Streptomyces/metabolismo , Transcrição GênicaRESUMO
Isoprenoids produced by bacteria are of particular interest as they encompass huge structural and functional diversity. A rapidly increasing number of bacterial derived isoprenoids have been reported in recent years, as many of the genes and biosynthetic gene clusters are responsible for their biosynthesis. Polyprenyl chains, synthesized by the condensation of isopentenyl diphosphate units, serve as the substrates for cyclases and subsequent tailoring enzymes. It is these enzymes, particularly the cyclases, which are responsible for the structural diversity of this chemical class. Recent studies have revealed novel insights into isoprenoid biosynthesis, and in several cases enzymatic mechanisms have been redefined.