Regulation of the biosynthesis of the glycopeptide antibiotic A40926 in Nonomuraea ATCC39727

Abstract

The actinomycete Nonomuraea produces the glycopeptide A40926, precursor of dalbavancin. Dalbavancin is a novel lipoglycopeptide agent against emerging resistant Gram-positive cocci with superior pharmacodynamics properties compared to vancomycin. Chemically, a glycopeptide antibiotic consists of a heptapeptide core constituted by proteinogenic and nonproteinogenic amino acids such as 3,5-dihydroxyphenylglycine (DPG) and 4-hydroxyphenylglycine (HPG). The heptapeptide is assembled by a nonribosomal peptide synthetase and modified by oxidative cross-linking of the electron-rich aromatic side chains, halogenation, sulfation, methylation, acylation, and glycosylation. The A40926 biosynthetic gene (dbv) cluster, which consists of 37 genes, encodes two putative regulators, Dbv3 and Dbv4, as well as the response regulator (Dbv6) and the sensor-kinase (Dbv22) of a putative two-component system. The purpose of this study is to investigate the role of these regulators in A40926 biosynthesis. Mutants in each regulator were generated. Bioassay and HPLC analysis revealed that Dbv3 and Dbv4 are necessary for antibiotic production. Dbv4, a StrR-like protein, was found to positively regulate the transcription of 13 genes, encoding DPG synthesizing enzymes and modification enzymes. Dbv3 is a putative LuxR-like regulator and the search for Dbv3-targeted genes is ongoing by transcriptional and biochemical analysis. Surprisingly, the response regulator Dbv6 does not seem to influence A40926 production. Chemical and biochemical analysis are in progress to elucidate its role. These studies will provide new insights in gene expression of gram-positive bacteria, in particular they will elucidate the complex regulatory network governing antibiotic production. Moreover, these analyses will offer a platform for the rational manipulation of Nonomuraea for increased production of A40926.