Expression of the Platencin Biosynthetic Gene Cluster in Heterologous Hosts Yielding New Platencin Congeners

Abstract

Platensimycin (PTM) and platencin (PTN) are potent and selective inhibitors of bacterial and mammalian fatty acid synthases and have emerged as promising drug leads for both antibacterial and antidiabetic therapies. We have previously cloned and sequenced the PTM-PTN dual biosynthetic gene cluster from S. platensis MA7327 and the PTN biosynthetic gene cluster from S. platensis MA7339, the latter of which is composed of 31 genes encoding PTN biosynthesis, regulation, and resistance. We have also demonstrated that PTM or PTN production can be significantly improved upon inactivation of the pathway specific regulator ptmR1 or ptnR1, in S. platensis MA7327 or MA7339, respectively. We now report engineered production of PTN and congeners in a heterologous Streptomyces host. Expression constructs containing the ptn biosynthetic gene cluster were engineered from SuperCos 1 library clones and introduced into five model Streptomyces hosts, and PTN production was achieved in Streptomyces lividans K4-114. Inactivation of ptnR1 was crucial for expression of the ptn biosynthetic gene cluster, thereby PTN production, in S. lividans K4-114. Six PTN congeners, five of which were new, were also isolated from the recombinant strain S. lividans SB12606, revealing new insights into PTN biosynthesis. Production of PTN in a model Streptomyces host provides new opportunities to apply combinatorial biosynthetic strategies to the PTN biosynthetic machinery for structural diversity.

A genetically amenable platensimycin- and platencin-overproducer as a platform for biosynthetic explorations: a showcase of PtmO4, a long-chain acyl-CoA dehydrogenase

“…Its 1 H and 13 C NMR data was identical with that of (4 S )-homoplatencinic acid, also called platencin SL5, a known intermediate in the heterologous biosynthesis of PTN in S. lividans SB12606 and SB12608. 15 Similar to 3 , the stereochemistry at C4 of platencin SL5 ( 11 ) is in the 4 S configuration. We isolated three additional PTN analogues including one known and two new compounds.…”

“…Following previously reported procedures, 8, 12, 13, 15 extraction of a 3.6 L culture with Amberlite XAD-16 resin and subsequent column chromatography resulted in the isolation of 14 compounds (Fig.1). Of these 14 compounds, ten are new and four were previously isolated from engineered PTN-producing strains ( 11 and 14 ) or plants ( 15 and 16 ).…”

“…Thus, the structure of 3 was fully elucidated. For consistency with PTN congeners, 15 3 was also named (4 S )-homoplatensic acid.…”

“…Their molecular weights were 16 Da more than 11 and likely due to one additional hydroxyl group. For 12 , the hydroxyl group was attached to C15, which caused downfield shifts of 5.1 and 4.4 ppm for the adjacent carbons C16 and C8, 15 respectively, and was further supported by key HMBC correlations of H15 ( δ 4.00, s) with C7 ( δ 153.6), C9 ( δ 38.2), C12 ( δ 36.6), C14 ( δ 20.7) and C17 ( δ 111.2). For 13 , the downfield shifts for C13 and C8 were 12.5 and 5.5 ppm, 15 respectively, indicating the hydroxyl group was attached at C14.…”

“…13, 14 Heterologous expression of the ptn cluster in Streptomyces lividans K4-114 also produced new PTN congeners as well as PTN, albeit at titers < 5 mg L −1 . 15 …”