Methylbenzene-Containing Polyketides from a Streptomyces that Spontaneously Acquired Rifampicin Resistance: Structural Elucidation and Biosynthesis

Abstract

Conventional screening for novel bioactive compounds in actinomycetes often results in the rediscovery of known compounds. In contrast, recent genome sequencing revealed that most of the predicted gene clusters for secondary metabolisms are not expressed under standard cultivation conditions. To explore the potential metabolites produced by these gene clusters, we implemented a cryptic gene activation strategy by screening mutants that acquire resistance to rifampicin. The induction of rifampicin resistance in 11 actinomycete strains generated 164 rifampicin-resistant mutants (rif mutants). The comparison of the metabolic profiles between the rif mutants and their wild-type strains indicated that one mutant (TW-R50-13) overproduced an unidentified metabolite (1). During the isolation and structural elucidation of metabolite 1, an additional metabolite was found; both are unprecedented compounds featuring a C5N unit and a methylbenzene moiety. Of these partial structures, the biosynthesis of the latter has not been reported. A feeding experiment using (13)C-labeled precursors demonstrated that the methylbenzene moiety is most likely synthesized by the action of polyketide synthase. The gene deletion experiments revealed that the genes for the methylbenzene moiety are located at a different locus than the genes for the C5N unit.

New Antimicrobial Phenyl Alkenoic Acids Isolated from an Oil Palm Rhizosphere-Associated Actinomycete, Streptomyces palmae CMU-AB204T

“…Previously, Thong et al [35] found two closely related compounds of 1-4, and they were isolated from a Streptomyces that had been spontaneously acquired rifampicin resistance. These compounds contained E-olefins and have both a methylbenzene unit and a 2-amino-3-hydroxycyclopent-2-enone (C 5 N) moiety.…”

“…Notably, the presence of a carboxylic acid moiety in the novel compounds, and a C 5 N moiety in the known compounds, may not be involved in the antimicrobial activity. Based on draft genome sequences of the rifampicin-resistant mutant (TW-R50-13), the methylbenzene moiety may be biosynthesized by the expression of polyketide synthase (PKS) genes that are located at a different locus from the biosynthetic genes for the C 5 N moiety [35]. The genes encoding for PKS have been disclosed to complex biosynthetic mechanisms, which were involved in the production of many metabolites in microorganisms [46].…”

“…The geometry of two substitutes of the aromatic ring was confirmed by NOE correlations, as is shown in Figure 2. From some Streptomyces strains, E-isomer of 1, (E)-5-(2-methylphenyl)-4-pentenoic acid was identified [33][34][35]; however, there was no report on the Z-isomer (1) obtained from natural sources. Therefore, it was concluded that 1 was a novel natural product.…”

Antifungal Streptomyces spp., Plausible Partners for Brood-Caring of the Dung Beetle Copris tripartitus

“…The phylogenetic tree was constructed using the Maximum Likelihood (ML) method applied to pairwise sequence distances calculated by the general time reversible (GTR) model with the MEGA program [ 31 , 32 ]. In addition, partial rpoB gene of antifungal isolates was obtained by PCR amplification using a primer set for rpoB -P2 [ 33 ] and purification, and sequenced. Additional phylogenetic tree of the concatenated sequences of 16S rRNA and rpoB genes was constructed using the ML method and GTR model.…”

Activation and Characterization of Lanthomicins A–C by Promoter Engineering in Streptomyces chattanoogensis L10

“…In the light of huge sequence data in the genomic era, several main strategies have been developed in an attempt to harness this tremendous chemical diversity. These include promoter engineering (Li et al, 2017 ), diversification of culture conditions (Zhang et al, 2014 ), ribosome engineering (Wang et al, 2008 ; Thong et al, 2016 ), manipulation of regulators (Chen et al, 2017 ), and others. Among them, promoter engineering has been proved as a powerful and efficient method to activate or improve the biosynthesis of interest products (Li et al, 2015 , 2017 ).…”