Abstract
Mycobacterial pathogenesis is hallmarked by lipidic polyketides that decorate the cell envelope and mediate infection. However, factors mediating persistence remain largely unknown. Dynamic cell wall remodeling could facilitate the different pathogenic phases. Recent studies have implicated type III polyketide synthases (PKSs) in cell wall alterations in several bacteria. Comparative genome analysis revealed several type III pks gene clusters in mycobacteria. In this study, we report the functional characterization of two novel type III PKSs, MMAR_2470 and MMAR_2474, in Mycobacterium marinum. These type III pkss belong to a unique pks genomic cluster conserved exclusively in pathogenic mycobacteria. Cell-free reconstitution assays and high-resolution mass spectrometric analyses revealed methylated polyketide products in independent reactions of both proteins. MMAR_2474 protein exceptionally biosynthesized methylated alkyl-resorcinol and methylated acyl-phloroglucinol products from the same catalytic core. Structure-based homology modeling, product docking, and mutational studies identified residues that could facilitate the distinctive catalysis of these proteins. Functional investigations in heterologous mycobacterial strain implicated MMAR_2474 protein to be vital for mycobacterial survival in stationary biofilms. Our investigations provide new insights into type III PKSs conserved in pathogenic mycobacterial species.
Highlights
Comparative genome analyses has revealed conservation of several biosynthetic pks genes across various mycobacterial species[11,12]
We investigated the occurrence of type III pks genes in Mycobacterium marinum (Mmar)
Several studies have provided understanding on polyketide synthases (PKSs) mediated mechanisms employed by pathogenic mycobacterial species to armor against the host defenses and establish virulent pathology
Summary
Comparative genome analyses has revealed conservation of several biosynthetic pks genes across various mycobacterial species[11,12]. Methylated polyketide quinones from Mycobacterium smegmatis (Msmeg) have been shown to facilitate anaerobic respiration of Msmeg cells in stationary biofilms[36] These molecules are produced by a genomic cluster of Msmeg homologous to the srs operon. Homology-based structural modeling, molecular docking and site directed mutagenesis identified catalytically crucial active site residue positions and revealed clues to the structure-function relationship in Mmar type III PKSs. High-resolution metabolomic investigations of heterologous complemented type III pks knockout mycobacterial strain revealed the physiological importance of methylated polyketides and implicated the biosynthetic proteins in aiding mycobacterial survival in biofilms
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