Self-recycling and partially conservative replication of mycobacterial methylmannose polysaccharides

Abstract The steep increase in nontuberculous mycobacteria (NTM) infections makes understanding their unique physiology an urgent health priority. NTM synthesize two polysaccharides proposed to modulate fatty acid metabolism: the ubiquitous 6-O-methylglucose lipopolysaccharide, and the 3-O-methylmannose polysaccharide (MMP) only detected in rapidly growing mycobacteria. The recent identification of a unique MMP methyltransferase implicated the adjacent genes in MMP biosynthesis. We report a wide distribution of this gene cluster in NTM, including slowly growing mycobacteria, which we reveal to produce MMP. Using a combination of MMP purification and chemoenzymatic syntheses of intermediates, we identified the biosynthetic mechanism of MMP, relying on two enzymes that we characterized biochemically and structurally: a unique α–endomannosidade that hydrolyses MMP into defined-sized mannoligosaccharides that prime the elongation of new daughter MMP chains by a rare α-(1>4)-mannosyltransferase. Therefore, MMP biogenesis occurs through a partially conservative replication mechanism, and disruption of the mechanism affected mycobacterial growth rate at low temperature.