Recognition of the lipid intermediate for arabinogalactan/arabinomannan biosynthesis and its relation to the mode of action of ethambutol on mycobacteria

Despite major advances in our understanding of the structure of mycobacterial cell walls, little is known of their biogenesis, and yet they are the site of action of many anti-tuberculosis drugs and implicated in much of the pathology of tuberculosis and leprosy. A family of monoglycosyl polyprenylphosphates was isolated from Mycobacterium smegmatis, containing arabinose, ribose, and mannose. The isoprenoid nature of the lipid components was established by 1H NMR, and fast atom bombardment mass spectroscopy (FAB-MS) demonstrated the presence of C50 decaprenyl-P derivatives and smaller amounts of the C35 octahydroheptaprenyl-P products. The configuration of the mycobacterial decaprenol was established as mono-trans, octa-cis, pointing to carriers of unusual structure. Combined gas chromatography (GC)/MS, FAB-MS/MS, and 1H NMR allowed characterization of one of the primary components as beta-D-arabinofuranosyl-1-monophosphodecaprenol. Pulse-chase metabolic labeling of cells with D-[14C]glucose indicated that the decaprenyl-P-arabinose is an active intermediate in the biosynthesis of the arabinan of cell wall arabinogalactan and arabinomannan. The identification of polyprenyl-P-ribose suggests the existence of ribose-containing polysaccharides in the cell walls of M. smegmatis or/and of a novel epimerase in the D-arabinose biosynthetic pathway. Ethambutol, a powerful anti-tuberculosis drug known to inhibit arabinogalactan and arabinomannan biosynthesis, results in the rapid accumulation of decaprenyl-P-arabinose, indicating that the drug interferes with either the transfer of arabinose from the donor or, alternatively, the synthesis of the arabinose acceptor itself.