title: Characterization of a putative alpha-mannosyltransferase involved in phosphatidylinositol trimannoside biosynthesis in Mycobacterium tuberculosis.
abstract: Phosphatidyl-myo-inositol mannosides (PIMs), lipomannan (LM) and lipoarabinomannan (LAM) are an important class of bacterial factors termed modulins that are found in tuberculosis and leprosy. Although their structures are well established, little is known with respect to the molecular aspects of the biosynthetic machinery involved in the synthesis of these glycolipids. On the basis of sequence similarity to other glycosyltransferases and our previous studies defining an alpha-mannosyltransferase from Mycobacterium tuberculosis, named PimB [Schaeffer, Khoo, Besra, Chatterjee, Brennan, Belisle and Inamine (1999) J. Biol. Chem. 274, 31625-31631], which catalysed the formation of triacyl (Ac(3))-PIM(2) (i.e. the dimannoside), we have identified a related gene from M. tuberculosis CDC1551, now designated pimC. The use of a cell-free assay containing GDP-[(14)C]mannose, amphomycin and membranes from Myobacterium smegmatis overexpressing PimC led to the synthesis of a new alkali-labile PIM product. Fast-atom-bombardment MS established the identity of the new enzymically synthesized product as Ac(3)PIM(3) (i.e. the trimannoside). The results indicate that pimC encodes an alpha-mannosyltransferase involved in Ac(3)PIM(3) biosynthesis. However, inactivation of pimC in Myobacterium bovis Bacille Calmette-Guérin (BCG) did not affect the production of higher PIMs, LM and LAM when compared with wild-type M. bovis BCG, suggesting the existence of redundant gene(s) or an alternate pathway that may compensate for this PimC deficiency. Further analyses, which compared the distribution of pimC in a panel of M. tuberculosis strains, revealed that pimC was present in only 22% of the clinical isolates examined.
authors.affiliation: Department of Microbiology and Immunology, University of Newcastle upon Tyne, Newcastle upon Tyne NE2 4HH, UK.
authors.forename: Laurent
authors.initials: L
authors.lastname: Kremer
authors.forename: Sudagar S
authors.initials: SS
authors.lastname: Gurcha
authors.forename: Pablo
authors.initials: P
authors.lastname: Bifani
authors.forename: Paul G
authors.initials: PG
authors.lastname: Hitchen
authors.forename: Alain
authors.initials: A
authors.lastname: Baulard
authors.forename: Howard R
authors.initials: HR
authors.lastname: Morris
authors.forename: Anne
authors.initials: A
authors.lastname: Dell
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Brennan
authors.forename: Gurdyal S
authors.initials: GS
authors.lastname: Besra
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D004269:DNA, Bacterial / Q000302:isolation & purification; D006155:Guanosine Diphosphate Mannose / Q000378:metabolism; D008070:Lipopolysaccharides / Q000378:metabolism; D008364:Mannosyltransferases / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D009163:Mycobacterium bovis / Q000235:genetics; D020102:Mycobacterium smegmatis / Q000201:enzymology; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics; D010716:Phosphatidylinositols / Q000378:metabolism*; D016415:Sequence Alignment; D016339:Spectrometry, Mass, Fast Atom Bombardment
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008070:Lipopolysaccharides; D010716:Phosphatidylinositols; C050016:lipoarabinomannan; C064351:lipomannan; C034154:phosphatidylinositol mannoside; D006155:Guanosine Diphosphate Mannose; D008364:Mannosyltransferases; C455962:pimC protein, Mycobacterium tuberculosis