title: Functional characterisation of three o-methyltransferases involved in the biosynthesis of phenolglycolipids in Mycobacterium tuberculosis.
abstract: Phenolic glycolipids are produced by a very limited number of slow-growing mycobacterial species, most of which are pathogen for humans. In Mycobacterium tuberculosis, the etiologic agent of tuberculosis, these molecules play a role in the pathogenicity by modulating the host immune response during infection. The major variant of phenolic glycolipids produced by M. tuberculosis, named PGL-tb, consists of a large lipid core terminated by a glycosylated aromatic nucleus. The carbohydrate part is composed of three sugar residues, two rhamnosyl units and a terminal fucosyl residue, which is per-O-methylated, and seems to be important for pathogenicity. While most of the genes responsible for the synthesis of the lipid core domain and the saccharide appendage of PGL-tb have been characterized, the enzymes involved in the O-methylation of the fucosyl residue of PGL-tb remain unknown. In this study we report the identification and characterization of the methyltransferases required for the O-methylation of the terminal fucosyl residue of PGL-tb. These enzymes are encoded by genes Rv2954c, Rv2955c and Rv2956. Mutants of M. tuberculosis harboring deletion within these genes were constructed. Purification and analysis of the phenolglycolipids produced by these strains, using a combination of mass spectrometry and NMR spectroscopy, revealed that Rv2954c, Rv2955c and Rv2956 encode the methyltransferases that respectively catalysed the O-methylation of the hydroxyl groups located at positions 3, 4 and 2 of the terminal fucosyl residue of PGL-tb. Our data also suggest that methylation at these positions is a sequential process, starting with position 2, followed by positions 4 and 3.
authors.affiliation: Centre national de la recherche scientifique, Institut de Pharmacologie et de Biologie Structurale, Toulouse, France.
authors.forename: Roxane
authors.initials: R
authors.lastname: Simeone
authors.forename: Gaëlle
authors.initials: G
authors.lastname: Huet
authors.forename: Patricia
authors.initials: P
authors.lastname: Constant
authors.forename: Wladimir
authors.initials: W
authors.lastname: Malaga
authors.forename: Anne
authors.initials: A
authors.lastname: Lemassu
authors.forename: Françoise
authors.initials: F
authors.lastname: Laval
authors.forename: Mamadou
authors.initials: M
authors.lastname: Daffé
authors.forename: Christophe
authors.initials: C
authors.lastname: Guilhot
authors.forename: Christian
authors.initials: C
authors.lastname: Chalut
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002384:Catalysis; D023061:Gene Order; D006017:Glycolipids / Q000096:biosynthesis* / Q000737:chemistry; D006801:Humans; D008745:Methylation; D008780:Methyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment
chemical_list: D006017:Glycolipids; D008780:Methyltransferases
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