title: Biosynthesis of mycobacterial lipoarabinomannan: role of a branching mannosyltransferase.
abstract: Lipoarabinomannan (LAM), one of the few known bacterial glycosylphosphoinositides (GPIs), occurs in various structural forms in Mycobacterium species. It has been implicated in key aspects of the physiology of Mycobacterium tuberculosis and the immunology and pathogenesis of tuberculosis. Yet, little is known of the biosynthesis of LAM. A bioinformatics approach identified putative integral membrane proteins, MSMEG4250 in Mycobacterium smegmatis and Rv2181 in M. tuberculosis, with 10 predicted transmembrane domains and a glycosyltransferase (GT) motif (DID), features that are common to eukaryotic mannosyltransferases (ManTs) of the GT-C superfamily that rely on polyprenyl-linked rather than nucleotide-linked sugar donors. Inactivation of M. smegmatis MSMEG4250 by allelic exchange resulted in altered growth and inability to synthesize lipomannan (LM) but accumulation of a previously uncharacterized, truncated LAM. MALDI-TOF/MS and NMR indicated a structure lower in molecular weight than the native molecule, a preponderance of 6-linked Manp residues, and the absence of 2,6-linked and terminal Manp. Complementation of the mutant with the corresponding ortholog of M. tuberculosis H37Rv restored normal LM/LAM synthesis. The data suggest that MSMEG4250 and Rv2181 are ManTs that are responsible for the addition of alpha(1-->2) branches to the mannan core of LM/LAM and that arrest of this branching in the mutant deters formation of native LAM. The results allow for the presentation of a unique model of LM and LAM biosynthesis. The generation of mutants defective in the synthesis of LM/LAM will help define the role of these GPIs in the immunology and pathogenesis of mycobacterial infections and physiology of the organism.
authors.affiliation: Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523, USA.
authors.forename: Devinder
authors.initials: D
authors.lastname: Kaur
authors.forename: Stefan
authors.initials: S
authors.lastname: Berg
authors.forename: Premkumar
authors.initials: P
authors.lastname: Dinadayala
authors.forename: Brigitte
authors.initials: B
authors.lastname: Gicquel
authors.forename: Delphi
authors.initials: D
authors.lastname: Chatterjee
authors.forename: Michael R
authors.initials: MR
authors.lastname: McNeil
authors.forename: Varalakshmi D
authors.initials: VD
authors.lastname: Vissa
authors.forename: Dean C
authors.initials: DC
authors.lastname: Crick
authors.forename: Mary
authors.initials: M
authors.lastname: Jackson
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Brennan
mesh_terms: D000483:Alleles; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D019295:Computational Biology; D005816:Genetic Complementation Test; D008070:Lipopolysaccharides / Q000096:biosynthesis*; D008364:Mannosyltransferases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D020102:Mycobacterium smegmatis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development*; D009169:Mycobacterium tuberculosis / Q000201:enzymology / Q000235:genetics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D001426:Bacterial Proteins; D008070:Lipopolysaccharides; C050016:lipoarabinomannan; C064351:lipomannan; D008364:Mannosyltransferases