Identification of an alpha(1-->6) mannopyranosyltransferase (MptA), involved in Corynebacterium glutamicum lipomanann biosynthesis, and identification of its orthologue in Mycobacterium tuberculosis.
Corynebacterium glutamicum and Mycobacterium tuberculosis share a similar cell wall architecture, and the availability of their genome sequences has enabled the utilization of C. glutamicum as a model for the identification and study of, otherwise essential, mycobacterial genes involved in lipomannan (LM) and lipoarabinomannan (LAM) biosynthesis. We selected the putative glycosyltransferase-Rv2174 from M. tuberculosis and deleted its orthologue NCgl2093 from C. glutamicum. This resulted in the formation of a novel truncated lipomannan (Cg-t-LM) and a complete ablation of LM/LAM biosynthesis. Purification and characterization of Cg-t-LM revealed an overall decrease in molecular mass, a reduction of alpha(1-->6) and alpha(1-->2) glycosidic linkages illustrating a reduced degree of branching compared with wild-type LM. The deletion mutant's biochemical phenotype was fully complemented by either NCgl2093 or Rv2174. Furthermore, the use of a synthetic neoglycolipid acceptor in an in vitro cell-free assay utilizing the sugar donor beta-D-mannopyranosyl-1-monophosphoryl-decaprenol together with the neoglycolipid acceptor alpha-D-Manp-(1-->6)-alpha-D-Manp-O-C8 as a substrate, confirmed NCgl2093 and Rv2174 as an alpha(1-->6) mannopyranosyltransferase (MptA), involved in the latter stages of the biosynthesis of the alpha(1-->6) mannan core of LM. Altogether, these studies have identified a new mannosyltransferase, MptA, and they shed further light on the biosynthesis of LM/LAM in Corynebacterianeae.
Mishra
Arun K
AK
School of Biosciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK.
Alderwick
Luke J
LJ
Rittmann
Doris
D
Tatituri
Raju V V
RV
Nigou
Jerome
J
Gilleron
Martine
M
Eggeling
Lothar
L
Besra
Gurdyal S
GS
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology; D048230:Corynebacterium glutamicum / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D016680:Genome, Bacterial; D006017:Glycolipids / Q000096:biosynthesis / Q000737:chemistry; D008070:Lipopolysaccharides / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D008364:Mannosyltransferases / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity
D001426:Bacterial Proteins; D006017:Glycolipids; D008070:Lipopolysaccharides; C064351:lipomannan; D008364:Mannosyltransferases
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