Structural analysis and biosynthesis gene cluster of an antigenic glycopeptidolipid from Mycobacterium intracellulare.
Mycobacterium avium-Mycobacterium intracellulare complex (MAC) is the most common isolate of nontuberculous mycobacteria and causes pulmonary and extrapulmonary diseases. MAC species can be grouped into 31 serotypes by the epitopic oligosaccharide structure of the species-specific glycopeptidolipid (GPL) antigen. The GPL consists of a serotype-common fatty acyl peptide core with 3,4-di-O-methyl-rhamnose at the terminal alaninol and a 6-deoxy-talose at the allo-threonine and serotype-specific oligosaccharides extending from the 6-deoxy-talose. Although the complete structures of 15 serotype-specific GPLs have been defined, the serotype 16-specific GPL structure has not yet been elucidated. In this study, the chemical structure of the serotype 16 GPL derived from M. intracellulare was determined by using chromatography, mass spectrometry, and nuclear magnetic resonance analyses. The result indicates that the terminal carbohydrate epitope of the oligosaccharide is a novel N-acyl-dideoxy-hexose. By the combined linkage analysis, the oligosaccharide structure of serotype 16 GPL was determined to be 3-2'-methyl-3'-hydroxy-4'-methoxy-pentanoyl-amido-3,6-dideoxy-beta-hexose-(1-->3)-4-O-methyl-alpha-L-rhamnose-(1-->3)-alpha-L-rhamnose-(1-->3)-alpha-L-rhamnose-(1-->2)-6-deoxy-alpha-L-talose. Next, the 22.9-kb serotype 16-specific gene cluster involved in the glycosylation of oligosaccharide was isolated and sequenced. The cluster contained 17 open reading frames (ORFs). Based on the similarity of the deduced amino acid sequences, it was assumed that the ORF functions include encoding three glycosyltransferases, an acyltransferase, an aminotransferase, and a methyltransferase. An M. avium serotype 1 strain was transformed with cosmid clone no. 253 containing gtfB-drrC of M. intracellulare serotype 16, and the transformant produced serotype 16 GPL. Together, the ORFs of this serotype 16-specific gene cluster are responsible for the biosynthesis of serotype 16 GPL.
Fujiwara
Nagatoshi
N
Department of Host Defense, Osaka City University Graduate School of Medicine, 1-4-3 Asahi-machi, Abeno-ku, Osaka 545-8585, Japan. fujiwara@med.osaka-cu.ac.jp
Nakata
Noboru
N
Naka
Takashi
T
Yano
Ikuya
I
Doe
Matsumi
M
Chatterjee
Delphi
D
McNeil
Michael
M
Brennan
Patrick J
PJ
Kobayashi
Kazuo
K
Makino
Masahiko
M
Matsumoto
Sohkichi
S
Ogura
Hisashi
H
Maeda
Shinji
S
D002240:Carbohydrate Sequence; D003837:Deoxy Sugars / Q000737:chemistry*; D005798:Genes, Bacterial; D006017:Glycolipids / Q000032:analysis* / Q000737:chemistry; D006020:Glycopeptides / Q000737:chemistry* / Q000235:genetics; D053858:Metabolic Networks and Pathways / Q000235:genetics; D005810:Multigene Family; D015269:Mycobacterium avium Complex / Q000737:chemistry* / Q000145:classification / Q000235:genetics* / Q000378:metabolism
D003837:Deoxy Sugars; D006017:Glycolipids; D006020:Glycopeptides