title: New-found phenolic glycolipids in Mycobacterium bovis BCG. Presence of a diglycosylated glycolipid.
abstract: A crude phenolic glycolipid extract from Mycobacterium bovis bacille Calmette-Guerin (BCG) was fractionated until homogeneity at the intact level into four phenolic glycolipids called B, B-1, B-2, and B-3 according to their polarity. The apolar one, which is the most abundant was assigned to the well-known mycoside B. The B-2 and B-3 phenolic glycolipids were purified by direct-phase high performance liquid chromatography using a 5 micron Spherisorb column but were only recovered in small amounts (3 mg). A linear gradient of 0-20% methanol in chloroform was used. The B-1, B-2, and B-3 glycolipids were subjected to suitable modern analytical techniques selected for their potential to elucidate the structure at the intact level. Desorption chemical ionization-mass spectrometry allowed the molecular mass of B-3 to be determined as 1652 Da for the major homolog establishing the molecular formula as C103H192O14. Thus, the B-3 polar phenolic glycolipid contained two deoxyhexoses, one molecule of phenolphthiocerol esterified by two molecules of mycocerosic acid. Using two-dimensional 1H NMR (correlated chemical shift and nuclear Overhauser effect spectroscopy) at the intact level the B-3 oligosaccharide structure was determined as an alpha-L-Rhap-(1----3)-2-O-Me-alpha-L-Rhap. This is the first report of a diglycosylated phenolic glycolipid in a nonpathogenic mycobacteria. The disaccharide unit, the antigenic determinant, appears to be characteristic of M. bovis BCG. This polar glycolipid B-3 and the apolar ones, B-1 and B-2, were reactive in enzyme-linked immunosorbent assay against serum from rabbit hyperimmunized with M. bovis BCG.
authors.affiliation: Centre de Recherche de Biochimie et Génétique Cellulaires du Centre National de la Recherche Scientifique, Toulouse, France.
authors.forename: A
authors.initials: A
authors.lastname: Vercellone
authors.forename: G
authors.initials: G
authors.lastname: Puzo
mesh_terms: D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D006017:Glycolipids / Q000032:analysis* / Q000276:immunology; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009163:Mycobacterium bovis / Q000032:analysis* / Q000276:immunology
chemical_list: D006017:Glycolipids