title: Specificity of the mannosyltransferase which initiates outer chain formation in Saccharomyces cerevisiae.
abstract: The in vitro specificity of the alpha 1-6 mannosyltransferase that initiates outer chain formation in Saccharomyces cerevisiae (Romero and Herscovics, J. Biol. Chem., 264, 1946-1950, 1989) was reassessed by fast atom bombardment mass spectrometry (FAB-MS). A particulate fraction from the mnn1 mutant was incubated with GDP-mannose and either Man9GlcNAc (M9T) isolated from thyroglobulin or Man8GlcNAc (M8Y) obtained by treatment of the M9T with the yeast specific mannosidase. The Man10GlcNAc (M10Y) and Man9GlcNAc (M9Y) oligosaccharides thus obtained, and the substrate oligosaccharides, were peracetylated or perdeuteroacetylated and submitted to FAB-MS using meta-nitrobenzylalcohol as the matrix. The latter was chosen as the matrix because it enhances the abundance of high-mass-fragment ions of peracetylated oligosaccharides and thereby facilitates the assignment of branching patterns. The results indicate that the alpha 1-6 mannosyltransferase catalyses the addition of mannose to the alpha 1-3 mannose residue, and thus provide additional new evidence to support the revised structure of yeast mannoproteins proposed by Hernandez et al. (J. Biol. Chem., 264, 11849-11856, 1989). [formula: see text] where Gn is N-acetylglucosamine, M is mannose and M is mannose added by the enzyme.
authors.affiliation: Department of Biochemistry, Imperial College of Science, Technology and Medicine, London, UK.
authors.forename: A J
authors.initials: AJ
authors.lastname: Reason
authors.forename: A
authors.initials: A
authors.lastname: Dell
authors.forename: P A
authors.initials: PA
authors.lastname: Romero
authors.forename: A
authors.initials: A
authors.lastname: Herscovics
mesh_terms: D002240:Carbohydrate Sequence; D008401:Gas Chromatography-Mass Spectrometry; D008361:Mannosidases / Q000378:metabolism; D008364:Mannosyltransferases / Q000378:metabolism*; D008956:Models, Chemical; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013379:Substrate Specificity; D013954:Thyroglobulin / Q000378:metabolism
chemical_list: D009844:Oligosaccharides; D013954:Thyroglobulin; D008364:Mannosyltransferases; D008361:Mannosidases