Characterization of an oligosaccharide-pyrophosphodolichol pyrophosphatase activity in yeast.
Use of Triton X-114 allowed us to develop a new method to separate hydrophilic oligosaccharidic material from hydrophobic oligosaccharide pyrophosphodolichols (oligosaccharide-PP-Dol). Taking advantage of this procedure we characterize, in yeast microsomal membranes, an enzymic activity that hydrolyses oligosaccharide-PP-Dol into oligosaccharidic material. H.p.l.c. analysis together with alkaline-phosphatase- and endo-N-acetyl-beta-D-glucosaminidase-susceptibility demonstrate that the oligosaccharidic released material is mainly composed of oligomannosides containing a chitobiose phosphate at the reducing end. The enzymic activity requires bivalent cations and is inhibited by pyrophosphate, NAD+ and bacitracin. As other, commercially available, pyrophosphatases have no action on lipid intermediates, the described pyrophosphatase activity appears to be the specific enzyme for oligosaccharide-PP-Dol. This enzymic splitting of the pyrophosphate bond might be the primary event in the catabolism of lipid intermediates.
Belard
M
M
Laboratoire de Chimie Biologique (Unité Associée au C.N.R.S. No 217), Université des Sciences et Techniques de Lille Flandres-Artois, Villeneuve d'Ascq, France.
Cacan
R
R
Verbert
A
A
D001414:Bacitracin / Q000494:pharmacology; D002413:Cations, Divalent / Q000494:pharmacology; D002850:Chromatography, Gel; D003902:Detergents / Q000494:pharmacology; D006031:Glycosylation; D007425:Intracellular Membranes / Q000201:enzymology; D008861:Microsomes / Q000201:enzymology; D009243:NAD / Q000494:pharmacology; D017830:Octoxynol; D009844:Oligosaccharides / Q000378:metabolism; D010710:Phosphates / Q000494:pharmacology; D011092:Polyethylene Glycols; D011104:Polyisoprenyl Phosphate Oligosaccharides / Q000378:metabolism; D011755:Pyrophosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D002413:Cations, Divalent; D003902:Detergents; D009844:Oligosaccharides; D010710:Phosphates; D011104:Polyisoprenyl Phosphate Oligosaccharides; C024962:dolichyl diphosphate oligosaccharides; D009243:NAD; D001414:Bacitracin; D011092:Polyethylene Glycols; D017830:Octoxynol; D011755:Pyrophosphatases; C057833:oligosaccharide-diphosphodolichol pyrophosphatase