title: Recovery of dolichyl diphosphate oligosaccharide in methanolic aqueous phase prepared from rat liver microsomal fractions.
abstract: The synthesis of dolichyl diphosphate oligosaccharide was studied by incubating rat liver microsomes (microsomal fractions) with GDP-[14C]mannose, UDP-glucose, UDP-N-acetylglucosamine and [3H]dolichol phosphate. The labelled products obtained by the first step of extraction of the microsomes in methanolic aqueous phase (MAP fraction in chloroform/methanol/water; 3:2:1, by vol.) and in CMW fraction (chloroform/methanol/water; 10:10:3, by vol.) obtained by extraction of the interphase after the first step of extraction were analysed on a DEAE-cellulose column. With the progress of incubation, the radioactivity in unchanged GDP-mannose decreased, whereas the labelled dol-P-P-oligo in the MAP fraction increased about 5-6-fold. The lipid oligosaccharide in this fraction accounted for about 50-60% of the GDP-mannose used, whereas the recovery of the labelled lipid oligosaccharide in the CMW fraction was about 10%. The lipid oligosaccharide from both reactions after mild acid hydrolysis were analysed by gel filtration on Bio-Gel P-4. The oligosaccharide from the MAP fraction gave a peak of higher Mr distinctly separate from the lower-Mr peak obtained from the CMW fraction. Microsomes incubated with labelled lipid oligosaccharide from the MAP fraction showed incorporation of the label into endogenous protein.
mesh_terms: D000818:Animals; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D008070:Lipopolysaccharides / Q000096:biosynthesis / Q000302:isolation & purification; D008862:Microsomes, Liver / Q000378:metabolism*; D009844:Oligosaccharides / Q000302:isolation & purification; D011104:Polyisoprenyl Phosphate Oligosaccharides / Q000302:isolation & purification*; D011105:Polyisoprenyl Phosphate Sugars / Q000302:isolation & purification*; D051381:Rats
chemical_list: D008070:Lipopolysaccharides; D009844:Oligosaccharides; D011104:Polyisoprenyl Phosphate Oligosaccharides; D011105:Polyisoprenyl Phosphate Sugars; C024962:dolichyl diphosphate oligosaccharides; C023023:lipid-linked oligosaccharides