Glycoprotein biosynthesis in Chlamydomonas. I. In vitro incorporation of galactose from UDP-(14C) galactose into membrane-bound protein

• Published in: Plant physiology (Bethesda) 1982-03, Vol.69 (3), p.678-681
• Main Author: Lang, W.C
• Format: Article
• Language: English
• Subjects: Algae; carbon; Cell walls; Chlamydomonas reinhardtii; Glycoproteins; Ions; isotopes; Particulate matter; Plant cells; Plants; Polymers; Product labeling; Radioactive decay; radionuclides
• ISSN: 0032-0889; 1532-2548
• DOI: 10.1104/pp.69.3.678

A crude membrane fraction from Chlamydomonas reinhardii was found to catalyze D-galactose transfer from UDP-galactose to endogenous proteins. Highest incorporation rates were achieved by incubation at 25°C and pH 7.5 in the presence of 10 millimolar Fe2+. Hydrolytic studies on the labeled polymer revealed that radioactivity was attached to protein via an alkali-stable and acid-labile linkage. Identification of galactose as the only labeled sugar in the acid hydrolysate and results of a tentative estimation of the molecular weight of the charged alkaline degradation product indicate that monomeric galactose units are transferred to form an O-glycosidic bond with peptidyl hydroxyproline. No indications were found for a similar linkage to serine which, in contrast to the hydroxyproline-O-glycoside linkage, is acid-stable but is cleaved by β-elimination. Chromatography of the sodium dodecyl sulfate-solubilized polymer on Sepharose-6B demonstrated that galactosyl residues are mainly associated with proteins which are of considerably higher molecular weight than are the majority of sodium dodecyl sulfate-denatured membrane proteins in this fraction.