REGULATION OF GLYCOPROTEIN BIOSYNTHESIS

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The work proposed involves the development and employment of an assay for dolichyl phosphate, a phospholipid involved in glycoprotein biosynthesis. The assay is based on the purification of dolichyl phosphate using DEAE cellulose and two-dimensional thin-layer chromatography prior to derivatization and quantitation using C14-phenyl chloroformate. Also proposed is a study using rat liver cells in which the incorporation of C14-acetate into cholesterol and dolichyl phosphate will be monitored as a function of the level of HMG-CoA reductase, which will be inhibited to varying degrees using the specific antibiotic compactin. Such studies will provide information on the factors which control the rate of dolichyl phosphate biosynthesis. Finally a protocol is outlined for affinity chromatography of UDPG1cNAc-1-P transferase using the specific inhibitor tunicamycin bound to an insoluble resin.