Topography of basal and stimulated biosynthesis of GlcNAc-P-P-dolichol and (GlcNAc)2-P-P-dolichol: variation in effect of proteolysis.
Evidence has recently appeared indicating the cytoplasmic orientation in microsomal vesicles of the GlcNAc-transferases that participate in the biosynthesis of GlcNAc-P-P-dolichol and (GlcNAc)2-P-P-dolichol. The topography of the stimulation of these activities brought about by dolichol-P-mannose and phosphatidylglycerol, however, is not known. The present report continues studies of the topography of these early reactions of the dolichol pathway, examined under basal and stimulatory conditions after proteolysis of intact microsomes isolated from livers of the embryonic chick. Under all conditions which were examined, the GlcNAc-transferase concerned with the biosynthesis of the chitobiosyl derivative was completely inhibited, consistent with its cytoplasmic orientation. The effect of proteolysis on the formation of GlcNAc-P-P-dolichol, however, varied with different proteolytic enzymes, ranging from no inhibition to over 90% inhibition. This variation in response suggests that the GlcNAc-transferase may be partially buried within the microsomal bilayer which only some preparations of proteolytic enzymes can penetrate. Evidence was obtained indicating the cytoplasmic orientation of the stimulation of GlcNAc-P-P-dolichol biosynthesis. These studies support the conclusion that both the catalytic and allosteric activating sites of the GlcNAc-transferase have a similar topography.
Department of Ophthalmology, Case Western Reserve University, Cleveland, OH 44106.
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Kean
D000818:Animals; D002642:Chick Embryo; D004288:Dolichol Phosphates / Q000378:metabolism*; D010450:Endopeptidases; D004789:Enzyme Activation; D008862:Microsomes, Liver / Q000378:metabolism*; D017351:N-Acetylglucosaminyltransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D011105:Polyisoprenyl Phosphate Sugars / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism*
D004288:Dolichol Phosphates; D011105:Polyisoprenyl Phosphate Sugars; D011506:Proteins; C026406:dolichol pyrophosphate; D017351:N-Acetylglucosaminyltransferases; C023553:mannose-6-phosphatase; D010744:Phosphoric Monoester Hydrolases; D010450:Endopeptidases