Effect of GTP on the dolichol pathway for protein glycosylation in rat liver microsomes.
Incubation of native rat liver microsomes with GTP resulted in enhanced incorporation of N-acetylglucosamine (GlcNAc) from UDP-GlcNAc into lipid acceptors. The stimulation of GlcNAc transfer by GTP was specific for GTP; ATP exerted no effect. The GTP effect was blocked by a non-hydrolysable GTP analogue guanosine 5'-[beta gamma-imido]triphosphate, indicating that GTP hydrolysis was crucial. Though dolichyl pyrophosphate NN'-diacetylchitobiose [Dol-PP-(GlcNAc)2] was the main radiolabelled product formed upon incubation of GTP-treated microsomes with UDP-GlcNAc, GTP selectively stimulated UDP-GlcNAc:dolichyl phosphate (Dol-P) N-acetylglucosaminyl 1-phosphotransferase (N-acetylglucosaminyl 1-phosphotransferase). This conclusion was reached on the basis of experiments in which tunicamycin was used to selectively inhibit N-acetylglucosaminyl 1-phosphotransferase. The enhanced transformation of Dol-P to dolichyl pyrophosphate N-acetylglucosamine (Dol-PP-GlcNAc) by GTP ultimately led to enhanced protein glycosylation. GTP-induced stimulation of GlcNAc incorporation in lipid and protein by GTP was observed also in microsomes fully permeabilized with Staph. aureus alpha-toxin. These findings refute the previous proposal [Godelaine, Beaufay, Wibo and Ravoet (1983) J. Cell Biol. 97, 340-350] that increased membrane permeability constitutes the mechanism whereby GTP activates the reactions of the dolichol pathway.
Bossuyt
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Laboratory of Biological Chemistry, Katholieke Universiteit Leuven, Belgium.
Blanckaert
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D000117:Acetylglucosamine / Q000378:metabolism; D000818:Animals; D002463:Cell Membrane Permeability; D004286:Dolichols / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism*; D006027:Glycosides / Q000378:metabolism; D006031:Glycosylation; D006160:Guanosine Triphosphate / Q000494:pharmacology*; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism*; D017351:N-Acetylglucosaminyltransferases / Q000378:metabolism; D051381:Rats; D017855:Transferases (Other Substituted Phosphate Groups) / Q000378:metabolism
D004286:Dolichols; D006023:Glycoproteins; D006027:Glycosides; D006160:Guanosine Triphosphate; D017351:N-Acetylglucosaminyltransferases; C037649:N-acetyllactosaminide beta-1,6-N-acetylglucosaminyltransferase; D017855:Transferases (Other Substituted Phosphate Groups); C023222:UDPacetylglucosamine-dolichyl-phosphate acetylglucosamine-1-phosphate transferase; D000117:Acetylglucosamine