Glycosylation site-binding protein is not required for N-linked glycoprotein synthesis.
In prior studies we identified a 57-kDa protein in the lumen of the endoplasmic reticulum that, in addition to having both protein disulfide isomerase and thyroid hormone-binding protein activities, bound a photoaffinity probe containing the N-glycosylation-site sequence Asn-Xaa-Ser/Thr. It was hypothesized that this multifunctional protein, called glycosylation site-binding protein (GSBP), participated in the process of N-glycosylation of proteins. To test this hypothesis we have employed various conditions to deplete the lumen of GSBP and then assess the level of N-glycosylation catalyzed by oligosaccharyltransferase (OTase). Although most conditions leading to depletion resulted in partial loss of OTase activity, this loss was independent of the extent of GSBP depletion. Indeed, virtually complete loss (greater than 99%) of GSBP with partial retention of OTase activity was frequently observed. Moreover, repletion of the microsomal lumen with GSBP did not restore OTase activity to control levels. Thus, no correlation between GSBP content and OTase activity before or after reconstitution was found. These results suggest that this multifunctional 57-kDa protein is not an essential component of the enzymatic reaction in which oligosaccharide chains are transferred from dolichyl-P-P-GlcNAc2Man9Glc3 to nascent polypeptides or to synthetic tripeptide acceptors.
Noiva
R
R
Department of Biochemistry and Molecular Biology, University of Texas-M.D. Anderson Cancer Center, Houston 77030.
Kaplan
H A
HA
Lennarz
W J
WJ
D000818:Animals; D001665:Binding Sites; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006023:Glycoproteins / Q000096:biosynthesis*; D006031:Glycosylation; D006602:Hexosyltransferases; D007425:Intracellular Membranes / Q000378:metabolism; D007535:Isomerases / Q000378:metabolism*; D007700:Kinetics; D008565:Membrane Proteins; D008862:Microsomes, Liver / Q000378:metabolism*; D019704:Protein Disulfide-Isomerases; D051381:Rats; D014166:Transferases / Q000378:metabolism*
D006023:Glycoproteins; D008565:Membrane Proteins; D014166:Transferases; D006602:Hexosyltransferases; C025620:dolichyl-diphosphooligosaccharide - protein glycotransferase; D007535:Isomerases; D019704:Protein Disulfide-Isomerases