Mammalian glycosyltransferases prefer glycosyl phosphoryl dolichols rather than glycosyl phosphoryl polyprenols as substrates for oligosaccharyl synthesis.
We have studied the effectiveness of polyprenyl-P-mannose and polyprenol-P-glucose as donor substrates for the dolichyl-P-mannose:Man5(GlcNAc)2-PP-dolichol mannosyltransferase and the dolichyl-P-glucose:Man9(GlcNAc)2-PP-dolichol glucosyltransferase, respectively. The polyprenol moiety differs from dolichol only in the unsaturation of the terminal isoprene unit of the molecule. Based on the kinetics of the reactions, we have found that both glycosyltransferases have higher apparent Kms and lower apparent Vmaxs using polyprenyl-P-monosaccharides as substrates rather than the dolichyl-P-monosaccharides. The products formed with the polyprenyl-P-sugars were the same as those formed by the dolichol-linked sugars, indicating that the polyprenol substrates could be utilized by the glycosyltransferases in vitro. The results also indicate that the dolichyl-P-sugars and the polyprenyl-P-sugars compete for the same binding site on the enzyme. These findings are significant in terms of understanding the glycosylation phenotypes of Chinese hamster ovary cell mutants of the Lec9 complementation group, which lack the ability to convert polyprenol into dolichol.
D'Souza-Schorey
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C
Department of Biochemistry, University of Arkansas for Medical Sciences, Little Rock 72205.
McLachlan
K R
KR
Krag
S S
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Elbein
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AD
D000818:Animals; D016466:CHO Cells; D002240:Carbohydrate Sequence; D002462:Cell Membrane / Q000201:enzymology; D006224:Cricetinae; D004287:Dolichol Monophosphate Mannose / Q000378:metabolism*; D005964:Glucosyltransferases / Q000378:metabolism*; D007700:Kinetics; D008364:Mannosyltransferases / Q000378:metabolism*; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000096:biosynthesis*; D011103:Polyisoprenyl Phosphate Monosaccharides / Q000378:metabolism*; D013379:Substrate Specificity
D009844:Oligosaccharides; D011103:Polyisoprenyl Phosphate Monosaccharides; D004287:Dolichol Monophosphate Mannose; C002968:dolichol-D-glucosylmonophosphate; D005964:Glucosyltransferases; D008364:Mannosyltransferases; C054676:dolichol phosphate glucose-dolichol diphosphate oligosaccharide glucosyltransferase; C066968:dolichyl-P-Man:Man(5)GlcNAc(2)-PP-dolichol alpha-1,3-mannosyltransferase