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The primary glycosylation defect in class e thy 1 negative mutant mouse lymphoma cells is an inability to synthesize dolichol phospho mannose

, : The primary glycosylation defect in class e thy 1 negative mutant mouse lymphoma cells is an inability to synthesize dolichol phospho mannose. Journal of Biological Chemistry 255(10): 4441-4446

Thy-1-- mutant mouse lymphoma cells of the class E complementation group are unable to synthesize the normal Glc3Man9GlcNAc2 lipid-linked oligosaccharide, but instead accumulate a smaller lipid-linked species with the structure Man.alpha.1 .fwdarw. 2Man.alpha.1 .fwdarw. 2Man.alpha.1 .fwdarw. 3(Man.alpha.1 .fwdarw. 6)Man.beta.1 .fwdarw. 4GlcNAc.beta.1 .fwdarw. 4GlcNAc. The primary defect in the Thy-1- cells is an inability to synthesize dolichol-P-mannose. Intact Thy-1- cells incubated with [2-3H]mannose failed to incorporate any detectable radioactiVity into dolicho-P-mannose and crude membrane preparations of Thy-1- cells were unable to transfer mannose from GDP-[3H]mannose to dolichol-P. These membrane preparations did incorporate [3H]mannose into lipid-linked oligosaccharides up to the size of Man5GlcNAc2, suggesting that these species are formed from mannosyl donors other than dolichol-P-mannose. When class E Thy-1- membrane preparations were incubated with exogenous dolicho-P-[3H]mannose, lipid-linked oligosaccharides ranging in size from Man6GlcNAc2 to Man9GlcNAc2 were formed. The mutant cells have the .alpha.1,3-mannosyltransferase necessary for the conversion of the Man5GlcNAc2 species to Man6GlcNAc2, but lack the appropriate mannosyl donor (dolichol-P-mannose) necessary for the formation of the larger lipid-linked oligosaccharides. At least 2 mannosyl donors are involved in the synthesis of lipid-linked oligosaccharides. GDP-mannose is the probable donor for the formation of the Man1-5GlcNAc2 species, while dolichol-P-mannose donates the 6th mannosyl residue and probably mannose residues 7 through 9.

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