Isolation of new nonconditional Saccharomyces cerevisiae mutants defective in asparagine-linked glycosylation

Glycobiology
P MañasL M Hernandez

Abstract

We describe the isolation and partial characterization of Saccharomyces cerevisiae nonconditional mutants that show defects in N-glycosylation of proteins. The selection method is based on the reduction of affinity for the ion exchanger QAE-Sephadex as a consequence of the decrease in the negative charge of the cell surface. This characteristic reflects a decrease in the incorporation of mannosylphosphate units into the N-linked oligosaccharides of the mannoproteins. The mutants exhibit low affinity for the basic dye alcian blue and for that reason we have called them Idb (low dye binding) mutants. Eight of the complementation groups seem to be new as shown by complementation studies with previously isolated mutants of similar phenotype. Four of the groups showed a significant reduction in the number and/or size of the N-linked oligosaccharides attached to secreted invertase. We have analyzed the N-linked oligosaccharides of Idb1 and Idb2, the mutants that show the most drastic reduction in the affinity for the alcian blue dye. In both cases, the purified endo H-released oligosaccharides from the mannoproteins lacked detectable amounts of phosphate groups as shown by ion exchange chromatography and the 1H NMR spectra. In additi...Continue Reading

Citations

Apr 20, 2006·Antonie van Leeuwenhoek·Isaac CorbachoLuis M Hernández
Jan 8, 1999·Biochimica Et Biophysica Acta·N Dean
Jan 8, 1999·Biochimica Et Biophysica Acta·Y Jigami, T Odani
Mar 25, 2000·FEMS Microbiology Letters·A B Vázquez-ReynaA Flores-Carreón
Feb 26, 2000·Molecular Biology of the Cell·B Santos, M Snyder
Jul 29, 2010·Glycobiology·Isaac CorbachoLuis M Hernández
Jan 4, 2012·FEMS Yeast Research·Isaac CorbachoLuis M Hernández
Apr 28, 2016·World Journal of Microbiology & Biotechnology·R Sidari, A Caridi
Jul 11, 2013·PloS One·Juergen H NettStephen R Hamilton
Jan 22, 2004·FEMS Yeast Research·Isaac CorbachoLuis M Hernández

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