Structural Analysis of Free N-Glycans in α-Glucosidase Mutants of Saccharomyces cerevisiae: Lack of the Evidence for the Occurrence of Catabolic α-Glucosidase Acting on the N-Glycans

Saccharomyces cerevisiae produces two different α-glucosidases, Glucosidase 1 (Gls1) and Glucosidase 2 (Gls2), which are responsible for the removal of the glucose molecules from N-g lycans (Glc 3 Man 9 GlcNAc 2 ) of glycoproteins in the endoplasmic reticulum. Whether any additional α-glucosidases playing a role in catabolizing the glucosylated N -glycans are produced by this yeast, however, remains unknown. We report herein on a search for additional α-glucosidases in S . cerevisiae . To this end, the precise structures of cytosolic free N -glycans (FNGs), mainly derived from the peptide: N -glycanase (Png1) mediated deglycosylation of N- glycoproteins were analyzed in the endoplasmic reticulum α-glucosidase-deficient mutants. 12 new glucosylated FNG structures were successfully identified through 2-dimentional HPLC analysis. On the other hand, non-glucosylated FNGs were not detected at all under any culture conditions. It can therefore be safely concluded that no catabolic α-glucosidases acting on N -glycans are produced by this yeast.