Figure 3
Glycosylation of CPY and Gas1p is impaired in the gdt1Δ mutant in the presence of 500 mM external Ca2+.
Total membrane protein extracts of the indicated strains were prepared from cultures grown to an OD600 of 1.2 in YD medium (YD) alone (A) or supplemented with 500 mM CaCl2 (Ca2+) (B), increasing Ca2+ concentrations (C), 500 μM MnCl2 (Mn2+) (D) or both 500 mM CaCl2 and 500 μM MnCl2 (Ca2+Mn2+) (E). Where indicated, the proteins were digested with endoglycosidase H (0.5 U/ml). Levels of CPY and Gas1p were then analyzed by SDS-PAGE followed by Western blotting analysis using specific antibodies. The different forms are indicated by arrows. All displayed results are representative of those seen in at least three replicates.
