Fig. 5

Comparative analysis of glycosylation occupancy and glycan patterns of W303A-pY26, W303A-RHO1, Δalg3Δoch1-pY26 and Δalg3Δoch1-RHO1. a Analysis of glycosylation occupancy of CPY proteins. 5 ml of yeast cells were incubated in SD-U medium for 48 h, and then transferred into fresh SD-U medium for another 24 h (30 °C, 230 rpm). 5 ml of growing cells were harvested and washed twice with deionized water. Intracellular proteins were extracted and analyzed by western blotting with anti-ScCPY antibody as described in the “Methods” section. The number of N-glycosylation sites is indicated by ‘n’. b Analysis of N-glycan profiles by HPLC. Δalg3Δoch1-pY26 (dotted line), Δalg3Δoch1-RHO1 (line segment), and Δalg3Δoch1 (solid line) were cultured in 5 ml of SD-U liquid medium at 30 °C for 48 h, and then transferred to fresh 50 ml of SD-U medium for 24 h. N-glycans were extracted from 5 ml of each culture. Three peaks represent Man₅GlcNAc₂, Man₆GlcNAc₂ and Man₇GlcNAc₂, respectively