N-glycosylation analysis of cellular and cell surface Hjv. Cotransfection of HEK293 cells with Hjv and Mt2 or mutants, as well as the biotinylation of cell surface proteins, was performed as described in the Figure 2A legend. Cell lysate (input) and biotinylated cell surface proteins were subjected to digestions by Endo-H (H) or PNGase F (F), followed by immunodetection with anti-FLAG HRP for Hjv, Mt2, and Mt2 mutants. Two images for cell surface proteins with different exposure times are presented. All experiments were repeated at least 3 times, with consistent results. Ctrl, control; IB, immunoblotting.