Figure 4.

Monitoring of opsin N-glycan formation and processing by pulse–chase. (A) Inducible HEK293S cell line. Opsin expression was induced, the total mixture pulsed with [³⁵S]methionine for 40 min, and chased with cold methionine for times indicated. ³⁵S-methionine-labeled opsin was purified from DM-solubilized cell extracts by using rho-1D4-Sepharose. Samples were examined by SDS/PAGE (lanes 2–9) and gels were dried before autoradiography. ¹⁴C-labeled markers (M) are in lanes 1 and 10. (B) The protocol was the same as in A except that the HEK293S GnTI⁻ cell line was used. For details, see the text.