Figure 1.
Protease protection of ER proteins in RM and EKRM. 4 eq. of either RM (A and C) or EKRM (B) were diluted to 20 μl in a buffer containing 25 mM K-Hepes, pH 7.2, 25 mM KOAc, and 2.5 mM Mg(OAc)2 at 4°C. Samples were treated with chymotrypsin (CT) at the indicated concentration for 30 min at 4°C and reactions quenched by addition of TCA to 10%. After centrifugation, samples were processed for SDS-PAGE and immunoblotted with antibodies directed against Sec61p, p180, and p34, as described in Materials and Methods. The migration of full-length Sec61p, p180, and p34 are indicated by arrows; a prominent limit digestion product of Sec61p is indicated by an asterisk.