FIGURE 5.

Time courses of Cu/Ph-induced cross-linking of mutant P28C/C154 at different temperatures. A and B, Western blots. RSO vesicles containing P28C/C154 LacY with tandem fXa sites were cross-linked on ice (A) or at room temperature (B) at the times given, and subsequently cleaved with fXa protease. LacY was detected with anti-C-terminal antibody as described under “Experimental Procedures.” C, time courses of cross-linking. Cross-linking is expressed as the density a band at a given time point relative to that of the undigested band. Solid line with a filled circle, room temperature; broken line with filled square, 0 °C.