Nonspecific cross-linking of membrane-bound wild-type SecYEG and SecA₇₉₅–Y₂₆₈EG. Proteoliposomes containing SecYEG or the cross-linked complex (A₇₉₅–Y₂₆₈EG) were cross-linked using Tris-bipyridylruthenium and exposure to light, as indicated in seconds (see Materials and methods), followed by treatment with DTT to quench the reaction and break the disulphide bond. Proteins were separated by SDS-PAGE and visualized by anti-SecY immunoblot. u/c refers to protein controls where no cross-linker was added and SecY-Y to a genetically fused SecY dimer (Duong, 2003). Bands corresponding to uncross-linked SecY, to two conjoined or cross-linked SecY subunits (Y-Y), and specifically/nonspecifically cross-linked SecA-SecY (A-Y) are shown.