FIG. 4.
Activity and localization of MalE::MzrA fusion proteins. (A) Western blot analysis of cell lysates obtained from strains expressing either no MzrA (pMalE-p2) or MalE::MzrA fusion proteins. Membrane blots were probed with OmpF/OmpC/OmpA antibodies. (B and C) Localization of MalE and MalE::MzrA fusion proteins. Cells were fractioned into whole-cell lysate (WCE), periplasm (Peri), or envelope (Env), as described in Materials and Methods. Protein samples from various fractions were subjected to Western blot analysis, and membrane blots were probed with MalE antibodies (B) and MalE and TolC antibodies (C). TolC, an outer membrane protein, serves as a control for the envelope fraction; TolC* refers to close approximation of TolC and MalE::MzrA fusion bands. ΔTM and Δ23 refer to the absence of the TM domain and the last 23 C-terminal residues from MzrA, respectively. Due to the leaky nature of the Ptac promoter, the effect of the recombinant proteins on OmpF and their cellular localization were studied from uninduced (without IPTG) cells.