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. 2003 Oct 13;163(1):35–44. doi: 10.1083/jcb.200306069

Figure 2.

Figure 2.

Site-specific incorporation of Tmd-Phe into pOmpA does not interfere with its translocation into membrane vesicles. In addition to wild-type pOmpA (wt), the indicated stop codon mutants located in the mature part and the signal sequence of OmpA were expressed in vitro by the help of suppressor tRNA charged with Tmd-Phe. Radiolabeled proteins were separated by SDS-PAGE and visualized by phosphorimaging. Addition of E. coli INVs and proteinase K (PK) is indicated. Precursor (pOmpA) and mature (OmpA) forms are labeled with open and closed arrowheads, respectively. Note the aberrant electrophoretic mobility of the mutant precursor carrying Tmd-Phe at position 11 within the signal sequence. The precursor of OmpA often resolves on SDS-PAGE into two closely spaced species, the reason for that being unknown. The background processing of the different pOmpAs obtained in the absence of added INVs (lanes 1, 5, 9, and 13) is most likely due to traces of INVs contaminating the ribosome preparation used.