Fig. 9.
Expression and secretion of YopD and YopE chimeras with a reciprocally exchanged N-terminal secretion signal. Overnight cultures were subcultured into BHI broth either containing (+) or lacking (−) calcium and grown at 26°C for 1 h and then at 37°C for 3 h. Protein samples were fractionated by 12% SDS-PAGE and then transferred onto a membrane support for immune detection. Expression fractions (upper panels) represent total protein associated with bacteria and also released into the culture supernatant. Secretion fractions (lower panels) signify protein freely released into the culture supernatant. Lanes: Parental (YopDwt), YPIII/pIB102; ΔyscU, ΔlcrQ (YopDwt), YPIII/pIB75-26; YopDB-Nterm, YPIII/pIB62577; YopEH-Nterm, YPIII/pIB578; YopDE-Nterm, YPIII/pIB62501; YopED-Nterm, YPIII/pIB577; YopDE-Nterm, YopED-Nterm, YPIII/pIB62501-577. Molecular mass values shown in parentheses were deduced from primary amino acid sequences.