Figure 5

In vitro protein-protein interactions of MecA, SigX and ClpC by co-elution experiments. A. Protein-protein interactions were initiated by incubating the protein mixtures as follows: (1) MecA-His and GST (control), (2) MecA-His and GST-SigX, (3) MecA-His and GST-ClpC and (4) all three proteins. The reactions were incubated at 37°C for 1 hour before applied to Ni-charged resin for co-elution. The elution samples were then analyzed by Western blotting using the anti-His or anti-GST antibody, respectively. B. Effects of deletion of the N-terminal domain or C-terminal domain of MecA on its interaction with SigX or ClpC. Lane 1 and 4 indicate the interactions of the full-length MecA with both SigX and ClpC. Lane 2 indicates an interaction of the N-terminal domain (NTD_1–82) with SigX, but not with ClpC (Lane 5). Lane 6 indicates an interaction of the C-terminal domain (CTD_123–240) with ClpC but not with SigX (Lane 3).