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. 2012 Aug 2;8(8):e1002843. doi: 10.1371/journal.ppat.1002843

Figure 4. Bap binds the cellular receptor Gp96.

Figure 4

A) For the co-immunoprecipitation (Co-IP) recombinant Gp96 was incubated with purified Bap protein and immunoprecipitated with anti-Bap and protein G sepharose beads. In the control assay, the incubation step with Bap or Gp96 was omitted. Immunoprecipitated proteins were separated in 10% SDS gel, transferred onto a nitrocellulose membrane and probed with anti-Gp96 (α-Gp96) or anti-Bap (α-Bap). B) Recombinant Gp96 binds to S. aureus expressing Bap. Bacteria expressing Bap (V329) and the Bap deleted mutant (Δbap) were subcultured with 5 µg/ml of recombinant Gp96 in the absence or presence of anti-Bap serum as indicated. Unbound Gp96 was removed by extensive washing and Gp96 bound to bacteria was detected using immunoblot analysis. C) Bap proteins of coagulase negative staphylococci(S. epidermidis C533, S. hyicus 12, S. chromogenes C483) bind Bap. Bacteria were subcultured with 5 µg/ml of recombinant Gp96. Unbound Gp96 was removed by extensive washing and Gp96 bound to bacteria was detected using immunoblot analysis (upper part). Bap expression in S. epidermidis C533, S. hyicus 12, S. chromogenes C483 by immunoblot using polyclonal anti-Bap (α-Bap) serum (lower part).