FIG. 3.

Immunoblot analysis of fusion proteins produced by E. coli DH5α harboring the indicated plasmid. Lanes: 1, pEHSTN24; 2, pEHSTC22; 3, pEHProSTC28; 4, pSTC17+pDSPH524. Recombinants were cultured at 37°C in LB liquid (A) or LB agar medium (B to D). (A) Supernatants were from broth cultures containing 1 × 10⁹ to 2 × 10⁹ CFU/ml. (B) Supernatants were from plate-gown cultures containing 5 × 10¹⁰ to 2 × 10¹¹ CFU/ml. Supernatant samples were boiled in Laemmli buffer with β-ME and loaded onto 0.1% SDS–15% polyacrylamide gels. After electrophoresis, proteins were electrotransferred to nitrocellulose membranes and incubated either with ClpG-specific antiserum (A and B) or with the STa-specific monoclonal antibody 11C (C) or 20C1 (D). HRP-labeled goat anti-rabbit IgG (A and B) or biotin-labeled goat anti-mouse IgG in concert with HRP-conjugated streptavidin (C and D) was used as secondary antibody. The arrows at left point to the position of ClpG as assessed by the simultaneous migration of ClpG-containing solid culture supernatant from E. coli DH5α(pEH524) (data not shown).