FIG 2.

Cofactor activity of FH bound to LcpA or to intact L. interrogans. C3b (500 ng) and FI (250 ng) were added to immobilized recombinant proteins (1 μg) or to bacteria (2 × 10⁸ cells) that had been preincubated with FH. The reaction mixtures were incubated for 1, 2, or 4 h at 37°C. The products were analyzed by SDS-PAGE, and the cleavage fragments of C3b were detected by Western blotting with anti-human C3 polyclonal antibodies. The presence of bands at 43, 46, and 68 kDa indicates that acquired FH was able to promote FI-mediated cleavage of C3b. Lanes: 1, purified C3b; 2, C3b, FH, and FI (positive control); 3, L. interrogans, FH, C3b, and FI; 4 to 7, LcpA, FH, C3b, and FI; 8 to 11, LcpA, C3b, and FI; 12 to 15, LIC10301, FH, C3b, and FI.