FIG 4 .

CspA binds to C9 and inhibits C9 polymerization. (A) C9 binds to immobilized CspA, and the effect is dose dependent. Binding of C9 (0.01 to 20 µg/ml) to immobilized CspA (5 µg/ml) was assayed by ELISA, and bound C9 was detected with polyclonal C9 antiserum followed by HRP-conjugated anti-goat antibody. (B) Heparin affects the CspA-C9 interaction. The effect of heparin (1 to 500 µg/ml) on binding of C9 (5 µg/ml) to immobilized CspA (5 µg/ml) was assayed. (C) NaCl inhibits the CspA-C9 interaction. The effect of NaCl (0.1 to 1 M) on binding of C9 (5 µg/ml) to immobilized CspA (5 µg/ml) was assayed. The mean values from three separate experiments are shown, and error bars show SD. *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001. (D) CspA inhibits polymerization of C9. ZnCl₂ induced C9 polymerization, and after incubation the samples were separated by SDS-PAGE. Following silver staining, C9 polymers and C9 monomers were identified by their mobility. C9 polymerizes in the presence of ZnCl₂ (lane 1). CspA (2.5 and 5 µg) blocks polymer formation (lanes 2 and 3). The borrelial immune evasion protein ErpC (5 µg) (lane 4) does not influence C9 polymerization. In the absence of ZnCl₂, C9 does not form polymers (lane 5). The data shown are representative of three independent experiments.