FIG 4.

Interaction of LcpA with vitronectin. (A) (Left) Purified recombinant proteins were subjected to SDS–15% PAGE under reducing conditions, transferred to a nitrocellulose membrane, and stained with Ponceau S. (Right) The membrane was incubated with 10% NHS, and vitronectin binding was detected with polyclonal goat antibodies recognizing human vitronectin. (B) Whole-cell lysates from L. interrogans serovar Kennewicki strain Fromm were separated by SDS–15% PAGE and transferred to nitrocellulose membranes. (Left) One membrane was incubated with 10% NHS, and vitronectin binding was detected with polyclonal goat antibodies recognizing human vitronectin. (Right) The other membrane was probed with LcpA antiserum (5). (C) The effect of heparin (0.1 to 10 μg/ml) on binding of vitronectin (10 μg/ml) to immobilized LcpA (10 μg/ml) was assayed. (D) The effect of NaCl (62.5 to 500 mM) on binding of vitronectin (10 μg/ml) to immobilized LcpA (10 μg/ml) was assayed. Each bar represents the mean absorbance value at 492 nm ± the SD for 3 independent experiments, each performed in triplicate. ***, P ≤ 0.001. Statistical analyses were performed using ANOVA.