Fig 6. Interaction of purified InlB with the native cell wall, WTA and LTA.
(A) Pulldown binding assay of His-tagged InlB to the indicated strains. WB of the entire Listeria pellet was performed, followed by detection using a His-tag antibody (control is 5 μg of purified His-tagged InlB only; blot is representative of three separate experiments, except for the complemented strain where n = 2). (B) Binding affinity response of immobilized InlB with WTA (upper) and the estimated KA (lower) for each interaction, as determined via BIAcore analysis. (C) Binding affinity response of immobilized InlB with LTA (upper) and the estimated KA (lower) for each interaction, as determined via BIAcore analysis. (RU: relative units; data for affinity responses of LTA/WTA is representative of three individual experiments, each time using newly purified InlB and WTA/LTA extracts). Estimated KA was determined by averaging four replicate titrations (see figure S9B for example) using two different chips with immobilized InlB; error bars represent SD for the four replicates; significance was determined via a paired one-way ANOVA; *P<0.05, ns = not significant).