Figure 5.

HK and cytokeratin interactions. (A) Microtiter plates were coated in 0.1 M Na₂CO₃ buffer (pH 9.6) with a purified cytokeratin mixture or BSA both at 1 μg/well. The ability of HK to bind to cytokeratin was determined by adding increasing concentrations of biotin–HK (2–80 nM) to wells coated with cytokeratin or BSA in the absence (−Zn) or presence (+Zn) of 50 μM Zn²⁺. The data presented are the mean ± SEM of three individual experiments at each point. (B) Investigation to determine whether cytokeratin blocks HK binding to HUVEC. Increasing concentrations of purified cytokeratin (0.002–3,000 nM) were incubated with biotin–HK (20 nM) in Hepes–Tyrode’s buffer containing 50 μM Zn²⁺ over confluent HUVEC in microtiter plates. After incubation for 1 h, the cells were washed and the degree of biotin–HK bound to the HUVEC was determined by procedures reported in Experimental Procedures. Each point is the mean ± SEM of three determinations.