FIGURE 1.

Kinetics and stoichiometry of serpin B8 inhibition of furin. A, progress curves for inhibition of 2.6 nm furin by increasing concentrations of serpin B8-5S monitored from the exponential decrease in rate of furin cleavage of fluorogenic substrate as a function of time. B, dependence of observed pseudo-first order rate constants (k_obs) derived from exponential fits of progress curves in panel A on the serpin B8 concentration corrected for fluorogenic substrate competition as described under “Experimental Procedures.” Data are shown for reactions of serpin B8-5S (●) and serpin B8-5S5A (○). The slope of linear regression fits of the data (solid lines) provided the apparent second order association rate constants (k_a,app) for the inhibition reaction. C, stoichiometric titrations of furin with serpin B8 analyzed from the furin activity remaining after complete reaction with increasing molar ratios of serpin B8 to furin. Shown are reactions with serpin B8-5S (●) and serpin B8-5S5A (○). The SI was obtained from the abscissa intercept of linear regression fits of titrations (solid lines).