Kinetic analysis of the interaction between HIV-1 protease and inhibitors using optical biosensor technology

The interaction between HIV-1 protease and reversible inhibitors was studie d by surface plasmon resonance biosensor technology, The steady-state bindi ng level and the time course of association and dissociation could be obser ved by measuring the binding of inhibitors injected in a continuous how of buffer to the immobilized enzyme. Fourteen low molecular weight inhibitors (500-700 Da), including the four clinically used HIV-1 protease inhibitors (indinavir, nelfinavir, ritonavir, and saquinavir), were analyzed. Affiniti es were estimated as B-50 values from a series of sensor-grams at different concentrations of inhibitors. These values were found to be correlated wit h inhibition constants (K-i) determined by an enzyme inhibition assay (r(2) = 0.84, logarithmic values). Dissociation rates were estimated at a single saturating concentration of the inhibitors as t(1/2,obs), but these values did not correlate with K-i (r(2) = 0.26, logarithmic values). Indinavir ha d the highest affinity (B-50 = 11 nM) and the fastest dissociation (t(1/2,o bs) = 500 s) among the clinically used inhibitors while saquinavir had a lo wer affinity (B-50 = 25 nM) and the slowest dissociation rate (t(1/2,obs) = 6500 s), Since these two inhibitors have similar Ki values, the difference s in dissociation rates reveal important characteristics in the interaction that cannot be obtained by the inhibition studies. The biosensor data are expected to be of greater in vivo relevance since the experiments were perf ormed in a buffer more similar to physiological conditions. (C) 2000 Academ ic Press.