Details

Autor(en) / Beteiligte

• Markgren, Per-Olof
• Hämäläinen, Markku
• Danielson, U.Helena

Titel

Kinetic Analysis of the Interaction between HIV-1 Protease and Inhibitors Using Optical Biosensor Technology

Ist Teil von

• Analytical biochemistry, 2000-03, Vol.279 (1), p.71-78

Ort / Verlag

United States: Elsevier Inc

Erscheinungsjahr

2000

Quelle

Access via ScienceDirect (Elsevier)

Beschreibungen/Notizen

• The interaction between HIV-1 protease and reversible inhibitors was studied by surface plasmon resonance biosensor technology. The steady-state binding level and the time course of association and dissociation could be observed by measuring the binding of inhibitors injected in a continuous flow 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. Affinities were estimated as B50 values from a series of sensorgrams at different concentrations of inhibitors. These values were found to be correlated with inhibition constants (Ki) determined by an enzyme inhibition assay (r2 = 0.84, logarithmic values). Dissociation rates were estimated at a single saturating concentration of the inhibitors as t1/2,obs, but these values did not correlate with Ki (r2 = 0.26, logarithmic values). Indinavir had the highest affinity (B50 = 11 nM) and the fastest dissociation (t1/2,obs = 500 s) among the clinically used inhibitors while saquinavir had a lower affinity (B50 = 25 nM) and the slowest dissociation rate (t1/2,obs = 6500 s). Since these two inhibitors have similar Ki values, the differences 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 performed in a buffer more similar to physiological conditions.