Synergistic binding of inhibitors to the protease from HIV type 1

Abstract

Inhibition of the protease in HIV is a potentially useful approach for the treatment of AIDS. In the course of evaluating inhibitors of the HIV-1 protease, we observed a strong synergism between certain inhibitors that might be expected to bind to different sites in this enzyme. The binding affinity of carbobenzyloxyisoleucinylphenylalaninol, for example, is increased 125-fold in the presence of carbobenzyloxyglutaminylisoamylamide. These synergistic effects between inhibitors have specific structural requirements that correlate well with the known substrate preference of the enzyme. The modular basis for this phenomenon remains to be elucidated but it could involve substrate-induced conformational change as part of the reaction mechanism. Similar effects have been reported previously for several zinc proteases. Thus this work extends the observation to a different class of enzymes and suggests that the phenomenon might be widespread.

Analysis of the interactions between an enzyme and multiple inhibitors using combination plots

“…In our experience with the above method [11][12][13][14], there are a number of important deficiencies. First, as a means of determining the value of α ",# , the method suffers from the fact that the family of lines do not generally intersect at a single point owing to the scatter in the data associated with experimental error (as shown in Figure 1a).…”

“…When the above authors considered nonexclusive inhibitors, they assumed (often without stating explicitly) that the interaction coefficient (α ",# ) was equal to 1. Our extensive work (covering seven different enzymes) [4,[11][12][13][14] indicates, in contrast, that this condition is rarely fulfilled. Instead α may range from less than 0.01 all the way to infinity.…”

Identification of broad-based HIV-1 protease inhibitors from combinatorial libraries

“…An alternative to competitive inhibitors has been the identification of inhibitors that target non-active site regions of the protease, such as the dimer interface [5, 6], flaps [7, 8], or other non-substrate active site regions [9]. Moreover, inhibitors that utilize non-competitive, uncompetitive, or mix-mode mechanisms have also been identified [5–8, 10, 11]. A potential advantage of a non-competitive mechanism will be the insensitivity to substrate concentrations, which may better maintain a therapeutic threshold in the substrate-rich virion.…”

Analysis of the combined effect of two linear inhibitors on a single enzyme

“…To maximize the affinity of an enzyme for an inhibitor, it is of interest to exploit all the potential interactions available. In proteases these interactions are numerous and intricate because of the highly extended nature of the active site [9]. The possible existence of interactions between subsites has been examined by analysing the combined effect of short peptides as competitive inhibitors of the protease, and substantial synergy has been found for some combinations [9][10][11].…”