Over 25 nitrogen-containing heterocycles were tested as inhibitors of sweet almond β-glucosidase (EC 18.104.22.168). Among the most potent of these are some imidazole derivatives. The pH dependence indicates that the unprotonated inhibitor binds most tightly to the catalytically active species of the enzyme. This is analogous to the situation with 1-deoxynojirimycin where the permanently cationic species, N,N-dimethyl-1-deoxynojirimycin, binds at least two orders of magnitude less tightly to the enzyme than does the unprotonated 1-deoxynojirimycin. The binding of imidazole derivatives show a general tendency of increasing affinity with increasing basicity (β≈0.4). One derivative which shows a significant positive deviation from this correlation (-log Ki vs. pKa) is 4-phenylimidazole. 4-Phenylimidazole is one of the most potent reversible inhibitors of β-glucosidase with a pH-independent Ki=0.8 μM. It is also fairly specific for β-glucosidase, binding at least three orders of magnitude less tightly to any of the other exoglycosidases tested. This inhibitor combines, in a mono-molecular species, the binding affinities of benzene, which binds at the hydrophobic aglycone binding site, and imidazole, which binds at the sugar binding site of β-glucosidase. The binding energy of 4-phenylimidazole can be attributed to the sum of the intrinsic binding energies of the phenyl and imidazole moieties. Thus, there is no significant entropic advantage of combining the component parts of phenylimidazole in a single species. This indicates that there is no significant uncompensated entropy loss upon binding of either benzene or imidazole to the enzyme. Nevertheless, the additivity of binding energy, even in the absence of an entropic advantage, results in the most powerful known inhibitor of the enzyme.
|Number of pages||6|
|Journal||Biochimica et Biophysica Acta (BBA)/Protein Structure and Molecular|
|State||Published - 21 Dec 1989|