SYNERGISTIC BINDING OF HYDROPHOBIC PROBES AND ZINC LIGANDS TO THERMOLYSIN

The strong synergism previously observed in the binding of inhibitors to two Zn-proteases, has also been found for thermolysin. As in earlie r cases, the effects are produced by a small Zn-ligand (e.g. a hydroxa mate) in the presence of another compound which contains the key struc tural features of specific substrates (a specificity probe). For therm olysin, the most effective specificity probes are hydrophobic derivati ves of amines and amino acids (e.g. carbobenzyloxy-L-alaninol). Even t he simple combination of benzyl alcohol and formohydroxamate displays considerable synergism. The above effects are temperature dependent an d correlate well with a thermally induced conformational isomerization reported recently for this enzyme. Our results seem to be related to previous observations of substrate synergism in the reverse reaction a nd to superactivation by chemical modification of this enzyme. All the se effects are consistent with a change in the environment of the cata lytically important zinc atom upon binding of the hydrophobic side cha in of the substrate. With the inclusion of thermolysin, binding synerg ism is now known to occur in an endopeptidase as well as in exopeptida ses of diverse specificity. The general occurrence of this phenomenon in zinc proteases and its possible significance are discussed in an ac companying study.