Inhibition of human carbonic anhydrase II by anions and some ‘neutral’ compounds

Abstract

Anions inhibit carbonic anhydrase-catalyzed reactions by binding to the zinc ion in the active center. The pH dependence of the inhibition of the esterase activity shows that anions predominantly bind to enzymes having a protonated catalytic group. Formally, anion binding can be described as a competition with OH − for a coordination site on the metal ion [1]. Pocker and Deits [2] recently showed that anions inhibit the CO 2 hydration catalyzed by bovine carbonic anhydrase at high pH values in an uncompetitive fashion, and they presented a kinetic scheme to explain this phenomenon. We have studied the anion inhibition of human carbonic anhydrase II (or C). We have confirmed the ▪ observations of Pocker and Deits that at least certain anions, such as SCN − [3], OCN −, and N − 3 , give rise to uncompetitive inhibition patterns of CO 2 hydration at high pH. This is illustrated for N − 3 in Fig. 1. However, we offer an alternative interpretation to that of Pocker and Deits. We will show that the uncompetitive pattern is a direct consequence of the kinetic mechanism which we have proposed previously [3, 4]. The crucial point is that a strongly anion binding enzyme species, HisEZn 2+OH 2 (see ref. [5]), precedes the rate-limiting step in catalysis. Recently, some organic compounds that bind at or near the metal ion have received attention. One of these is phenol which has been shown to act as a competitive inhibitor of CO 2 hydration and as a mixed noncompetitive inhibitor of HCO − 3 dehydration [6]. We have also studied o-nitrophenol, 1,2,4-triazole and tetrazole. However, these inhibitors behave kinetically like anions and give rise to mixed uncompetitive-noncompetitive patterns in the CO 2 hydration reaction at high pH.