Kinetic study of catalytic CO 2 hydration by water-soluble model compound of carbonic anhydrase and anion inhibition effect on CO 2 hydration

Abstract

A kinetic study of CO 2 hydration was carried out using the water-soluble zinc model complex with water-soluble nitrilotris(2-benzimidazolylmethyl-6-sulfonate) L1S, [L1SZn(OH 2)] −, mimicking the active site of carbonic anhydrase, in the presence and absence of anion inhibitors NCS − and Cl −. The obtained rate constants k cat for CO 2 hydration were 5.9×10 2, 1. 7×10 3, and 3.1×10 3 M −1 s −1 at 5, 10, and 15 °C, respectively: the k cat=ca. 10 4 M −1 s −1 extrapolated towards 25 °C has been the largest among the reported k cat using zinc model complexes for carbonic anhydrase. It was also revealed that NCS −, Cl − and acetazolamide play a role of inhibitors by the decrease of k cat: 7×10 2 and 2×10 3 M −1 s −1 for NCS − and Cl − at 15 °C, respectively. The sequence of their magnitudes in k cat is Cl − ∼acetazolamide>NCS −, where the sequence Cl −>NCS − is confirmed for native carbonic anhydrase. The difference of k cat or k obs between NCS − and Cl − resulted from that between the stability constants K st=2×10 3 for [L1SZn(NCS)] 2− and 1×10 2 M −1 for [L1SZnCl] 2− in D 2O: for water-insoluble tris(2-benzimidazolylmethyl)amine L1, K st=1.8×10 4 for [L1Zn(NCS)] 2− and 1.5×10 3 M −1 for [L1ZnCl] 2−in CD 3CN/D 2O (50% v/v). The crystal structure of anion-binding zinc model complexes [L1Zn(OH 2)] 0.5[L1ZnCl] 0.5 (ClO 4) 1.5 1 0.5 2 0.5(ClO 4) 1.5 was revealed by X-ray crystallography. The geometry around Zn 2+ in 1 and 2 was tetrahedrally coordinated by three benzimidazolyl nitrogen atoms and one oxygen atom of H 2O, or Cl −.