The major aim of this work has been to gain further insights into the catalytic mechanism of carbonic anhydrase (carbonate hydro-lyase, EC 4.2.1.1). One approach has been to replace the essential Zn(II) ion by Cd(II) which has favourable spectroscopic properties. The Cd(II)-enzymes have appreciable 4-nitrophenyl acetate hydrolase activities. These activities increase with pH as if dependent on the basic form of a group with pKa near 10. The Cd(II)-carbonic anhydrases also have significant carbon dioxide hydration activities. Jhe Cd(II) derivatives are strongly inhibited by monovalent anions. The 113-Cd(II) derivatives have also been studied by 113-Cd NMR as a function of pH and bicarbonate or inhibitor concentration. Plots of chemical shift versus pH give sigmoidal titration curves in the studied pH range, 10.3. The p«a values vary from 9.2 to 9.7 correlating reasonably well with the activity profiles. When bicarbonate is added to the samples the 113-Cd resonances shift upfield to new characteristic positions. The inhibitors CN", SH", and SCN” bind directly to the metal ion with their C, S, and N atoms, respectively. The results are best explained by assuming a rapid exchange between three species in which the open coordination site of the metal ion is occupied by'hydroxide, water, or bicarbonate. Another approach has been to study kinetic properties of the active en­zyme. A number of monovalent anions were investigated as inhibitors of carbon dioxide hydration catalyzed by human carbonic anhydrase II. Predominantly uncompetitive inhibition patterns were observed at pH near 9 in all cases. The inhibition of human carbonic anhydrase II by the organic compounds tetrazole, 1,2,4-triazole, 2-nitrophenol, and chloral hydrate was also investigated. These inhibitors, together with phenol, can be classified in three groups depending upon the kinetic patterns of inhibition of carbon dioixde hydration at pH near 9. The first group, represented by tetrazole and 2-nitrophenol, yields predominantly uncompetitive inhibition under these conditions in analogy with simple, inorganic anions. The second group, represented by 1,2,4-triazole and chloral hydrate gives rise to essentially noncompetitive inhibition patterns whereas phenol, representing the third group, is a competitive inhibitor of carbon dioxide hydration. These results are analyzed in terms of two rivaling mechanism models, a kinetic scheme originally proposed by Steiner et al. (Eur. 3. Biochem. (1975) 59, 253-259) and a rapid-equilibrium kinetic scheme proposed by Pocker and Deits (3. Am. Chem. Soc. (1982) 104, 2424-2434). It is concluded that the observed steady-state inhibition patterns are compatible with both models, but hat discriminatory data, strongly favouring the model of Stêiner et al., are available in the literature. / <p>Diss. (sammanfattning) Umeå : Umeå universitet, 1984, härtill 4 uppsatser</p> / digitalisering@umu