35CL NUCLEAR MAGNETIC RESONANCE STUDIES OF A ZINC METALLOENZYME CARBONIC ANHYDRASE

The interaction of chloride ions with the Zn(II) metal of bovine carbonic anhydrase has been studied by 35Cl nuclear magnetic resonance spectroscopy. It has been shown that there exists one zinc coordination site available for chloride interaction. This zinc-chloride interaction can be inhibited by cyanide and acetazolamide. The zinc-chloride interaction is highly pH dependent decreasing to zero above pH 10. The apparent pKa of the chloride broadening is chloride concentration dependent and measurements at several chloride concentrations yield, on extrapolation to zero chloride, a value of 7.0 ± 0.1. The 35Cl nuclear magnetic resonance spectrum has been observed in the presence of the apoenzyme. The apoenzyme is approximately 25 times less effective in broadening the chloride line than carbonic anhydrase. Carbonic anhydrase is approximately a 1000-fold more effective than aquo Zn(II) ions in broadening the chloride line. The effectiveness of carbonic anhydrase, as compared to aquo Zn(II) ions, in changing the 35Cl relaxation rate is attributed to the fact that both the chloride exchange process and the molecular rotational interactions have correlation times of the order of 10-8 sec, whereas for aquo Zn(II) ions the molecular rotational process dominates the relaxation mechanism with a correlation time of the order of 10-11 sec. © 1969, American Chemical Society. All rights reserved.