Product Inhibition Study on Carbonic Anhydrase Using Spectroscopy and Calorimetry

Abstract

Kinetic and thermodynamic studies have been made on the effect of the p -nitrophenol product on the activity of bovine carbonic anhydrase in 50 mM Tris buffer pH 7.5, at 300 K using UV spectrophotometry and isothermal titration calorimetry (ITC). A competitive inhibition was observed for p -nitrophenol as a product of the enzymatic reaction. A graphical fitting method was used for determination of the binding constant and enthalpy of inhibitor binding using ITC data. The dissociation binding constant was 0.10 mM by the microcalorimetric method, which is in good agreement with the value of 0.11 mM for the inhibition constant obtained from the spectrophotometric method.

Keywords::

• Carbonic Anhydrase
• p -Nitrophenol
• Product Inhibition
• Inhibition Constant
• Isothermal Titration Calorimetry