Application Of Kinetic Analysis Of Enzyme Inhibit Reaction Process

The first part: To investigate the prerequisite of the integrated method for characterizing inhibitors with the inhibition of xanthine on Candida species uricase as the model. Uricase reaction was monitored by absorbance at 293 nm. The apparent kinetic parameters (apparent Michaelis-Menten constant, K_mapp , and apparent maximal reaction rate, V_mapp) were estimated by the integrated method. The inhibition type and inhibition constant (Ki) were determined based on the responses of apparent kinetic parameters to xanthine concentrations. Outliers in reaction curve greatly reduced the reliability of parameters obtained by this integrated method. With residual substrate below one-sixth of K_mapp and initial substrate high enough, this integrated method usually showed deviation below 20% for K_mapp, and smaller deviation for V_mapp. By this integrated method with the same range of data for analysis, K_mapp was consistent to that by Lineweaver-Burk plot, and the response of K_mapp to xanthine concentrations gave inhibition constant of (5.4±0.8)μmol/L (n=3), also consistent to that by Lineweaver-Burk plot. This integrated method was feasible for fast screening of special inhibitors with data of higher precision...