Kinetic and Thermodynamic Studies on Afp Binding to Afp Antibody in Colorectal Tumor Homogenates

Abstract

Kinetic and thermodynamic parameters associated with the binding of ¹²⁵I- anti AFP Antibody to AFP in both crude colorectal homogenate and partially- purified fractions were investigated. It was shown that the reaction in all studied groups follow pseudo- first order reaction kinetics.The value of kinetic parameters K_a, K_d, K_obs, K₊₁, K_-1, (t_1/2) _ass., (t_1/2)_diss. and maximal binding capacity (B_max) at 25^oC for the binding of anti ¹²⁵I-AFP antibody with its cytosolic antigen in benign colorectal tumor were found to be : 0.0543 x 10¹⁰ M^-1, 15.064 x 10^-10M, 0.0158 min^-1, 4.086 x 10⁶ M^-1.min^-1, 64.76x10^-4 min^-1, 43 min, 104.28 min and 29.14 Pmol/mg protein respectively, while 0.0538 x 10¹⁰ M^-1, 15.142 x 10^-10 M, 0.0163 min^-1, 4.301 x 10⁶ M^-1.min^-1, 67.63 x 10^-4 min^-1, 40 min, 79.66 min and 46.25 Pmol/mg respectively for the binding of anti ¹²⁵I-AFP antibody with its cytosolic antigen in tissues of malignant colorectal tumor and at 4^oC.  The maximum binding of partially purified AFP occurred at 25^oC. The values ofK_a and K₊₁ increased with increasing temperature. The Van^’t Hoff plot demonstrated linear relationship between lnKa and 1/T, using crude colorectal homogenate partially purified AFP as AFP source. Plotting between ln K₊₁ and 1/T gave linear relationship called Arrheniuns relationship. The thermodynamic parameters ΔΗ˚, ΔG˚ and ΔЅ˚ for the formation of (¹²⁵I- anti AFP Antibody / AFP) complex at the standard state had been determinedas well asEa, ΔΗ*, ΔG* and ΔЅ* which representing the transition state.