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 Ka, Kd, Kobs, K+1, K-1, (t1/2) ass., (t1/2)diss. and maximal binding capacity (Bmax) at 25oC 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 4oC. The maximum binding of partially purified AFP occurred at 25oC. The values ofKa 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.