Kinetics and mechanism of the interaction between human serum albumin and monomeric haemin

Abstract

The interaction of human serum albumin with monomeric haemin has been investigated by detailed kinetic analysis in dimethyl sulphoxide/water (3:5, v/v). The results obtained under conditions of albumin saturation of haemin and under pseudo-single turnover conditions indicate that methaemalbumin is formed in a two-stage, single-intermediate process. The initial association between the haemin and human serum albumin is a chemically controlled process (k1 = 1.7 X 10(5) mol-1 . s-1 . dm3 at 24 degrees C); the variation of K1 with pH exhibited a well defined pK of 5.9. The overall equilibrium constant, calculated by using microscopic rate constants, is 1.1 (+/- 0.5) X 10(8) mol-1 at 24 degrees C. The data and conclusions are consistent with a general binding mechanism for albumin in which intermediate formation is followed by an entropy-controlled internalization of the ligand.