Binding Constant Measurement by Hyper-Rayleigh Scattering: Bilirubin-Human Serum Albumin Binding as a Case Study

Abstract

In this paper, a new application of the hyper-Rayleigh scattering technique in determining multiple binding constants of a small molecule like bilirubin to a macromolecule like the protein human serum albumin has been demonstrated. Human serum albumin has two binding sites for bilirubin, and the binding constants have been measured by carrying out a second harmonic titration of the protein against bilirubin and vice versa. The measured binding constants K_1 = 1.5 \pm 0.43 \times 10^7 M^-^1 and K_2 = 1.01 \pm 0.16 \times 10^6 M^-^1 agree well with the reported values obtained by other methods