Nanosecond spectroscopy of retinol bound to proteins

Abstract

Nanosecond fluorescence techniques have been employed for studying the mode of binding of retinol to the retinol binding protein (RBP), β-lactoglobulin, and human serum albumin. A substantial increase in the fluorescence decay time of bound retinol relative to that in hexane is attributed to a change in its conformation upon binding. It is demonstrated that retinol bound to RBP and to β-lactoglobulin lacks flexibility in the nanosecond scale. The latter protein, which is shown to be globular, induces the appearance of structure in the absorption spectrum of the ligand but not in its fluorescence spectrum. It is demonstrated that the mirror-image relationship between the modified absorption and fluorescence spectra of retinol bound to this protein breaks down. This implies the existence of a profound difference between the nuclear configurations of the ground and excited singlet electronic states of bound retinol.