THE INTERACTIONS BETWEEN CELLULAR RETINOL-BINDING PROTEIN (CRBP-I) AND RETINAL - A VIBRATIONAL SPECTROSCOPIC STUDY

Preresonance Raman difference spectra have been obtained for all-trans retinal in dilute CCl4 solution, complexed with cellular retinol-bind ing protein (CRBP-I) and retinol-binding protein (RBP). These spectra indicate that retinal is of a slightly more planar conformation within the binding pocket of CRBP-I than in solution or hydrophobically comp lexed with RBP. Compared to retinal in solution or bound to RBP, the c onformation of the polyene tail of the retinal chromophore is perturbe d from C8 through C11. This perturbation is probably due to the close proximity of the Lys40 in the CRBP-I binding pocket to the above-menti oned carbons. The C=O stretching vibration of bound retinal carbonyl h as been found to shift from 1664 cm(-1) solubilized in CCl4 to 1650 an d 1645 cm(-1) in RBP and CRBP-I, respectively, and significantly broad ened in both cases. The frequency shift and broadening have been attri buted to hydrogen bonding. These have been compared to calibrations of frequency shift (Delta nu(C=O)) vs. Delta H and Delta G of all-trans retinal complexed with a series of phenol derivatives of incremental p roton-donating ability as obtained by the relationship of van't Hoff. By this relationship, the binding enthalpy of the all-trans retinal ca rbonyl moiety bound to CRBP-I and RBP is -28.1 kJ/mol (-6.7 kcal/mol) and -23.5 kJ/mol (-5.6 kcal/mol), respectively. The free energy of bin ding of the retinal carbonyl bound to CRBP-I and RBP has been determin ed to be -10.5 kJ/mol (-2.5 kcal/mol) and -7.2 kJ/mol (-1.7 kcal/mol), respectively. The hydrogen-bonded C=O moiety of retinal complexed wit h CRBP-I accounts for a substantial (25%) but not overriding amount of the binding energy of CRBP-I for retinal, and it also accounts for th e protein's preference for binding retinol. (C) 1997 John & Wiley & So ns, Inc.