Structure and ligand-binding modes of human serum albumin studied by UV resonance Raman spectroscopy

Ultraviolet resonance Raman spectra have been measured for human serum albumin (HSA) and its complexes with three types of ligands. Tyr and Trp Raman bands of ligand-free HSA are strongly enhanced with 240-nm excitation compared with those of aqueous amino acids, indicating that the side chains of some Tyr residues and the unique Trp at position 214 are hydrogen bonded in hydrophobic environments. The high intensity of Tyr and Trp Raman scattering remains unchanged in a wide pH range of 3.5-9.0 and the microenvironments of the Tyr and Trp residues seem conserved in this pH range. The Tyr Raman intensity of HSA increases upon binding of palmitic acid, indicating hydrophobic interactions between the Tyr phenol ring and the aliphatic chain of the bound fatty acid. Binding of warfarin, on the other hand, does not affect the Tyr Raman intensity. Instead, an increase is observed for the Raman intensity of Trp-214, which is located at the opening of the warfarin binding site. Concomitantly a conformation-marker Raman band of Trp-214 changes in frequency and the warfarin Raman intensity increases, suggesting direct hydrophobic interactions between Trp-214 and warfarin, Complexation of ibuprofen and HSA increases the Raman intensity of Tyr but not of ibuprofen, which is ascribed to the formation of a hydrogen bond between the carboxylate of ibuprofen and the phenolic OH of Tyr-411, the unique Tyr residue at the ibuprofen binding site. Hydrophobic interactions and hydrogen bonding of Tyr and Trp residues play a key role in ligand binding of HSA. (C) 1996 John Wiley & Sons, Inc.