title: Bile salt recognition by human liver fatty acid binding protein.
abstract: Fatty acid binding proteins (FABPs) act as intracellular carriers of lipid molecules, and play a role in global metabolism regulation. Liver FABP (L-FABP) is prominent among FABPs for its wide ligand repertoire, which includes long-chain fatty acids as well as bile acids (BAs). In this work, we performed a detailed molecular- and atomic-level analysis of the interactions established by human L-FABP with nine BAs to understand the binding specificity for this important class of cholesterol-derived metabolites. Protein-ligand complex formation was monitored using heteronuclear NMR, steady-state fluorescence spectroscopy, and mass spectrometry. BAs were found to interact with L-FABP with dissociation constants in the narrow range of 0.6-7 μm; however, the diverse substitution patterns of the sterol nucleus and the presence of side-chain conjugation resulted in complexes endowed with various degrees of conformational heterogeneity. Trihydroxylated BAs formed monomeric complexes in which single ligand molecules occupied similar internal binding sites, based on chemical-shift perturbation data. Analysis of NMR line shapes upon progressive addition of taurocholate indicated that the binding mechanism departed from a simple binary association equilibrium, and instead involved intermediates along the binding path. The co-linear chemical shift behavior observed for L-FABP complexes with cholate derivatives added insight into conformational dynamics in the presence of ligands. The observed spectroscopic features of L-FABP/BA complexes, discussed in relation to ligand chemistry, suggest possible molecular determinants of recognition, with implications regarding intracellular BA transport. Our findings suggest that human L-FABP is a poorly selective, universal BA binder.
authors: Department of Biotechnology, University of Verona, Italy.
authors: Filippo
authors: F
authors: Favretto
authors: Carlo
authors: C
authors: Santambrogio
authors: Mariapina
authors: M
authors: D'Onofrio
authors: Henriette
authors: H
authors: Molinari
authors: Rita
authors: R
authors: Grandori
authors: Michael
authors: M
authors: Assfalg
mesh_terms: D001647:Bile Acids and Salts / Q000737:chemistry*; D001665:Binding Sites; D050556:Fatty Acid-Binding Proteins / Q000737:chemistry*; D006801:Humans; D006900:Hydroxylation; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular; D011485:Protein Binding; D017433:Protein Structure, Secondary; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D001647:Bile Acids and Salts; D050556:Fatty Acid-Binding Proteins
keywords: NMR spectroscopy; bile salts; fatty acid binding protein; mass spectrometry; protein ligand binding