Three-Dimensional Structure of Bovine Heart Fatty-acid-binding Protein with Bound Palmitic Acid, Determined by Multidimensional NMR Spectroscopy

• DOI: 10.1111/j.1432-1033.1995.0266i.x
• Publication Date: 2005-03-04T14:32:38Z
• Authors: Dirck Lassen; Christian Lucke; Marina Kveder; Azita Mesgarzadeh; Jurgen M. Schmidt; Bernfried Specht; Axel Lezius; Friedrich Spener; Heinz Ruterjans
• Keywords: 0301 basic medicine; 0303 health sciences; Magnetic Resonance Spectroscopy; Molecular Sequence Data; Palmitic Acid; Palmitic Acids; Fatty Acid-Binding Proteins; Protein Structure, Secondary; Recombinant Proteins; Neoplasm Proteins; 03 medical and health sciences; Animals; Cattle; distance geometry ; energy minimization ; protein-ligand interaction ; isotope enrichment; Amino Acid Sequence; Carrier Proteins

The three-dimensional structure of the holo form of recombinant cellular bovine heart fatty-acid-binding protein (H-FABPc), a polypeptide of 133 amino acid residues with a molecular mass of 15 kDa, has been determined by multidimensional homonuclear and heteronuclear NMR spectroscopy applied to uniformly 15N-labeled and unlabeled protein. A nearly complete set of 1H and 15N chemical shift assignments was obtained. A total of 2329 intramolecular distance constraints and 42 side-chain chi 1 dihedral-angle constraints were derived from cross-relaxation and J coupling information. 3D nuclear Overhauser enhancement and exchange spectroscopy combined with heteronuclear multiple-quantum coherence (NOESY-HMQC) experiments, performed on a sample of uniformly 13C-labeled palmitic acid bound to unlabeled cellular heart fatty-acid-binding protein revealed 10 intermolecular contacts that determine the orientation of the bound fatty acid. An ensemble of protein conformations was calculated with the distance-geometry algorithm for NMR applications (DIANA) using the redundant dihedral-angle constraint (REDAC) strategy. After docking the fatty acid into the protein, the protein-ligand arrangement was subject to distance-restrained energy minimization. The overall conformation of the protein is a beta-barrel consisting of 10 antiparallel beta-strands which form two nearly orthogonal beta-sheets of five strands each. Two short helices form a helix-turn-helix motif in the N-terminal region of the polypeptide chain. The palmitic acid is bound within the protein in a U-shaped conformation close to the two helices. The obtained solution structure of the protein is consistent with a number of fatty-acid-binding-protein crystal structures.