title: NMR and modeling studies of protein-carbohydrate interactions: synthesis, three-dimensional structure, and recognition properties of a minimum hevein domain with binding affinity for chitooligosaccharides.
abstract: HEV32, a 32-residue, truncated hevein lacking eleven C-terminal amino acids, was synthesized by solid-phase methodology and correctly folded with three cysteine bridge pairs. The affinities of HEV32 for small chitin fragments--in the forms of N,N',N"-triacetylchitotriose ((GlcNAc)3) (millimolar) and N,N',N",N"',N"",N""'-hexaacetylchitohexaose ((GlcNAc)6) (micromolar)--as measured by NMR and fluorescence methods, are comparable with those of native hevein. The HEV32 ligand-binding process is enthalpy driven, while entropy opposes binding. The NMR structure of ligand-bound HEV32 in aqueous solution was determined to be highly similar to the NMR structure of ligand-bound hevein. Solvated molecular-dynamics simulations were performed in order to monitor the changes in side-chain conformation of the binding site of HEV32 and hevein upon interaction with ligands. The calculations suggest that the Trp21 side-chain orientation of HEV32 in the free form differs from that in the bound state; this agrees with fluorescence and thermodynamic data. HEV32 provides a simple molecular model for studying protein-carbohydrate interactions and for understanding the physiological relevance of small native hevein domains lacking C-terminal residues.
authors.affiliation: Department of Protein Structure and Function, Centro de Investigaciones Biológicas, CSIC, Ramiro de Maeztu 9, 28040 Madrid, Spain.
authors.forename: Nuria
authors.initials: N
authors.lastname: Aboitiz
authors.forename: Miquel
authors.initials: M
authors.lastname: Vila-Perelló
authors.forename: Patrick
authors.initials: P
authors.lastname: Groves
authors.forename: Juan Luis
authors.initials: JL
authors.lastname: Asensio
authors.forename: David
authors.initials: D
authors.lastname: Andreu
authors.forename: Francisco Javier
authors.initials: FJ
authors.lastname: Cañada
authors.forename: Jesús
authors.initials: J
authors.lastname: Jiménez-Barbero
mesh_terms: D000485:Allergens / Q000737:chemistry*; D027721:Amaranthus / Q000737:chemistry; D000595:Amino Acid Sequence; D000890:Anti-Infective Agents / Q000737:chemistry / Q000494:pharmacology; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry*; D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000737:chemistry*; D002851:Chromatography, High Pressure Liquid; D018360:Crystallography, X-Ray; D020315:Latex Hypersensitivity; D037102:Lectins / Q000737:chemistry; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry*; D037121:Plant Lectins / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D011506:Proteins / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D013816:Thermodynamics
chemical_list: D000485:Allergens; D000890:Anti-Infective Agents; D023181:Antimicrobial Cationic Peptides; D002241:Carbohydrates; D037102:Lectins; D008024:Ligands; D009844:Oligosaccharides; D037121:Plant Lectins; D011506:Proteins; C045875:hevein