title: Hydrogen bonding geometry of a protein-bound carbohydrate from water exchange-mediated cross-relaxation.
abstract: We present heteronuclear two-dimensional methods for the analysis of the geometry of exchangeable protons on a protein-bound carbohydrate. By using a water-selective NOESY-HSQC, we observed cross-relaxation between carbohydrate hydroxyl protons and non-exchangeable ring protons in the complex of [13C6]-alpha-methyl-D-mannopyranoside with recombinant rat mannose binding protein. Using a simple kinetic model, we were able to explain the differences in the initial slopes of the resulting cross-relaxation buildup curves in terms of the geometry of the hydroxyl protons in the bound state. The hydroxyl rotamers consistent with our cross-relaxation data fit very well with predictions based on the crystal structure of MBP bound to a mannose-rich oligosaccharide. These methods should be applicable to other systems where both ligand exchange and water exchange are fast relative to the rate of cross-relaxation.
authors.affiliation: Department of Pharmacology, Yale University, New Haven, CT 06510, USA.
authors.forename: E W
authors.initials: EW
authors.lastname: Sayers
authors.forename: J L
authors.initials: JL
authors.lastname: Weaver
authors.forename: J H
authors.initials: JH
authors.lastname: Prestegard
mesh_terms: D000818:Animals; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000737:chemistry*; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D006860:Hydrogen Bonding; D066298:In Vitro Techniques; D007700:Kinetics; D008024:Ligands; D009682:Magnetic Resonance Spectroscopy; D037241:Mannose-Binding Lectins; D008766:Methylmannosides / Q000737:chemistry / Q000378:metabolism; D008956:Models, Chemical; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014867:Water / Q000737:chemistry / Q000378:metabolism
chemical_list: D002241:Carbohydrates; D002352:Carrier Proteins; D008024:Ligands; D037241:Mannose-Binding Lectins; D008766:Methylmannosides; D009844:Oligosaccharides; D011506:Proteins; D011994:Recombinant Proteins; D014867:Water; C008466:methylmannoside