Sayers E W, Weaver J L, Prestegard J H
Department of Pharmacology, Yale University, New Haven, CT 06510, USA.
J Biomol NMR. 1998 Aug;12(2):209-22. doi: 10.1023/a:1008220522409.
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.
