ABSTRACT
Free-energy perturbation (FEP) simulations have been applied to a series of analogues of the natural trisaccharide epitope of Salmonella serotype B bound to a fragment of the monoclonal anti-Salmonella antibody Se155-4. This system was selected in order to assess the ability of free-energy perturbation (FEP) simulations to predict carbohydrate-protein interaction energies. The ultimate goal is to use FEP simulations to aid in the design of synthetic high affinity ligands for carbohydrate-binding proteins. The molecular dynamics (MD) simulations were performed in the explicit presence of water molecules, at room temperature. The AMBER force field, with the GLYCAM parameter set for oligosaccharides, was employed. In contrast to many modeling protocols, FEP simulations are capable of including the effects of entropy, arising from differential ligand flexibilities and solvation properties. The experimental binding affinities are all close in value, resulting in small relative free energies of binding. Many of the DeltaDeltaG values are on the order of 0-1 kcal mol(-1), making their accurate calculation particularly challenging. The simulations were shown to reasonably reproduce the known geometries of the ligands and the ligand-protein complexes. A model for the conformational behavior of the unbound antigen is proposed that is consistent with the reported NMR data. The best agreement with experiment was obtained when histidine 97H was treated as fully protonated, for which the relative binding energies were predicted to well within 1 kcal mol(-1). To our knowledge this is the first report of FEP simulations applied to an oligosaccharide-protein complex.
ABSTRACT
The conformational properties of oligosaccharides are important in determining their biological properties, such as recognition by proteins. The structural and dynamic properties of many oligosaccharides are poorly understood both because of a lack of experimental data (usually obtained from solution NMR parameters) and because of gross approximations frequently invoked in theoretical models. To characterise the oligomannose oligosaccharide Man,GlcNAc2 we have acquired a more extensive NMR data set and performed the first unrestrained molecular dynamics (MD) simulation in water of this large oligosaccharide (employing the GLYCAM_93 parameter set with the AMBER force field). Good agreement is seen between the computed dynamics data and the results of both an isolated spin pair (ISPA) analysis of short mixing time NOE data and NOE build-up curves for mixing times from 100 to 2000 ms. The number of experimental conformational constraints obtained in this study are in principle sufficient to fully define a rigid structure. The fact that this could not be done indicates a high degree of internal flexibility and/or the presence of multiple conformations about the glycosidic linkages. Independently, the same conclusions are reached from an analysis of the MD results. In addition, the theoretical results allow the overall topology of the molecule and its intra-molecular and solvent-mediated hydrogen bonding pattern to be defined. Extensive re-organisation of solvent and inter-residue hydrogen bonds is shown to be required for significant conformational changes to occur, resulting in relatively long life-times for distinct glycosidic linkage conformations, despite the high local flexibility of the glycosidic linkages. This factor is also seen in the overall topology of the molecule, where the considerable internal flexibility is not translated into gross changes in structure. The control exerted by the solvent over both the flexibility and overall topology of an oligosaccharide has important implications for recognition processes and for the conformational properties of glycans attached to glycoproteins.
