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Investigation of the Binding of a Carbohydrate‐Mimetic Peptide to its Complementary Anticarbohydrate Antibody by STD‐NMR Spectroscopy and Molecular‐Dynamics Simulations
Author(s) -
Szczepina Monica G.,
Bleile Dustin W.,
Pinto B. Mario
Publication year - 2011
Publication title -
chemistry – a european journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.687
H-Index - 242
eISSN - 1521-3765
pISSN - 0947-6539
DOI - 10.1002/chem.201100222
Subject(s) - chemistry , molecular dynamics , nuclear magnetic resonance spectroscopy , ligand (biochemistry) , proton , spectroscopy , crystallography , two dimensional nuclear magnetic resonance spectroscopy , stereochemistry , computational chemistry , physics , receptor , biochemistry , quantum mechanics
Saturation transfer difference (STD)‐NMR spectroscopy was used to probe experimentally the bioactive solution conformation of the carbohydrate mimic MDWNMHAA 1 of the O‐polysaccharide of Shigella flexneri Y when bound to its complementary antibody, mAb SYA/J6. Molecular dynamics simulations using the ZymeCAD™ Molecular Dynamics platform were also undertaken to give a more accurate picture of the conformational flexibility and the possibilities for bound ligand conformations. The ligand topology, or the dynamic epitope, was mapped with the CORCEMA‐ST (COmplete Relaxation and Conformational Exchange Matrix Analysis of Saturation Transfer) program that calculates a total matrix analysis of relaxation and exchange effects to generate predicted STD‐NMR intensities from simulation. The comparison of these predicted STD enhancements with experimental data was used to select a representative binding mode. A protocol that employed theoretical STD effects calculated at snapshots during the entire course of a molecular dynamics (MD) trajectory of the peptide bound to the Fv portion of the antibody, and not the averaged atomic positions of receptor–ligand complexes, was also examined. In addition, the R factor was calculated on the basis of STD (fit) to avoid T1 bias, and an effective R factor, R eff , was defined such that if the calculated STD (fit) for proton k was within error of the experimental STD (fit) for proton k , then that calculated STD (fit) for proton k was not included in the calculation of the R factor. This protocol was effective in deriving the antibody‐bound solution conformation of the peptide which also differed from the bound conformation determined by X‐ray crystallography; however, several discrepancies between experimental and calculated STD (fit) values were observed. The bound conformation was therefore further refined with a simulated annealing refinement protocol known as STD‐NMR intensity‐restrained CORCEMA optimization (SICO) to give a more accurate representation of the bound peptide epitope. Further optimization was required in this case, but a satisfactory correlation between experimental and calculated STD values was obtained. Attempts were also made to obtain STD enhancements with a synthetic pentasaccharide hapten, corresponding to the O‐polysaccharide, while bound to the antibody. However, unfavorable kinetics of binding in this system prevented sufficient STD build‐up, which, in turn, hindered a rigorous analysis via full STD build‐up curves.