10] Nuclear magnetic resonance for studying peptide — Antibody complexes by transferred nuclear overhauser effect difference spectroscopy

Abstract

This chapter describes an approach to study ligand–protein interaction using two-dimensional (2D) transferred nuclear Overhauser effect (NOE) difference spectroscopy. The methodology developed in laboratory allows obtaining detailed information concerning the interactions among residues in a peptide antigen and residues in the combining site of a monoclonal antibody. Transferred NOE difference spectroscopy, in conjunction with the specific deuteration of the antibody, provides parameters that give information about distances among the above residues. Unfortunately, the number of NOE cross-peaks is not sufficient to calculate an unequivocal structure for the bound antigen. The comparison of nuclear magnetic resonance (NMR) data with calculated models allows to examine the validity of a given model and to exclude bad models that result from choosing improper segments from the known three-dimensional structures of other antibodies. In all cases to date, information forthcoming from such studies requires comparison with crystal structures to rationalize interactions that are apparent in the NMR spectra.