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NMR observation of interactions in the combining site region of an antibody using a spin‐labeled peptide antigen and NOESY difference spectroscopy
Author(s) -
Scherf Tali,
Hiller Reuben,
Anglister Jacob
Publication year - 1995
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.9.1.7821751
Subject(s) - two dimensional nuclear magnetic resonance spectroscopy , peptide , antigen , nuclear magnetic resonance , spin (aerodynamics) , antibody , spectroscopy , chemistry , physics , biology , immunology , quantum mechanics , thermodynamics
A spin‐labeled peptide antigen (TEMPO‐VEVPGSQHIDSQ) was used to measure NOESY difference spectra that show interactions in the binding site region of the Fab fragment of the anti‐cholera toxin peptide antibody TE33. In addition to identification of peptide‐Fab interactions and interactions within the bound peptide, these difference spectra show well‐resolved cross peaks due to interactions within the large Fab fragment (50 kDa). These difference spectra indicate that the conformational changes in the Fab upon peptide binding are confined to the combining site region of the antibody. The NOESY difference spectra of selectively deuterated Fab molecules were used in combination with HOHAHA measurements to assign the interactions to amino acid type and to identify the interactions within the Fab as either inter‐ or intraresidue interactions. The assignment of interactions within the Fab to corresponding aromatic residues in the Fab sequence was facilitated by an earlier NMR‐derived model calculated on the basis of NOE restraints on Fab‐peptide and intra‐boundpeptide distances. The new restraints on distances within the Fab, combined with the previously obtained restraints, were used to generate a refined NMR‐derived model for the TE33‐peptide complex.—Scherf, T., Hiller, R., Anglister, J. NMR observation of interactions in the combining site region of an antibody using a spin‐labeled peptide antigen and NOESY difference spectroscopy. FASEB J. 9, 120‐126 (1995)