Fast and efficient characterization of an anti-gliadin monoclonal antibody epitope related to celiac disease using resin-bound peptides

TY - JOUR

T1 - Fast and efficient characterization of an anti-gliadin monoclonal antibody epitope related to celiac disease using resin-bound peptides

AU - Petersen, Nicole Hartwig

AU - Hansen, Paul Robert

AU - Houen, Gunnar

PY - 2011/2/28

Y1 - 2011/2/28

N2 - Celiac disease is a permanent intolerance to gluten. A strong indication for celiac disease is the presence of antibodies specific for gliadin, a main component of gluten. Using a deamidated immunogenic gliadin fragment, corresponding to amino acid residues 58-73 (LQPFPQPQLPYPQPQ) of gliadin a monoclonal anti-gliadin antibody has previously been generated.In this study we present an alternative approach for fast and efficient epitope mapping of the monoclonal anti-gliadin antibody, using gliadin peptide (Lys57)-Glu65-[gliadin 58-73] as template. This approach involves generation of a number of N-terminally truncated peptide analogues in one synthesis, and the use of these resin-bound peptides for rapid screening, in order to identify the minimal peptide length required for antibody recognition. The peptides were assembled on a TentaGel resin equipped with an acid-stable HMBA linker, which allows cleavage of the side-chain protecting groups but leaves the peptide attached to the resin. If required, the peptides may be cleaved from the support under mild basic conditions.Using this approach and competitive inhibition assays we identified the immunodominant epitope as the PELPYPQPQ sequence, which was located in the C-terminal end of the gliadin peptide. Our results show that the N-terminal proline residue and the C-terminal glutamine residues are essential for antibody recognition in addition to the deamidated glutamine residue.

AB - Celiac disease is a permanent intolerance to gluten. A strong indication for celiac disease is the presence of antibodies specific for gliadin, a main component of gluten. Using a deamidated immunogenic gliadin fragment, corresponding to amino acid residues 58-73 (LQPFPQPQLPYPQPQ) of gliadin a monoclonal anti-gliadin antibody has previously been generated.In this study we present an alternative approach for fast and efficient epitope mapping of the monoclonal anti-gliadin antibody, using gliadin peptide (Lys57)-Glu65-[gliadin 58-73] as template. This approach involves generation of a number of N-terminally truncated peptide analogues in one synthesis, and the use of these resin-bound peptides for rapid screening, in order to identify the minimal peptide length required for antibody recognition. The peptides were assembled on a TentaGel resin equipped with an acid-stable HMBA linker, which allows cleavage of the side-chain protecting groups but leaves the peptide attached to the resin. If required, the peptides may be cleaved from the support under mild basic conditions.Using this approach and competitive inhibition assays we identified the immunodominant epitope as the PELPYPQPQ sequence, which was located in the C-terminal end of the gliadin peptide. Our results show that the N-terminal proline residue and the C-terminal glutamine residues are essential for antibody recognition in addition to the deamidated glutamine residue.

U2 - 10.1016/j.jim.2010.12.019

DO - 10.1016/j.jim.2010.12.019

M3 - Journal article

C2 - 21195715

SN - 0022-1759

VL - 365

SP - 174

EP - 182

JO - Journal of Immunological Methods

JF - Journal of Immunological Methods

IS - 1-2

ER -