INTEREST OF ANTI-PEPTIDE ANTIBODIES DIRECTED AGAINST OMEGA-GLIADINS TO DETECT SPECIFIC SEQUENCES AND EVALUATE CONFORMATIONAL CHANGES.

s.Denery-Papini, 2M.F. Samson and 2J.C. Autran.

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INRA, 1Unité de Biochimie et de technologie des protéines, B.P. 71627, 44316 2 nantes cedex 3, France, Unité de technologie des Céréalef 2 place Viala, 34060 Montpellier cedex 1. Keys words : omega-gliadins, anti-peptide antibodies, bread wheat, durum wheat, thermal treatment. Omega-gliadins are thermostable proteins of great interest as they can be used as tracers in food products submitted to heating. We have obtained anti-peptide antibodies directed against the differents classes ofwheat prolamin s some ofwhich specifically detec tro -gliadins (Denery-Papini et al., 1994). The objectives of this study ~to explore the interest of the anti-ro-gliadin antibodies to detect sequences specific of bread wheat and to follow the influence of thennal treatment on these proteins. Material and methods : Pasta with different bread wheat contents were used as models. The y were submitted to different temperatures of drying (60, 85 or 100°C) and some of them were heated in boiling water. Antibodies directed against N-terminal peptides and to a repetitive peptide found in the central domain of ro-gliadins were used in this study. The reactivity of these antibodies against proteins contained in 50% propanol extracts were analysed by immunoblotting after SOS-PAGE and by competitive ELISA Omega-gliadin content in propanol extracts were obtained by RP-HPLC. Results : One of the anti-peptide antisera (anti-NTl-ro) directed against an ro-gliadin N-termin;il sequence found in bread wheat permitted a specific detection of this speci_es. No reac tion was observed in irnmunoblottin g with proteins extracted from 26 durum wheat semolina whereas a 60 kd component was recognized in the extracts of 36 bread wheat flours. Detection of the presence of bread wheat in pasta was possible with competitive ELISA. The influence of thermal treatement on ro-gliadin immunodetection was observed in competitive r-..)' ELISA. The anti-peptide antiserum (anti-R-gli adin) directed against _! repetitive seque nce / l?_~tte reco gn ized proteins extracted from pasta dried at high temperatu re (100°C) than those extracted from pas ta dried at 60°C. On the contrary, no difference was seen with the anti-NTl-ro antibodies. Both ··antisera detected better proteins extracted from pasta after a step ofh_eatlniJ!! boilin,g wât~; Conclusions : Anti-peptide antibodies are gene rally directed against lin!tar epitopes that are not so much affe cted by COnformational changes than CO!}formationaJ _epitopes and therefore present ·mtere st to etect and qûantify protein s in food submitted to proc ess ing. We show in this study that these antibodie s ca~ be used to observe structurf modific ations that occur in particular domains and that drying or heating differently affects N-termin al or repetitive reg ions. In addition, thanks to its narrow specificity , the anti-NTl-ro antiserum could be used to detect bread wheat additions in durum ---·wheat products.

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S. Denery-Papini , Y. Popineau, L. Quillien and M.H.V. Van Regenmortel. Immunological differentiation ofvarious gliad ins and low mol ecu lar weight subunits of g lutenin usin g ant i-peptide anti sera. Journal ofCereal Science (1994), 20, 1- 14.