Non-irradiated WGA was separated

by RP-HPLC into only one peak. The chromatography analysis revealed changes after exposure at 1 kGy, as indicated Selleckchem Tariquidar in Fig. 1c. The appearance of a previous peak (Fig. 1c – arrow) to the main peak indicates partial fragmentation of the WGA at this dose. Conformational stability of WGA was investigated using fluorescence and CD spectroscopy. The shift in tryptophan fluorescence intensity and ellipticity at ∼225 nm were observed with increasing doses (Fig. 1d and Fig. 2b). At a dose of 10 kGy, the protein possibly unfolds into non-native states that are prone to aggregation. Intense fluorescence due to bis-ANS bound to the WGA was observed at 10 and 25 kGy while 1 kGy shows significantly less binding (Fig. 2a). Oligomerisation yields the basis for the multivalency necessary for typical lectin activities (Sharon & Lis, 1993). Therefore, any perturbation in protein structure that may affect the intrinsic activity after irradiation must be clarified. The positive band centred Selleckchem HSP inhibitor at ∼225 nm in the far-UV CD spectra of WGA is characteristic of cystine residues immersed in an asymmetric environment (Drenth, Low, Richardson, & Wright, 1980). Its relatively elevated intensity is due to the high density of disulphide bridges, as well as the lack of secondary-structure

repetitive elements. Radiation damage to sulphur-containing amino acids has been reported (Xu & Chance, 2005). This particular effect on disulphide bridges was observed in WGA and suggests that irradiation does not only compromise the dimeric structure but also produces a mixture of partially unfolded species at various stages of unfolding and large amorphous aggregates, after low and high doses of radiation, respectively. Such events were proven by the decrease of intrinsic fluorescence 5-Fluoracil ic50 and high binding of bis-ANS to amorphous aggregates. The current understanding about allergenicity of a plant food protein is determined by a sum of factors, including its abundance, the stability

to processing and digestion and the protective effect of food matrix (Breiteneder & Mills, 2005b). One aspect of food allergens that remains to be elucidated is the influence of the food matrix on the immune responses to food proteins. It has been hypothesised that the food body, consisting of fats, carbohydrates, and other proteins, may affect allergenic potential of proteins (Van Wijk et al., 2005). The main problem behind the conformational changes of proteins is that these are not always perceived by the general methods of analysis, which complicates the structural analysis if the food matrix is involved. However, these considerations should be investigated to clarify the contribution of food matrix to immune responses against irradiated food allergens.