Methods: The proteomic profile was studied using a label-free shotgun proteomics approach. As the genome sequence of macadamia nut allergens is not available, homologies to other known allergens and affiliations to protein families were determined. The results from the allergenomic screening method were used to predict potential allergenic proteins and cross-reactivity with other plants particularly. The molecular weight distribution of proteins was determined by gel electrophoresis. Following in-gel digestion with trypsin, proteins were subjected to liquid chromatography coupled tandem mass spectrometry. Based on the shared peptide evidence, the identified proteins were clustered and the allergenic proteins were identified. Immunoreactive proteins were identified by immunoblotting with three patient sera confirmed to exhibit IgE-mediated reaction.
Results: Peptides matched to the sequences of 21 allergenic proteins belonging to different protein families such as seed storage proteins (conglutins and vicilins), rubber elongation factor proteins, phosphate binding proteins and detoxifying methylglyoxalases were identified. This included peptide sequence homologies to 5 conglutins, which are known allergens from Lupin angustifolius.
Conclusion: Allergenic proteins were confirmed to be seed storage proteins belonging to 11S, 7S and 2S proteins. Significant number of peptide sequence homologies to conglutins from Lupin angustifolius was observed, suggesting that there may be cross-reaction between macadamia and lupin allergens