A Computational Study of B-Cell Epitopes of Macadamia (Macadamia Integrifolia) Allergens and Identication of its IgE Binding Residues

In contemporary research, biological computational tools have emerged to play a pivotal role in facilitating both cost and time-ecient research. One such domain is addressing the prevailing food allergy issues, where these computational tools have been proven of vital importance. The present study is done as part of an internship at Jozbiz technologies. In the present study, we discuss the identication of IgE(Immunoglobulin E) binding allergy-causing B-Cell epitopes of macadamia (Macadamia integrifolia) allergens, namely AMP23_MACIN, AMP1_MACIN, AMP21_MACIN, MATK_MACIN and 11S1_MACIN. Using seven web servers (ABCPred, ElliPro, BepiPred 1.0b, BcePred, BCPred, CBTOPE and Disco Tope 2.0) twenty-one epitopes and seventeen conformational epitopes were predicted in the present study. The predicted epitopes are analysed in terms of residues having hydrophilicity, polar nature and having exposed surfaces. The unavailable 3-d structure of proteins was developed by homology modelling. Cross-reactivity of Macadamia integrifolia with other food items has also been listed.


Introduction
In the growing age of digital advancement and fast-paced life, things are getting personalised with an increase in its speci city. Every day the eld of computational biology is also making advancements in easing the experimenting in wet labs and increasing the pace of development of the desired products. With the boon of in silico methods advancements, this study aims to identify and list the B-cell epitopes of macadamia and its IgE binding residues.
Before moving further with this it is important to understand the concept of allergy. Allergy is an immune response that occurs when any foreign material gets introduced into our bodies. These are generally perceived as harmful to organisms. Several common symptoms of allergy are rashes, swollen bronchi, stomach aches and anaphylaxis which might be fatal. Some common allergens are animal dander, pollen, dust and certain foods too. Food allergens can be categorized into two parts a)Immunoglobulin E (IgE) mediated and b) Non-IgE mediated.
Wheat allergy is an IgE mediated allergy where the antibody IgE is produced by the human immune system in response to certain allergens present in macadamia. the person having IgE mediated allergy will also be allergic to foods with a similar amino acid sequence as the macadamia.
Macadamia the allergen for the study is from the family of nuts and has its bene ts as well as certain specialities. Scienti cally known as Macadamia integrifolia is indigenous to Australia with a shelf life of a few months. It is widely used in delicacies. It's not only a decorative food but also has great nutritional bene ts too. It is rich in vitamin B6 and antioxidants to get rid of free radicals. Regular intake of macadamia nuts lowers the risk of metabolic syndromes, diabetes and aids in weight loss too. The avonoids and tocotrienols present in them are considered anti-cancer properties.
The macadamia nuts are not always a blessing to all. A certain population has shown an allergic response to it. The cases reporting macadamia nut allergy are quite low. In 2000, two allergy cases were reported and later the patients were tested in the clinic [2,7]. The skin prick test upon induced allergy proved the cause of it being the macadamia nuts. Along with the reaction on the skin the other effects include decreased gut health with abdominal cramps and vomiting in some cases.
The manually curated proteins namely AMP23_MACIN, AMP1_MACIN, AMP21_MACIN, MATK_MACIN and 11S1_MACIN from the UniProt database are used for the identi cation of the epitopes. These are processed using web servers that work on different principles. The epitopes identi ed are linear and conformational.

Materials And Methods
The Uniprot database(https://www.uniprot.org/) is used in retrieving the different amino acid sequences of macadamia nuts namely AMP23_MACIN, AMP1_MACIN, AMP21_MACIN, MATK_MACIN and 11S1_MACIN respectively. Several web servers are benchmarked for the prediction of linear and conformational epitopes. These are ABCpred [10], Bepipred [4], CBTope [1], DiscoTope2.0, Bcepred [9] and Ellipro[8]. These web servers work on principles of binding require the 3-d model. The unavailability of structures was resolved using the modelling server. The modelled and edited structure of the AMP1_MACIN is validated using the Rampage web servers. The residues are found to be in favourable regions making it an e cient model for use and refer to the Fig. 1.   The predicted linear and conformational epitopes are listed below. The use of computational tools gives the in-silico results which are asked to be proved using in vivo and in vitro experiments. These in silico results aid as the rst hand-processed data. The linear and the conformational epitopes were assessed by consensus for the most potent epitopes listed in Table III. The physio-chemical properties of the predicted continuous/linear epitopes The physicochemical properties also give an interesting insight into the binding of the Ab molecules. The hydrophilicity is proportional to the power of IgE binding.

Conclusion
Foods are not always good for the body. Some foods are perceived as foreign material and our immune system reacts to it causing an allergic reaction. The study is about the in silico identi cation of the potent epitopes and their cross-reactivities. For this experiment web servers like ABC pred, Bepipred, CBTope, DiscoTope2.0, Bcepred and Ellipro were used. The linear and conformational epitopes were recorded. The macadamia(Macadamia integrifolia) proteins used were manually curated namely AMP23_MACIN, AMP1_MACIN, AMP21_MACIN, MATK_MACIN and 11S1_MACIN.