1155 Novel Transcriptomic and Immunoproteotomic Approaches in Identifying Cross-Reactive Allergens Between Crustacean and Molluscs

Wednesday, 14 October 2015
Hall D1 Foyer (Floor 3) (Coex Convention Center)

Andreas/Ludwig Lopata, PhD , Cell & Molecular Biology, James Cook University, Townsville, Australia

Kyall Zenger, PhD , College of Marine & Environmental Sciences, James Cook University, Townsville, Australia

Sandip Kamath, PhD , Cell & Molecular Biology, James Cook University, Townsville, Australia

Roni Nugraha , Cell & Molecular Biology, James Cook University, Townsville, Australia

Background: Utilizing crude protein extracts is currently inadequate to identify allergenic proteins in shellfish. Currently, only three proteins – tropomyosin, arginine kinase, and paramyosin – have been fully identified and characterized as potential cross-reactive allergens between crustacean and mollusks. Bioinformatics analysis could provide a powerful and versatile tool for the in-depth molecular characterization of biological species with unknown allergenicity. Bioinformatics approaches have recently been used to identify putative allergens in rice, chickpea and Johnson grass pollen. Therefore this study was aimed to identify putative and cross-reactive allergens using in-silico analysis of the transcriptome from the mollusk Pacific oyster (Crassostrea gigas).

Methods: Several allergen sequences related to shellfish allergy including from Black tiger prawn, lobster and abalone were documented from different databases. TBLASTN analysis was performed on these sequences against the genomic database of Pacific oyster to retrieve putative and cross-reactive allergen sequence. The retrieved sequences were used to generate phylogenetic trees using the Neighbor joining method in Mega6 software, after association using the MUSCLE multiple alignment program. Following this, structure model of the proteins were built using Chimera 1.9 program. In addition whole protein extracts where analysed by mass spectroscopy as well as IgE binding protein from shellfish allergic patients.

Result: Based on amino acid sequence similarity, cross-reactive allergens and several putative allergen genes were identified after detailed in silicoanalysis of the genomic data of Pacific oyster. Some of the putative oyster allergens demonstrated the potential to cross-react to crustacean allergens, based on conserved domains and similar structural features. This method also revealed the presence of various isoforms of the oyster allergens. The phylogenetic trees of six different allergens demonstrate that the Pacific oyster clusters with the corresponding allergens from other mollusk, however is grouped distinct separately from crustacean allergens.

Conclusion: Bioinformatics approaches reveal several putative and cross reactive allergens that have not been fully elucidated using less sensitive immuno-chemical IgE-based methods. These findings are of importance for the development of specific allergy diagnostics for shellfish allergens. In-silico approaches, in combination with recombinant allergen generation, will be an eminent method for the comprehensive assesment of allergenicity and opens an additional path for efficient allergy diagnostics and immunotherapeutics.