CHARACTERIZATION OF MAJOR ALLERGENS OF MACROBRACHIUM ROSENBERGII (GIANT RIVER PRAWN) BY ALLERGENOMIC APPROACH AND THEIR THERMOSTABILITY PROPERTIES

Prawn allergy is a common cause of allergic reactions in countries where crustacean is a famous seafood. Macrobrachium rosenbergii (giant river prawn) is declared as a local major food allergens, causes severe symptoms like asthma and anaphylactic shock. Therefore, the goal concerning this research is to identify the allergenicity of heat treated of M. rosenbergii. Prawn extracts have been prepared using fresh raw prawn and three heat-treated prawn flesh (boiled, steamed, fried), then afterward analyzed by the usage of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in conformity with their protein profiles. Allergenic proteins were identified by means of immunoblotting by the use of sera from 30 prawn-allergic patients. The identities of selective major allergens were then determined by mass-spectrometry analysis. The raw prawn has 27 protein fractions between 6 to 207 kDa, while the heat-treated prawns have reduced number of protein bands. Six prominent bands at 72, 65, 48, 38, 36, and 30 kDa were considered as the major allergens of raw extract. Meanwhile, after heat remedies applied, most of the IgE-binding bands were disappeared except for three major allergens at 38, 36 or 30 kDa which were still remain to be seen, suggesting as highly thermostable allergens. Among heat-treated prawns, steamed and boiled elicited more allergenic bands, contrast to fried prawn extract. Mass spectrometry analysis identified two selected major allergens at 36 and 48 kDa as tropomyosin and arginine kinase, respectively. As a conclusion, this study indicated that both thermostable and thermolabile proteins from M. rosenbergii were allergenic. Tropomyosin and arginine kinase were identified as the most important major allergens. Thus, the knowledge on thermostability of prawn allergens are crucial for improving diagnosis and management of prawn allergic patients in this county.

IDENTIFICATION OF TROPOMYOSIN AS THE MAJOR ALLERGEN OF BLACK TIGER PRAWN (PENAEUS MONODON) BY AN ALLERGENOMIC APPROACH

Shellfish has been recognized as one of the leading causes of food allergy in both adults and children in Asia Pacific region. In Malaysia, black tiger prawn (Penaeus monodon) is among the most widely consumed species. Our previous studies have successfully identified several major allergens including a thermostable protein of 36 kDa. Thus the aim of this study was to identify the 36 kDa major allergen by an allergenomic approach. Protein extracts of raw prown were prepared and resolved by 2-dimensional electrophoresis (2-DE). Immunoblotting was then performed using sera from patients with prawn allergy. Selected spot from 2-DE was then excised, digested and analyzed by mass spectrometry. The 2-DE profile of the extract revealed approximately 100 protein spots between pH of ~4 to 10 and the size range between (<10 to 250 kDa). The 2-DE immunoblotting has detected numerous IgE-binding spots at 36 kDa. Mass spectrometry analysis of the major IgE-binding spot (spot 1a) has identified the 36 kDa spot as tropomyosin. Our findings indicated that tropomyosin play a major role in allergic reaction to black tiger prawn among local patients with prawn allergy, and should be included in diagnostics and therapeutic strategies of this allergy.