Journal
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
Volume 58, Issue 24, Pages 12964-12969Publisher
AMER CHEMICAL SOC
DOI: 10.1021/jf103316h
Keywords
Dasyatis akajel; parvalbumin; purification; characterization
Funding
- National Natural Scientific Foundation of China [20872049, 30871947]
- Natural Scientific Foundations of Fujian Province [2008J0067, 2010J01044]
- Jimei University [2010A005]
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Fish has received increasing attention because it induces IgE-meidated food allergy. Parvalbumin (PV) represents the major allergen of fish, and IgE cross-reactivity to PV in various teleost fish species has been shown, while little information is available about allergens in elesmobranch fish. In this study, two PV isoforms (named as PV-I and PV-II) from red stingray (Dasyatis akajei) were purified to homogeneity by a series of procedures including ammonium sulfate precipitation and column chromatographies of DEAE-Sepharose and Sephacryl S-200. Purified PVs revealed a single band on tricine sodium dodecyl sulfate polyacryalmide gel electrophoresis. The molecular masses of PV-I and PV-II were 12.29 and 11.95 kDa, respectively, as determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Western blot using antifrog PV monoclonal antibody (PARV-19) showed positive reactions to the two proteins, confirming that they were PVs, although their immunological reactivities were weaker than those of PV from silver carp. The N-terminal amino acid sequence of PV-I was determined, and comparison with PVs from other fish species showed low homology between teleost and elasmobranch fish. The isoelectric points of PV-I and PV-I1 were 5.4 and 5.0, respectively, as determined by two-dimensional electrophoresis (2-DE), suggesting that both isoforms belong to the a-group. IgE immunoblotting analysis showed that sera from fish-allergic patients reacted to both PV-I and PV-II from red stingray. Thermal stability revealed that PV-I easily formed oligomers than PV-II, which might contribute to the maintenance of its allerginicity during heat processing.
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