Purification, Expression, and Characterization of Sarcoplasmic Calcium Binding Protein: A Novel Allergen of Portunus trituberculatus

Abstract

Swimming crab (Portunus trituberculatus), a crucial valuable crustacean, is a common factor causing food allergy. However, studies on allergens of P. trituberculatus are scarce. In this study, the sarcoplasmic calcium binding protein (SCP) of P. trituberculatus was expressed in Escherichia coli, purified with affinity chromatography, and the IgE-binding activity was evaluated through serological analyses. Further, the structure, physicochemical properties, and cross-reactivity were assessed via bioinformatics, immunologic, and spectroscopy techniques. The results indicated that P. trituberculatus SCP was an allergen displaying strong IgEbinding capacity, composed of 60% α-helix. It presented good immunologic and structural stability at 4−70 °C and pH 3−10, and only exhibited high IgG cross-reactivity among crustaceans, without cross-reactivity with other species tested. These results establish the foundations for further studies on SCP and are promising to promote the development of specific crustacean allergen detection and precise allergy diagnosis.

Genomics of Shrimp Allergens and Beyond

“…At the protein level, the protein length of SCP is approximately 192 AA, with a MW of 20-25 kDa. It has been reported that SCP exhibits high resistance to acidalkali conditions and heat [65]. Zhao et al [66] examined the immunological properties and structural changes of the recombinant Lit v 4 (rLit v 4) under various temperature conditions.…”

“…They demonstrated that crustacean SCP showed high sequence identities ranging from 77% to 96%. The sequence homology of crustacean SCPs with those of insects, mites (and their relative, the horseshoe crab), and mollusks was generally lower, ranging from 14% to 55% [65]. Thus far, SCP shows cross-reactivity in crab (Scy p 4), oyster (Cra a 4), and lobster (Pon l 4) (Table 3).…”

Mutation of Disulfide Bond Sites Reduces the Immunoreactivity of Cra a 4 by Changing the Structural Characteristics

“…angulata, including tropomyosin, arginine kinase, and sarcoplasmic calcium-binding protein (SCP) . SCP is considered to be a stable protein. − The study had shown that SCP can be recognized by specific IgE in sera of allergic patients after the heat treatment. , SCP from C. angulata was registered in the WHO/IUIS allergen database in our previous research, which was named Cra a 4.…”