Sarcoplasmic calcium-binding protein is an EF-hand-type protein identified as a new shrimp allergen

Abstract

Background: Shellfish allergy is a long-lasting disorder usually persisting throughout life. Despite its high prevalence, there is limited information about allergenic shrimp proteins.

Objective: Characterization of shrimp allergens.

Methods: Fifty-two adults and children with a history of immediate allergic reactions to shrimp and elevated serum IgE to shrimp were selected for this study. Tryptic digests from a 20-kd IgE-binding protein were analyzed by LC-MS/MS, identifying the protein as a sarcoplasmic-calcium-binding protein. cDNA encoding sarcoplasmic calcium-binding protein (SCP) from a shrimp cDNA library (Litopenaeus vannamei) was amplified by PCR, cloned into an expression vector, and sequenced. Recombinant SCP was tested with patients' sera. ELISA inhibition experiments determined the fraction of total shrimp IgE recognizing SCP. A functional assay with a rat basophilic leukemia cell line was used to determine the capacity for mediator release induced by SCP.

Results: Immunoblotting demonstrated IgE binding by 31 of 52 (59.6%) of the sera to a 20-kd shrimp protein. The protein was identified as a SCP. Amplified cDNA encoding SCP was isolated and sequenced. Open reading frame translation provided the complete amino acid sequence of shrimp SCP. Recombinant SCP was recognized by serum IgE from 20 of 52 (38.4%) subjects, of whom 17 of 20 (85%) were children. ELISA inhibition of pooled sera IgE reactivity to BS extract using recombinant SCP was significant (as high as 79%). For some subjects, mediator release induced by recombinant SCP was higher than that induced by recombinant tropomyosin.

Conclusion: We have identified and cloned a new shrimp allergen, Lit v 4.0101, an SCP, which appears to be of particular importance in the pediatric population.