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


CONCLUSIONS
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.
Ayuso
Rosalía
R
Division of Allergy and Immunology and the Jaffe Food Allergy Research Institute, Mount Sinai School of Medicine, One Gustave L. Levy Place, Box 1198, New York, NY 10029, USA. rosalia.ayuso@mssm.edu
Grishina
Galina
G
Ibáñez
María Dolores
MD
Blanco
Carlos
C
Carrillo
Teresa
T
Bencharitiwong
Ramon
R
Sánchez
Silvia
S
Nowak-Wegrzyn
Anna
A
Sampson
Hugh A
HA
D000293:Adolescent; D000328:Adult; D000368:Aged; D000485:Allergens / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D000595:Amino Acid Sequence; D000818:Animals; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D002460:Cell Line; D002648:Child; D002675:Child, Preschool; D003429:Cross Reactions; D020832:EF Hand Motifs; D015180:Electrophoresis, Gel, Two-Dimensional; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D006967:Hypersensitivity; D007073:Immunoglobulin E / Q000097:blood; D008875:Middle Aged; D008969:Molecular Sequence Data; D033561:Penaeidae; D011485:Protein Binding; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000737:chemistry* / Q000145:classification / Q000235:genetics; D016415:Sequence Alignment; D012758:Shellfish; D055815:Young Adult
D000485:Allergens; D002135:Calcium-Binding Proteins; D011994:Recombinant Proteins; D007073:Immunoglobulin E; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases