Cloning, expression, and mapping of allergenic determinants of alphaS1-casein, a major cow's milk allergen.

Abstract

Milk is one of the first components introduced into human diet. It also represents one of the first allergen sources, which induces IgE-mediated allergies in childhood ranging from gastrointestinal, skin, and respiratory manifestations to severe life-threatening manifestations, such as anaphylaxis. Here we isolated a cDNA coding for a major cow's milk allergen, alphaS1-casein, from a bovine mammary gland cDNA library with allergic patients' IgE Abs. Recombinant alphaS1-casein was expressed in Escherichia coli, purified, and characterized by circular dichroism as a folded protein. IgE epitopes of alphaS1-casein were determined with recombinant fragments and synthetic peptides spanning the alphaS1-casein sequence using microarrayed components and sera from 66 cow's milk-sensitized patients. The allergenic activity of ralphaS1-casein and the alphaS1-casein-derived peptides was determined using rat basophil leukemia cells transfected with human FcepsilonRI, which had been loaded with the patients' serum IgE. Our results demonstrate that ralphaS1-casein as well as alphaS1-casein-derived peptides exhibit IgE reactivity, but mainly the intact ralphaS1-casein induced strong basophil degranulation. These results suggest that primarily intact alphaS1-casein or larger IgE-reactive portions thereof are responsible for IgE-mediated symptoms of food allergy. Recombinant alphaS1-casein as well as alphaS1-casein-derived peptides may be used in clinical studies to further explore pathomechanisms of food allergy as well as for the development of new diagnostic and therapeutic strategies for milk allergy.

• Authors: Ulrike Schulmeister, Heidrun Hochwallner, Ines Swoboda, Margarete Focke-Tejkl, Beate Geller, Mats Nystrand, Annika Härlin, Josef Thalhamer, Sandra Scheiblhofer, Walter Keller, Bodo Niggemann, Santiago Quirce, Christoph Ebner, Adriano Mari, Gabrielle Pauli, Udo Herz, Rudolf Valenta, Susanne Spitzauer
• Journal: Journal of immunology (Baltimore, Md. : 1950) (J Immunol) Vol. 182 Issue 11 Pg. 7019-29 (Jun 01 2009) ISSN: 1550-6606 [Electronic] United States
• PMID: 19454699 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Allergens
• Caseins
• DNA, Complementary
• Epitopes
• Peptide Fragments
• Receptors, IgE
• Immunoglobulin E

Topics

• Allergens (immunology)
• Animals
• Basophils (physiology)
• Caseins (immunology)
• Cattle
• Cell Degranulation
• Cell Line, Tumor
• Cloning, Molecular
• DNA, Complementary
• Epitope Mapping
• Epitopes (genetics, immunology)
• Humans
• Immunoglobulin E
• Milk (immunology)
• Milk Hypersensitivity (immunology)
• Peptide Fragments (chemical synthesis, immunology)
• Rats
• Receptors, IgE