Cloning, expression, and mapping of allergenic determinants of alphaS1-casein, a major cow's milk allergen.
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.
Schulmeister
Ulrike
U
Institute of Medical and Chemical Laboratory Diagnostics, Medical University of Vienna, Austria.
Hochwallner
Heidrun
H
Swoboda
Ines
I
Focke-Tejkl
Margarete
M
Geller
Beate
B
Nystrand
Mats
M
Härlin
Annika
A
Thalhamer
Josef
J
Scheiblhofer
Sandra
S
Keller
Walter
W
Niggemann
Bodo
B
Quirce
Santiago
S
Ebner
Christoph
C
Mari
Adriano
A
Pauli
Gabrielle
G
Herz
Udo
U
Valenta
Rudolf
R
Spitzauer
Susanne
S
D000485:Allergens / Q000276:immunology*; D000818:Animals; D001491:Basophils / Q000502:physiology; D002364:Caseins / Q000276:immunology*; D002417:Cattle; D015550:Cell Degranulation; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D018076:DNA, Complementary; D018604:Epitope Mapping; D000939:Epitopes / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007073:Immunoglobulin E; D008892:Milk / Q000276:immunology*; D016269:Milk Hypersensitivity / Q000276:immunology; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000276:immunology; D051381:Rats; D017455:Receptors, IgE
D000485:Allergens; D002364:Caseins; D018076:DNA, Complementary; D000939:Epitopes; D010446:Peptide Fragments; D017455:Receptors, IgE; D007073:Immunoglobulin E