title: Visualization of clustered IgE epitopes on alpha-lactalbumin.
abstract: BACKGROUND
  alpha-Lactalbumin (alpha-La) is a major cow's milk (CM) allergen responsible for allergic reactions in infants.
  
  
  OBJECTIVE
  We performed molecular, structural, and immunologic characterization of alpha-La.
  
  
  METHODS
  Recombinant alpha-lactalbumin (ralpha-La) was expressed in Escherichia coli, purified to homogeneity, and characterized by means of mass spectrometry and circular dichroism, and its allergenic activity was studied by using microarray technology, as well as in a basophil histamine release assay. IgE epitope mapping was performed with synthetic peptides.
  
  
  RESULTS
  According to circular dichroism analysis, ralpha-La represented a folded protein with a high thermal stability and refolding capacity. ralpha-La reacted with IgE antibodies from 57.6% of patients with CM allergy (n = 66) and induced the strongest basophil degranulation with sera from patients with CM allergy who had exhibited gastrointestinal symptoms or severe systemic reactions on CM exposure. ralpha-La contained sequential and conformational IgE epitopes. Superposition of IgE-reactive peptides onto the 3-dimensional structure of alpha-La revealed a close vicinity of the N- and C-terminal peptides within a surface-exposed patch.
  
  
  CONCLUSIONS
  ralpha-La can be used for the diagnosis of patients with severe allergic reactions to CM and serves as a paradigmatic tool for the development of therapeutic strategies for CM allergy.
authors.affiliation: Department of Medical and Chemical Laboratory Diagnostics, Medical University of Vienna, Vienna, Austria.
authors.forename: Heidrun
authors.initials: H
authors.lastname: Hochwallner
authors.forename: Ulrike
authors.initials: U
authors.lastname: Schulmeister
authors.forename: Ines
authors.initials: I
authors.lastname: Swoboda
authors.forename: Margit
authors.initials: M
authors.lastname: Focke-Tejkl
authors.forename: Vera
authors.initials: V
authors.lastname: Civaj
authors.forename: Nadja
authors.initials: N
authors.lastname: Balic
authors.forename: Mats
authors.initials: M
authors.lastname: Nystrand
authors.forename: Annika
authors.initials: A
authors.lastname: Härlin
authors.forename: Josef
authors.initials: J
authors.lastname: Thalhamer
authors.forename: Sandra
authors.initials: S
authors.lastname: Scheiblhofer
authors.forename: Walter
authors.initials: W
authors.lastname: Keller
authors.forename: Tea
authors.initials: T
authors.lastname: Pavkov
authors.forename: Domen
authors.initials: D
authors.lastname: Zafred
authors.forename: Bodo
authors.initials: B
authors.lastname: Niggemann
authors.forename: Santiago
authors.initials: S
authors.lastname: Quirce
authors.forename: Adriano
authors.initials: A
authors.lastname: Mari
authors.forename: Gabrielle
authors.initials: G
authors.lastname: Pauli
authors.forename: Christof
authors.initials: C
authors.lastname: Ebner
authors.forename: Nikolaos G
authors.initials: NG
authors.lastname: Papadopoulos
authors.forename: Udo
authors.initials: U
authors.lastname: Herz
authors.forename: Eric A F
authors.initials: EA
authors.lastname: van Tol
authors.forename: Rudolf
authors.initials: R
authors.lastname: Valenta
authors.forename: Susanne
authors.initials: S
authors.lastname: Spitzauer
mesh_terms: D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D002942:Circular Dichroism; D003001:Cloning, Molecular; D018985:Epitopes, B-Lymphocyte / Q000737:chemistry / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005240:Feasibility Studies; D006636:Histamine Release / Q000276:immunology; D006801:Humans; D007073:Immunoglobulin E / Q000097:blood / Q000276:immunology; D007768:Lactalbumin / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D013058:Mass Spectrometry; D046228:Microarray Analysis; D016269:Milk Hypersensitivity / Q000097:blood / Q000175:diagnosis* / Q000276:immunology*; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D018985:Epitopes, B-Lymphocyte; D010446:Peptide Fragments; D011994:Recombinant Proteins; D007073:Immunoglobulin E; D007768:Lactalbumin