title: Designing hypoallergenic derivatives for allergy treatment by means of in silico mutation and screening.
abstract: BACKGROUND
  The incidence of allergic diseases in developed countries has been increasing constantly in the last 2 decades. The only curative treatment available thus far is specific immunotherapy (SIT). The problematic side effects that can occur during SIT with allergen extracts led to the search for safer alternatives, such as recombinant hypoallergenic proteins with reduced allergenic potential and preserved T-cell immunogenicity.
  
  
  OBJECTIVE
  We created hypoallergenic variants of the allergens Bet v 1a and Phl p 5b by using in silico mutation and screening and characterized the biochemical and immunologic properties of selected mutant proteins.
  
  
  METHODS
  Knowledge-based potentials were used to estimate structural changes of the protein structures under sequence variation. IgE antibodies and their cross-linking capacity were determined by using a basophil release assay, binding of human birch pollen-specific IgE was investigated by means of ELISA, and ELISPOT assays were performed to examine the T-cell immunogenicity.
  
  
  RESULTS
  Selected mutated proteins showed significantly reduced IgE-binding and cross-linking ability but retained their T cell-stimulating properties. Immunization with the hypoallergenic mutants induced blocking antibodies against murine and human IgE epitopes.
  
  
  CONCLUSIONS
  In silico calculation and selection of mutations that render a protein hypoallergenic represents a novel and rapid tool to create candidate molecules for SIT with recombinant allergens.
authors.affiliation: Department of Molecular Biology, Division of Allergy and Immunology, University of Salzburg, Salzburg, Austria.
authors.forename: Theresa
authors.initials: T
authors.lastname: Thalhamer
authors.forename: Heidi
authors.initials: H
authors.lastname: Dobias
authors.forename: Tatjana
authors.initials: T
authors.lastname: Stepanoska
authors.forename: Martina
authors.initials: M
authors.lastname: Pröll
authors.forename: Hanno
authors.initials: H
authors.lastname: Stutz
authors.forename: Oliver
authors.initials: O
authors.lastname: Dissertori
authors.forename: Peter
authors.initials: P
authors.lastname: Lackner
authors.forename: Fatima
authors.initials: F
authors.lastname: Ferreira
authors.forename: Michael
authors.initials: M
authors.lastname: Wallner
authors.forename: Josef
authors.initials: J
authors.lastname: Thalhamer
authors.forename: Arnulf
authors.initials: A
authors.lastname: Hartl
mesh_terms: D000485:Allergens / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology; D000818:Animals; D052179:Antigens, Plant / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D003888:Desensitization, Immunologic; D015195:Drug Design; D005260:Female; D006801:Humans; D006967:Hypersensitivity / Q000188:drug therapy*; D007073:Immunoglobulin E / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D009154:Mutation; D010940:Plant Proteins / Q000235:genetics / Q000276:immunology; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology; D012260:Ribonucleases / Q000235:genetics / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology
chemical_list: D000485:Allergens; D052179:Antigens, Plant; D010940:Plant Proteins; D011994:Recombinant Proteins; D007073:Immunoglobulin E; C102306:Phl p Vb protein, Phleum pratense; D012260:Ribonucleases