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Hypoallergenic mutants of the Timothy grass pollen allergen Phl p 5 generated by proline mutations.
Wald, Martin; Kahlert, Helga; Reese, Gerald; Krontal, Nina; Zafred, Domen; Keller, Walter; Cromwell, Oliver; Fiebig, Helmut; Nandy, Andreas.
Afiliação
  • Wald M; Research and Development Division, Allergopharma Joachim Ganzer KG, Reinbek, Germany. martin.wald@allergopharma.de
Int Arch Allergy Immunol ; 159(2): 130-42, 2012.
Article em En | MEDLINE | ID: mdl-22653364
ABSTRACT

BACKGROUND:

Phl p 5 is a major allergen of Timothy grass (Phleum pratense). A recombinant native Phl p 5 has already been used in clinical trials of allergen-specific immunotherapy as a component of a cocktail of allergens. Recombinant hypoallergenic allergens should further improve the treatment by reducing the risk of anaphylactic reactions at an increased therapeutic dosage. Native Phl p 5 is formed by α-helical regions separated by regions containing prolines. In order to generate hypoallergenic mutants, we studied the effect of proline mutations in single and multiple regions.

METHODS:

All mutants were analyzed by IgE inhibition assays and size exclusion chromatography with on-line mass determination. Selected mutants were additionally analyzed by field-flow fractionation, dynamic light scattering, circular dichroism spectroscopy, basophil activation and T-cell proliferation assays.

RESULTS:

Variants lacking prolines in a single region were obtained as soluble monomers. Six of eight molecules showed a slightly reduced IgE-binding capacity. Mutants carrying proline deletions in multiple regions formed monomers, dimers or insoluble aggregates. The mutant MPV.7 with five proline deletions and a substitution of proline 211 to leucine is monomeric, shows a strongly diminished IgE binding and maintains T-cell reactivity. The hydrodynamic radius and the content of the α-helical structure of MPV.7 are well comparable with the wild-type allergen.

CONCLUSIONS:

The hypoallergenic Phl p 5 variant MPV.7 combines multiple proline deletions with a substitution of proline 211 to leucine and meets basic demands for a pharmaceutical application. MPV.7 is a promising candidate for grass pollen immunotherapy with a cocktail of recombinant hypoallergens.
Assuntos

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Proteínas de Plantas / Pólen / Alérgenos Limite: Adult / Aged / Female / Humans / Male / Middle aged Idioma: En Ano de publicação: 2012 Tipo de documento: Article

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Proteínas de Plantas / Pólen / Alérgenos Limite: Adult / Aged / Female / Humans / Male / Middle aged Idioma: En Ano de publicação: 2012 Tipo de documento: Article