Antigenic Determinants of the Bilobal Cockroach Allergen Bla g 2

Woodfolk, Judith A.; Glesner, Jill; Wright, Paul W.; Kepley, Christopher L.; Li, Mi; Himly, Martin; Muehling, Lyndsey M.; Gustchina, Alla; Wlodawer, Alexander; Chapman, Martin D.; Pomés, Anna

doi:10.1074/jbc.M115.702324

Title: Antigenic Determinants of the Bilobal Cockroach Allergen Bla g 2

Journal Article · Mon Dec 07 00:00:00 EST 2015 · Journal of Biological Chemistry

DOI:https://doi.org/10.1074/jbc.M115.702324· OSTI ID:1244723

Woodfolk, Judith A. ^[1]; Glesner, Jill ^[2]; Wright, Paul W. ^[1]; Kepley, Christopher L. ^[3]; Li, Mi ^[4]; Himly, Martin ^[5]; Muehling, Lyndsey M. ^[1]; Gustchina, Alla ^[6]; Wlodawer, Alexander ^[6]; Chapman, Martin D. ^[3]; Pomés, Anna ^[7]

Univ. of Virginia, Charlottesville, VA (United States)
INDOOR Biotechnologies, Inc., Charlottesville, VA (United States)
Univ. of North Carolina, Greensboro, NC (United States)
National Inst. of Health (NIH), Frederick, MD (United States); Frederick National Lab., MD (United States)
Univ. of Salzburg (Austria)
National Inst. of Health (NIH), Frederick, MD (United States)
Univ. of North Carolina, Greensboro, NC (United States); Indoor Biotechnologies, Inc., Charlottesville, VA (United States)

Bla g 2 is a major indoor cockroach allergen associated with the development of asthma. Antigenic determinants on Bla g 2 were analyzed by mutagenesis based on the structure of the allergen alone and in complex with monoclonal antibodies that interfere with IgE antibody binding. The structural analysis revealed mechanisms of allergen-antibody recognition through cation-π interactions. Single and multiple Bla g 2 mutants were expressed in Pichia pastoris and purified. The triple mutant K132A/K251A/F162Y showed an ~100-fold reduced capacity to bind IgE, while preserving the native molecular fold, as proven by x-ray crystallography. This mutant was still able to induce mast cell release. T-cell responses were assessed by analyzing Th1/Th2 cytokine production and the CD4+ T-cell phenotype in peripheral blood mononuclear cell cultures. Although T-cell activating capacity was similar for the KKF mutant and Bla g 2 based on CD25 expression, the KKF mutant was a weaker inducer of the Th2 cytokine IL-13. Furthermore, this mutant induced IL-10 from a non-T-cell source at higher levels that those induced by Bla g 2. Lastly, our findings demonstrate that a rational design of site-directed mutagenesis was effective in producing a mutant with only 3 amino acid substitutions that maintained the same fold as wild type Bla g 2. These residues, which were involved in IgE antibody binding, endowed Bla g 2 with a T-cell modulatory capacity. The antigenic analysis of Bla g 2 will be useful for the subsequent development of recombinant allergen vaccines.

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Research Organization:: Argonne National Lab. (ANL), Argonne, IL (United States)

Sponsoring Organization:: National Inst. of Health

Grant/Contract Number:: R01AI077653; R01AI77653-01A2S; HHSN26120080001E

OSTI ID:: 1244723

Journal Information:: Journal of Biological Chemistry, Vol. 291, Issue 5; ISSN 0021-9258

Publisher:: American Society for Biochemistry and Molecular BiologyCopyright Statement

Country of Publication:: United States

Language:: ENGLISH

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Cited by: 15 works

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59 BASIC BIOLOGICAL SCIENCES
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asthma
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immunotherapy
site-directed mutagenesis
x-ray crystallography
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cockroach allergy

Title: Antigenic Determinants of the Bilobal Cockroach Allergen Bla g 2

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