Molecular cloning of a class IV chitinase allergen from Japanese cedar (Cryptomeria japonica) pollen and competitive inhibition of its immunoglobulin E-binding capacity by latex C-serum

Authors

  • T. Fujimura,

    1. Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Kagamiyama, Higashi-Hiroshima, Japan,
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    • 1Present address: Research Center for Allergy and Immunology, Yokohama Institute, RIKEN (The Institute of Physical and Chemical Research), 1-7-22 Suehiro, Tsurumi, Yokohama, Kanagawa 230-0045, Japan.

  • S. Shigeta,

    1. Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Kagamiyama, Higashi-Hiroshima, Japan,
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  • T. Suwa,

    1. Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Kagamiyama, Higashi-Hiroshima, Japan,
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  • S. Kawamoto,

    1. Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Kagamiyama, Higashi-Hiroshima, Japan,
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  • T. Aki,

    1. Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Kagamiyama, Higashi-Hiroshima, Japan,
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  • M. Masubuchi,

    1. Hiroshima Bunkyo Woman's University, Hiroshima, Japan,
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  • T. Hayashi,

    1. Takanobashi Central Hospital, Hiroshima, Japan
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  • M. Hide,

    1. Graduate School of Biomedical Science, Hiroshima University, Hiroshima, Japan
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  • K. Ono

    1. Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Kagamiyama, Higashi-Hiroshima, Japan,
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Dr Seiko Shigeta, Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, 1-3-1 Kagamiyama, Higashi-Hiroshima 739-8530, Japan. E-mail: shigeta@hiroshima-u.ac.jp

Summary

Background Japanese cedar (Cryptomeria japonica) pollinosis is one of the most prevalent allergic diseases in Japan. Only three C. japonica allergens, Cry j 1, Cry j 2, and CJP-6, have been characterized. The full IgE-binding spectrum of C. japonica pollen allergens demonstrates that many allergens remain to be identified.

Objective The aim of this study was to characterize a novel allergen with a high frequency of IgE binding.

Methods The cDNA coding for a high-frequency IgE-binding protein, designated CJP-4, was cloned from the total mRNA of C. japonica pollen. The corresponding native allergen was purified by affinity precipitation with colloidal chitin and gel chromatography. The IgE-binding ability of purified native CJP-4 was characterized by ELISA and ELISA inhibition.

Results The CJP-4 cDNA encoded 281 amino acids with significant sequence homology to class IV chitinases. Purified native CJP-4, migrated as a homogeneous 34-kDa protein on SDS-PAGE, revealed endochitinase activity on native PAGE. The purified protein displayed the ability to bind IgE from all patients tested (31/31) in ELISA, whereas Cry j 1 bound to IgE at a 71% frequency (22/31). Pre-incubation with latex C-serum completely inhibited the reaction of pooled sera IgE from patients with C. japonica pollinosis and/or latex allergy to purified CJP-4.

Conclusion We identified CJP-4 as a novel and fourth C. japonica chitinase allergen with high IgE-binding frequency. The competitive IgE-binding profile between C. japonica chitinase and latex C-serum indicated that C. japonica chitinase should be an important pan-allergen in C. japonica pollen.

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