Molecular cloning and immunochemical characterization of a novel major Japanese cedar pollen allergen belonging to the aspartic protease family
| Autor: | Isao Oiso, Satoshi Rikimaru, Kazuhisa Ono, Nobukazu Onishi, Kunihiko Hashimoto, Seiji Kawamoto, Yayoi Shimada, Takaharu Hayashi, Ahmed Ragaa Nour Ibrahim, Tsunehiro Aki, Elfadil Elfadl Babiker |
|---|---|
| Rok vydání: | 2009 |
| Předmět: |
Aspartic Acid Proteases
medicine.medical_treatment Cryptomeria Immunology Blotting Western Molecular Sequence Data Molecular cloning medicine.disease_cause Japonica Antibodies Microbiology Hemoglobins Allergen Pollen Complementary DNA Catalytic Domain otorhinolaryngologic diseases medicine Immunology and Allergy Humans Protease Inhibitors Amino Acid Sequence Cloning Molecular Phylogeny Cloning Genetics Enzyme Precursors Protease biology Sequence Homology Amino Acid food and beverages Rhinitis Allergic Seasonal General Medicine Antigens Plant Hydrogen-Ion Concentration Immunoglobulin E biology.organism_classification Recombinant Proteins Biocatalysis |
| Zdroj: | International archives of allergy and immunology. 152(3) |
| ISSN: | 1423-0097 |
| Popis: | Background:Japanese cedar (Cryptomeria japonica) pollen is a major cause of seasonal pollinosis in Japan. Protease activity in the pollen grains may trigger pro-allergic responses but no such proteases have yet been identified as pollen allergens. Objectives:We report the molecular cloning and immunochemical characterization of a novel C. japonica pollen allergen belonging to the aspartic protease family. Methods:We focused on the C. japonica pollen allergen spot No. 63 (CPA63, 47.5% IgE binding frequency) on our 2-dimensional IgE immunoblot map. The internal amino acid sequences were determined using time-of-flight mass spectrometry. Full-length cpa63 cDNA was cloned by rapid amplification of cDNA ends (RACE)-PCR. Recombinant CPA63 (r-CPA63) was expressed using the baculovirus-insect cell culture system and its IgE binding capacity was analyzed by enzyme-linked immunosorbent assay (ELISA). The proteolytic activity of r-CPA63 was also assessed using a putative mature enzyme produced upon autolysis. Results: cpa63 cDNA encoded a 472 amino acid polypeptide showing about 40% sequence identity to members of the plant atypical aspartic protease family. ELISA showed that r-CPA63 was recognized by IgE antibodies in the serum of 58% (18/31) of Japanese cedar pollinosis patients. We also demonstrated an aspartic protease-like enzyme activity of the putative mature r-CPA63. Conclusions:We have identified the first plant aspartic protease allergen from Japanese cedar pollen. The availability of the CPA63 sequence and its recombinant allergen production system are useful not only for pharmaceutical applications but also for further examination of the role of protease activity in the pathogenesis of cedar pollinosis. |
| Databáze: | OpenAIRE |
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