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.
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.


CONCLUSIONS
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.
Fujimura
T
T
Department of Molecular Biotechnology, Graduate School of Advanced Sciences of Matter, Hiroshima University, Kagamiyama, Higashi-Hiroshima, Japan.
Shigeta
S
S
Suwa
T
T
Kawamoto
S
S
Aki
T
T
Masubuchi
M
M
Hayashi
T
T
Hide
M
M
Ono
K
K
D000485:Allergens / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D000937:Antigen-Antibody Reactions / Q000187:drug effects; D052179:Antigens, Plant; D001483:Base Sequence; D001667:Binding, Competitive; D003001:Cloning, Molecular; D003429:Cross Reactions; D032485:Cryptomeria; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007073:Immunoglobulin E / Q000378:metabolism; D020315:Latex Hypersensitivity / Q000276:immunology; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D011058:Pollen; D016415:Sequence Alignment; D017385:Sequence Homology
D000485:Allergens; D052179:Antigens, Plant; C501259:CJP-4 protein, Cryptomeria japonica; D010940:Plant Proteins; D007073:Immunoglobulin E