J Korean Med Sci.  2006 Aug;21(4):652-655. 10.3346/jkms.2006.21.4.652.

Antigen-binding Characteristics of Circulating IgG Autoantibodies to Cytokeratin 18 Protein in Patients with Nonallergic Asthma

• Yim H
• Kim JE
• Shin JY
• Ye YM
• Park HS
• Nahm DH

• KMID: 2157816
• DOI: http://doi.org/10.3346/jkms.2006.21.4.652

Abstract

Cytokeratin 18 (CK18) protein was identified as an airway epithelial cell autoantigen associated with nonallergic asthma. Cleavage of CK18 protein by caspase-3 is a marker of early apoptosis in epithelial cells. It has been shown that the expression of active caspase-3 was increased in bronchial epithelial cells of asthmatic patients, when compared with healthy controls. To investigate the antigen-binding characteristics of IgG autoantibodies to CK18 protein in nonallergic asthma, the bindings of IgG autoantibodies to the fragments of CK18 protein cleaved by caspase-3 were analyzed by Western blot using serum samples from three patients with nonallergic asthma. Recombinant human CK18 protein was treated by caspase-3 and cleaved into N-terminal fragment (1-397 amino acids) and C-terminal fragment (398-430 amino acids). The binding capacity of IgG autoantibodies to N-terminal fragment of CK18 was maintained in one patient and reduced in other two patients. IgG autoantibodies from all three patients did not bind to C-terminal fragment of CK 18. In conclusion, IgG autoantibodies to CK18 protein from patients with nonallergic asthma seems to preferentially bind to the whole molecule of CK18 protein and their antigen-binding characteristics were heterogeneous among the patients with nonallergic asthma.

Keyword

Asthma; Autoantibodies; Keratin; caspase-3

MeSH Terms

Protein Binding
Peptide Fragments/immunology/metabolism
Male
Keratins/chemistry/*immunology/metabolism
Immunoglobulin G/blood/*immunology/metabolism
Hydrolysis
Humans
Female
Epitopes/immunology/metabolism
Caspases/metabolism
Caspase 3
Blotting, Western
Autoantibodies/blood/*immunology/metabolism
Asthma/*blood/immunology
Antigen-Antibody Reactions
Antibodies, Monoclonal/immunology/metabolism
Aged
Adult

Figure

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  Fig. 1 Cleavage of cytokeratin 18 protein by caspase-3 at the sequence 394DALD/S (arrow). Cleavage of 49-kDa cytokeratin 18 protein (A) by caspase-3 results in 45-kDa fragment (B) and 4-kDa fragment (C).

• 

  Fig. 2 Protein stain of cytokeratin 18, caspase-3, and fragments of cytokeratin 18 treated by caspase-3. 0.25 µg of caspase-3 only (lane 1), 0.25 µg of cytokeratin 18 treated by 0.05 µg of caspase-3 (lane 2), molecular weight marker (lane 3), 0.25 µg of cytokeratin 18 only (lane 4), and 0.25 µg of cytokeratin 18 treated by 0.25 µg of caspase-3 (lane 5).

• 

  Fig. 3 Binding of IgG autoantibodies from three patients with nonallergic asthma to the fragments of cytokeratin 18 protein treated by caspase-3. Lane 1, 3, 5, and 7: 0.2 µg of cytokeratin 18 protein not treated by caspase. Lane 2, 4, 6, and 8: 0.2 µg of cytokeratin 18 protein treated by 0.2 µg of caspase-3. mAb: mouse monoclonal antibody to human cytokeratin 18 (clone no. CY-90, Sigma Co., St. Louis, MO, U.S.A.) known to recognize an epitope located in the region of 312-356 amino acids of cytokeratin 18 protein.

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