Isolation and partial characterization of a 46-kD allergen of Bermuda grass pollen |
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Authors: | Wen-Chia Wu Ming F Tam Ho-Jen Peng Lai-Chin Tsai Chin-Wen Chi Zo-Nan Chang |
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Institution: | Institute of Biotechnology in Medicine, National Yang-Ming University, Taipei, Taiwan, ROC. |
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Abstract: | Cyn d Bd46K, a 46-kD component of Bermuda grass (Cynodon dactylon) pollen, had been identified as an allergenic constituent. In the present study two-dimensional (2D) gel electrophoresis illustrated the presence of five acidic isoforms in Cyn d Bd46K, and this molecule was purified by monoclonal antibody (MAb) affinity chromatography for further characterization. Using a digoxigenin-labeled lectin-binding assay, the elucidating protein was disclosed to be a glycoprotein with terminal mannose. The involvement of a carbohydrate moiety in the allergenicity and antigenicity of the elucidated molecule was demonstrated with sodium-periodate-treated Cyn d Bd46K, which reduced binding to its specific MAb and human IgE. We were unable to identify the N-terminal amino acid sequences of Cyn d Bd46K, but some internal amino acid sequences were disclosed by microsequencing some fragments cleaved by Achromobacter protease I and fractionated by reversed-phase column chromatography. The amino acid sequences of 4 identified Cyn d Bd46K internal peptide fragments were found to be 25-71% identical with that of cytochrome c oxidase III from corn grass pollen. The present study provided important information for future experiments on the molecular cloning of the elucidated allergen. |
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Keywords: | Bermuda grass pollen Cyn d Bd46K Monoclonal antibody Affinity chromatography Internal sequencing |
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