Isolation and partial characterization of a 46-kd allergen of Bermuda grass pollen.
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.
Wu
W C
WC
Institute of Biotechnology in Medicine, National Yang-Ming University, Taipei, Taiwan, ROC.
Tam
M F
MF
Peng
H J
HJ
Tsai
L C
LC
Chi
C W
CW
Chang
Z N
ZN
D000485:Allergens / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D052179:Antigens, Plant; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D015180:Electrophoresis, Gel, Two-Dimensional; D037102:Lectins / Q000378:metabolism; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000737:chemistry; D010504:Periodic Acid / Q000737:chemistry; D037121:Plant Lectins; D006109:Poaceae / Q000276:immunology*; D011485:Protein Binding
D000485:Allergens; D000911:Antibodies, Monoclonal; D052179:Antigens, Plant; C437644:Cyn d Bd46K allergen; D037102:Lectins; D010446:Peptide Fragments; D037121:Plant Lectins; D010504:Periodic Acid