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 acidi c isoforms in Cyn d Bd46K, and this molecule was purified by monoclonal ant ibody (MAb) affinity chromatography for further characterization. Using a d igoxigenin-labeled lectin-binding assay, the elucidating protein was disclo sed to be a glycoprotein with terminal mannose. The involvement of a carboh ydrate moiety in the allergenicity and antigenicity of the elucidated molec ule was demonstrated with sodium-periodate-treated Cyn d Bd46K, which reduc ed binding to its specific MAb and human IgE. We were unable to identify th e N-terminal amino acid sequences of Cyn d Bd46K, but some internal amino a cid sequences were disclosed by microsequencing some fragments cleaved by A chromobacter protease I and fractionated by reversed-phase column chromatog raphy. The amino acid sequences of 4 identified Cyn d Bd46K internal peptid e fragments were found to be 25-71% identical with that of cytochrome c oxi dase III from corn grass pollen. The present study provided important infor mation for future experiments on the molecular cloning of the elucidated al lergen. Copyright (C) 2001 National Science Council, ROC and S. Karger AG, Basel.