Characterization and Structural Analysis of Bermuda Grass Pollen Allergy Protein BG60

• Main Authors: Donna Y. Lee, 李雅婷
• Other Authors: Shwu-Hey Liaw
• Format: Others
• Language: zh-TW
• Published: 1999
• Online Access: http://ndltd.ncl.edu.tw/handle/02870645162089551608

碩士 === 國立臺灣大學 === 分子醫學研究所 === 87 === BG60, having an apparent molecular weight of 60 kD, is one of the major allergens in Bermuda grass. SDS-PAGE analysis revealed that BG60 contained at least 3 components with slight differences in molecular weight (designated L and S for large and small forms respectively). The peptide sequences show that L and S are isomers. Database searching with the peptide sequences revealed high similarity to a plant reticuline oxidase (BBE), especially a putative FAD binding motif. BG60 exhibited a fluorescence emission maximum at 450 nm, and denaturation BG60 failed to eliminate the auto-fluorescence. These suggest that BG60 is a putative FAD covalent linked flavoprotein. Prediction of the secondary structure by CD suggested thatβ-sheet is the major secondary structure of BG60. Limited proteolysis suggested that L consists of an N-terminal segment, a 33 kD central domain, and a 22 kD C-terminal domain, whereas S lacks the N-terminal segment. The lower protease susceptibility, the higher melting temperature, and the preference for crystal growth of L forms than S forms implied the importance of the N-terminal segment in the structural stabilization. Green crystals of BG60 were obtained in 30% PEG4K and 25% isopropanol. The crystals diffract beyond 2 A resolution and belong to a tetragonal space group (P422) with the unit cell dimensions a = b = 86 A, c = 310 A. There are two monomers in an asymmetric unit. It still needs more efforts to find the heavy atom derivative to resolve the three dimensional structure of BG60.