題名: A novel conformation-dependent monoclonal antibody specific to the native structure of beta-lactoglobulin and its application
作者: Chen WL;Liu WT;Yang MC;Hwang MT;Tsao JH;Mao SJT 貢獻者: Department of Biotechnology
日期: 2006-03
上傳時間: 2009-11-26T01:43:11Z 出版者: Asia University
摘要: Molten globules are thought to be general intermediates in protein folding and unfolding. beta-lactoglobulin (beta-LG) is one of the major bovine whey proteins, constituting approximately 10 to 15% of total milk proteins. We have recently identified beta-LG as a superior marker for evaluating thermally processed milk. Strand D of beta-LG participates in irreversible thermal unfolding as probed by a monoclonal antibody (mAb) specific to thermally denatured beta-LG. In the present study, we used native beta-LG as an immunogen to test the hypothesis that a specific mAb against the native beta-LG could be established. As result, a mAb (4H11E8) directed against the native structure of beta-LG was made. The antibody did not recognize the heat-denatured form of beta- LG, such as its dimer and aggregates. Immunoassay using this "native"
mAb showed that the stability of beta-LG was at temperatures < or =70 degrees C. beta-Lactoglobulin began to deteriorate between 70 and 80 degrees C over time. The denaturation was correlated with the transition temperature of beta-LG. Further chemical modification of Cys
(carboxymethylation) or positively charged residues (acetylation) of beta-LG totally abolished its immunoreactivity, confirming the
conformation-dependent nature of this mAb. Using competitive ELISA, the 4H11E8 mAb could determine the native beta-LG content in
commercially processed milks. Concentrations of native beta-LG varied significantly among the local brands tested. From a technological
standpoint, the mAb prepared in this study is relevant to the design and operation of appropriate processes for thermal sanitation of milk and of other dairy products.
