The contribution of non-native structure with recombinant cobrotoxin to its immunoreactivity toward anti-cobrotoxin antibodies
碩士 === 國立中山大學 === 生物醫學研究所 === 97 === To induce the production of antibodies, exogenous antigens are taken up and degraded in antigen presenting cells in vivo. Since this process inevitably lead to distort antigen’s structure, it is likely that some arising antibodies following immunization may not r...
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ndltd-TW-097NSYS51140052019-05-15T19:27:45Z http://ndltd.ncl.edu.tw/handle/pkc977 The contribution of non-native structure with recombinant cobrotoxin to its immunoreactivity toward anti-cobrotoxin antibodies 臺灣眼鏡蛇神經毒重組蛋白非原生態結構與抗體反應性之研究 Sheng-che Ding 丁聖哲 碩士 國立中山大學 生物醫學研究所 97 To induce the production of antibodies, exogenous antigens are taken up and degraded in antigen presenting cells in vivo. Since this process inevitably lead to distort antigen’s structure, it is likely that some arising antibodies following immunization may not react appropriately with native protein. In the present study, comparative studies on the reactivity of cobrotoxin and recombinant cobrotoxin toward anti-cobrotoxin antibodies were carried out. CD spectra and acrylamide quenching of Trp fluorescence showed that global structure of recombinant cobrotoxin was different from that of native toxin. Results of ELISA and dot blotting assay revealed that recombinant cobrotoxin had a superior reactivity toward anti-cobrotoxin antibodies than native toxin did. Reactivity with antibody fractions specifically against N-terminal region or C-terminal region of cobrotoxin also showed the same results. The binding of recombinant cobrotoxin with antibodies was stronger than that of cobrotoxin as revealed by ammonium thiocyanate elution assay. Recombinant protein was susceptible to reduce its antigenicity after tryptic digestion compared to cobrotoxin. Distorting disulfide linkages at C-terminus caused a marked decrease in immunoreactivity of recombinant cobrotoxin, indicating that anti-cobrotoxin antibodies mostly recognized conformation-dependent epitopes. Moreover, cobrotoxin and recombinant cobrotoxin showed a similar immunoreactivity under denaturing condition. Taken together, these results suggest that native conformation with cobrotoxin may unfavorably impede the interaction of some epitope(s) with anti-cobrotoxin antibodies. Long-Sen Chang 張榮賢 2009 學位論文 ; thesis 64 zh-TW |
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碩士 === 國立中山大學 === 生物醫學研究所 === 97 === To induce the production of antibodies, exogenous antigens are taken up and degraded in antigen presenting cells in vivo. Since this process inevitably lead to distort antigen’s structure, it is likely that some arising antibodies following immunization may not react appropriately with native protein. In the present study, comparative studies on the reactivity of cobrotoxin and recombinant cobrotoxin toward anti-cobrotoxin antibodies were carried out. CD spectra and acrylamide quenching of Trp fluorescence showed that global structure of recombinant cobrotoxin was different from that of native toxin. Results of ELISA and dot blotting assay revealed that recombinant cobrotoxin had a superior reactivity toward anti-cobrotoxin antibodies than native toxin did. Reactivity with antibody fractions specifically against N-terminal region or C-terminal region of cobrotoxin also showed the same results. The binding of recombinant cobrotoxin with antibodies was stronger than that of cobrotoxin as revealed by ammonium thiocyanate elution assay. Recombinant protein was susceptible to reduce its antigenicity after tryptic digestion compared to cobrotoxin. Distorting disulfide linkages at C-terminus caused a marked decrease in immunoreactivity of recombinant cobrotoxin, indicating that anti-cobrotoxin antibodies mostly recognized conformation-dependent epitopes. Moreover, cobrotoxin and recombinant cobrotoxin showed a similar immunoreactivity under denaturing condition. Taken together, these results suggest that native conformation with cobrotoxin may unfavorably impede the interaction of some epitope(s) with anti-cobrotoxin antibodies.
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author2 |
Long-Sen Chang |
author_facet |
Long-Sen Chang Sheng-che Ding 丁聖哲 |
author |
Sheng-che Ding 丁聖哲 |
spellingShingle |
Sheng-che Ding 丁聖哲 The contribution of non-native structure with recombinant cobrotoxin to its immunoreactivity toward anti-cobrotoxin antibodies |
author_sort |
Sheng-che Ding |
title |
The contribution of non-native structure with recombinant cobrotoxin to its immunoreactivity toward anti-cobrotoxin antibodies |
title_short |
The contribution of non-native structure with recombinant cobrotoxin to its immunoreactivity toward anti-cobrotoxin antibodies |
title_full |
The contribution of non-native structure with recombinant cobrotoxin to its immunoreactivity toward anti-cobrotoxin antibodies |
title_fullStr |
The contribution of non-native structure with recombinant cobrotoxin to its immunoreactivity toward anti-cobrotoxin antibodies |
title_full_unstemmed |
The contribution of non-native structure with recombinant cobrotoxin to its immunoreactivity toward anti-cobrotoxin antibodies |
title_sort |
contribution of non-native structure with recombinant cobrotoxin to its immunoreactivity toward anti-cobrotoxin antibodies |
publishDate |
2009 |
url |
http://ndltd.ncl.edu.tw/handle/pkc977 |
work_keys_str_mv |
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