Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/3906
標題: 聚乙二醇化對酵素活性影響之探討
Effects of PEGylation on enzyme activity
作者: 謝元榜
Hsieh, Yuan-Pang
關鍵字: PEGylation;聚乙二醇;intertwinement;amphiphilic polymers;纏繞;雙性分子
出版社: 化學工程學系所
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摘要: 
在此研究中,我們利用4-nitrophenyl chloroformate活化mPEG,使其接附至酵素上,經由實驗數據之分析來觀測其對酵素活性及穩定性之影響。
然本實驗選用之酵素:L-N-carbamoylase(L-Ncar)及Horseradish Peroxidase(HRP),二者經聚乙二醇化(pegylation)後,在活性上卻有相反之效果。
L-Ncar經聚乙二醇化後,在活性上明顯有下降之趨勢,其原因我們推估可能是接附至酵素上之mPEG長鏈對酵素發生纏繞現象,致使酵素構形改變或是遮蔽其活化位置。
HRP經聚乙二醇化後,在活性上卻有明顯之提升,其原因我們推測可能與mPEG發生水合作用有關,因mPEG屬雙性分子,在溶液中可能因水合作用將水分子往自身高分子方向吸引,將基質往外推送,因而在微觀下形成一類似兩相之狀態,致使產生活性提升之結果;另在實驗中亦發現HRP經聚乙二醇化後,其熱穩定性有明顯之提升及其得以在低酵素濃度下長時間仍保有高活性,我們推測應與接附高分子之酵素,其剛性增加,致使構形不易改變有關。

In this thesis, the activity and stability of PEGylated enzyme were studied. The activated mPEG (mPEG*) prepared by 4-nitrophenyl chloroformate was attached to the enzyme, and the activity of PEGylated enzymes, L-N-carbamoylase (L-Ncar) and Horseradish Peroxidase (HRP), were compared. The opposite effect of activity between two enzymes indicated the different conjugated chemistry.
The PEGylated L-Ncar shows lower activity compared to native one, and the suspicious root cause could be the long chain of mPEG causing the intertwinement with the enzyme. The long chain would cause the transformation of enzyme conformation or shield the active sites of enzyme.
On the other hand, PEGylated HRP shows higher activity with activated mPEG. This opposite result could be attributed to the hydration of mPEG. Because mPEG is an amphiphilic polymer, the effect of hydration would cause water molecules attracting to itself in the solution and, at the same time, exclude substrates from itself. It might become aqueous two-phase system in microenvironment, and it could lead to higher activity. In addition, we also found the PEGylated HRP could enhance the thermal stability and maintain high activity even in low concentration for a long term. This result implied that PEGylated HRP have more rigid conformation to resist tranformation from experimental environment.
URI: http://hdl.handle.net/11455/3906
其他識別: U0005-1708201114210000
Appears in Collections:化學工程學系所

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