標題: 功能性奈米金粒子對增強酵素催化活性之動力學探討
An Enzymatic kinetics Investigation into the Significantly Enhanced Activity of Functionalized Gold Nanoparticles
作者: 吳中書
Chung-Shu Wu
柯富祥
Fu-Hsiang Ko
材料科學與工程學系奈米科技碩博士班
關鍵字: 金;奈米粒子;酵素動力學;催化活性;轉脂酶;gold;nanoparticles;enzyme kinetic;catalytic activity;lipase
公開日期: 2007
摘要: 在本篇研究中,我們利用奈米金粒子本身對生物分子的選擇性將酵素固定於粒子的表面並探討其酵素行為的差異,在之前有關這方面的研究,都會有修飾奈米粒子表面的步驟,然而這些動作不但費時且複雜,對於酵素的影響也難以估計,更重要的是,這樣的方法無法確定奈米粒子的表面覆蓋率而造成錯估其動力學參數,因此我們在保持金奈米粒子的穩定性下,發展不需表面修飾的量測系統以減少對酵素活性的影響並找出生物分子與奈米粒子之間的最佳比例。 我們發現酵素固定於奈米金上其活性的確有顯著性的增強,進ㄧ步的實驗結果顯示其增強的行為是由於反應物與酵素之間親合力發生變化所導致,這樣的結果支持奈米金粒子在這反應中為一項影響酵素催化的因子,像這樣基礎性理論的探討對於日後在奈米生物領域上的發展有所貢獻。
Enzymes immobilize onto gold nanoparticles (AuNPs) in the absence of a linker, using rapid and uncomplicated processes, generally possess higher activity bound to the surfaces through chemical modification. In previous reports, catalytic activity of enzyme-functionalized AuNPs has been investigated with the surface modification of linkers. Although the surface modification of linkers protects AuNPs from aggregation upon reactant’s entering, it will be perplexed at examination the coverage of the enzyme onto nanoparticles because less or much enzyme can not be identified by salt titration. Herein, linker-free immobilization is beneficial for developing precise activity assays. AuNP-bound lipase exhibits significantly enhanced catalytic activity relative to that of the free enzyme. In this work, we have systematically investigated the interactions between the NP monolayer and the affected substrates by quantifying the kinetic parameters kcat and KM to understand the enzymatic behavior of AuNP-bound lipase. Investigation of the kinetic parameters reveals that these two systems operate with the same value of kcat; i.e., the AuNPs exert no influence on the process of product release in the rate-limiting step. The Michaelis–Menten curves revealed that the AuNP-bound lipase provided the lower value of KM. Thus, the addition of AuNPs is an efficacious means of tuning enzyme–substrate association. We suspect that such fundamental research will aid in the development of new nanobiotechnological applications.
URI: http://140.113.39.130/cdrfb3/record/nctu/#GT009552510
http://hdl.handle.net/11536/39448
Appears in Collections:Thesis


Files in This Item:

  1. 251001.pdf