本文關(guān)鍵詞： 銀納米顆粒 牛血清蛋白 離散型共聚物 熒光淬滅 構(gòu)象變化　出處：《內(nèi)蒙古農(nóng)業(yè)大學(xué)》2016年博士論文　論文類型：學(xué)位論文

【摘要】：有機(jī)分子特別是生物大分子與無(wú)機(jī)納米顆粒形成的納米結(jié)構(gòu)具有優(yōu)良特性,在生物醫(yī)藥研究及應(yīng)用領(lǐng)域引起的關(guān)注日益增多,尤其是在血液中的檢測(cè)、呈遞、診療等方面得到了迅速的發(fā)展。血漿中含量最豐富的成分之一是血清白蛋白,無(wú)論是內(nèi)源性物質(zhì)還是外源性物質(zhì),包括納米顆粒都容易與其發(fā)生相互作用。納米顆粒作為藥物載體進(jìn)入血液后會(huì)不可避免地附著血清白蛋白,其功能不可避免會(huì)受到影響。因此,納米顆粒與蛋白質(zhì)相互作用研究是設(shè)計(jì)和構(gòu)建基于納米顆粒的生物傳感器的先決條件,對(duì)于探索納米材料在生命科學(xué)領(lǐng)域的應(yīng)用具有重要意義。為了實(shí)現(xiàn)納米顆粒在血液環(huán)境下的應(yīng)用,本文對(duì)銀納米顆粒一一牛血清白蛋白暈圈這種復(fù)合物的特性及形成機(jī)制進(jìn)行了系統(tǒng)的研究。利用雙親性聚合物(Amphiphilic polymer, AP)進(jìn)行表面改性可以使疏水銀納米顆粒具有水溶性,在此過(guò)程中顆粒粒徑變化較大,并引起此外最大吸收峰的輕微藍(lán)移,然而外部形貌特征仍然比較規(guī)則且分散均勻。在水相環(huán)境中與血液相關(guān)的不同蛋白質(zhì)混合,常溫下反應(yīng),牛血清白蛋白(Bovine serum albuinin, BSA)表現(xiàn)出與雙親性聚合物包裹的銀納米顆粒具有突出的吸附特性,通過(guò)調(diào)節(jié)混合比例,能夠得到吸附BSA數(shù)量可控的復(fù)合物。結(jié)合這種通用的表面修飾策略和物理吸附形成的納米顆粒一蛋白復(fù)合物,雙親性聚合物的外殼不僅可以為納米顆粒表面提供親水基團(tuán),同時(shí)也形成了一個(gè)較厚的隔離層,蛋白分子并不直接接觸金屬納米顆粒,弱化了金屬本身與蛋白之間的相互作用,對(duì)于維持納米顆粒一蛋白復(fù)合物穩(wěn)定性至關(guān)重要。與化學(xué)耦合方法相比,物理吸附介導(dǎo)蛋白——納米顆粒復(fù)合物擁有容易合成,結(jié)合穩(wěn)定的以及可逆的偶聯(lián)一解離等諸多的優(yōu)勢(shì),在納米生物醫(yī)藥領(lǐng)域具有更加廣泛應(yīng)用前景。納米顆粒與離散蛋白質(zhì)僅可以通過(guò)一個(gè)簡(jiǎn)單的混合過(guò)程,隨后進(jìn)行瓊脂糖凝膠電泳分離,由此產(chǎn)生基于物理吸附形成穩(wěn)定的復(fù)合物足以抵抗常見(jiàn)的凝膠電泳等分離技術(shù),而不會(huì)造成納米顆粒表面BSA的脫離。這種強(qiáng)烈相互作用,源于雙親性聚合物外殼賦予銀顆粒的水溶性與顆粒表面同BSA分子形成物理吸附,兩個(gè)因素之間相互平衡、共同作用。結(jié)合Stern-Volmer和Hill等方程并采用熒光猝滅的方法,系統(tǒng)地研究了兩親性聚合物包覆的納米銀顆粒與牛血清白蛋白之間的相互作用。這種結(jié)合力為1.30x107M-1并且其相互作用從熱力學(xué)的觀點(diǎn)來(lái)說(shuō)是自發(fā)的通過(guò)物理吸附實(shí)現(xiàn)的。雙親性聚合物修飾的銀納米顆粒表面會(huì)形成蛋白暈圈,當(dāng)精確控制摩爾比混合時(shí)這樣的結(jié)構(gòu)可以形成離散的納米顆粒一蛋白共聚物。同時(shí)使用熒光光譜的方法研究結(jié)合納米顆粒引起的蛋白構(gòu)象的變化,結(jié)果顯示納米顆粒引起B(yǎng)SA的強(qiáng)烈物理吸附,蛋白的空間構(gòu)象存在輕微的改變。
[Abstract]:Organic molecules, especially biomolecules and inorganic nanoparticles, have excellent properties, and have attracted increasing attention in biomedical research and applications, especially in the detection and presentation of blood. One of the most abundant components in plasma is serum albumin, whether endogenous or exogenous. Nanoparticles are easy to interact with them. As drug carriers, nanoparticles will inevitably adhere to serum albumin, and its function will inevitably be affected. The study of the interaction between nanoparticles and proteins is a prerequisite for the design and construction of biosensors based on nanoparticles. In order to realize the application of nanoparticles in blood environment, it is of great significance to explore the application of nanomaterials in the field of life science. In this paper, the characteristics and formation mechanism of silver nanoparticles, bovine serum albumin halo ring, were systematically studied. The amphiphilic polymer Amphiphilic polymer was used. The surface modification of APs can make the hydrophobic silver nanoparticles water-soluble, and the particle size changes greatly during the process, which also leads to the slight blue shift of the maximum absorption peak. However, the external morphologies are still regular and evenly dispersed. In aqueous environment, different proteins associated with blood are mixed and reacted at room temperature. Bovine serum albumin (BSA) exhibited outstanding adsorption properties with silver nanoparticles coated with amphiphilic polymers. By adjusting the mixing ratio, the adsorbed BSA complex with controllable number can be obtained, which combines the general surface modification strategy with the physical adsorption to form a nano-particle protein complex. The shell of amphiphilic polymer can not only provide hydrophilic groups on the surface of nanoparticles, but also form a thick isolation layer. Protein molecules do not directly contact metal nanoparticles. It weakens the interaction between metal and protein, which is very important to maintain the stability of nanoparticles and protein complexes, compared with the chemical coupling method. Physical adsorption mediated protein-nanoparticle complexes have many advantages, such as easy synthesis, stable and reversible coupling dissociation and so on. Nanoparticles and discrete proteins can only be separated by a simple mixing process and then separated by agarose gel electrophoresis. The formation of stable complexes based on physical adsorption is sufficient to resist common separation techniques such as gel electrophoresis without causing the separation of BSA on the surface of nanoparticles. The water solubility of silver particles is attributed to the amphiphilic polymer shell, and the surface of the particles is physically adsorbed with BSA molecules, and the two factors balance each other. Combined with Stern-Volmer and Hill equations and fluorescence quenching method. The interaction between amphiphilic polymer-coated silver nanoparticles and bovine serum albumin (BSA) has been systematically studied. The binding force is 1.30x107M-1 and the interaction is from a thermodynamic point of view. Amphiphilic polymer modified silver nanoparticles will form protein halos on the surface. When the molar ratio is accurately controlled, this structure can form a discrete nano-particle protein copolymer. Meanwhile, the conformational changes of proteins induced by the combination of nanoparticles are studied by fluorescence spectroscopy. The results showed that BSA was strongly physically adsorbed by nanoparticles and the spatial conformation of protein changed slightly.
【學(xué)位授予單位】：內(nèi)蒙古農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類號(hào)】：TB383.1;Q51

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本文編號(hào)：1492664