蛋白質元器件的合成與組裝

冷艷　門冬

摘 要：利用合成生物學建立人工合成生物體系與生物制造是一個系統工程，人造功能器件在其中發揮著重要作用。當前的合成生物學能夠組織不同來源的順序催化酶分子在細胞內制造全新的合成途徑，但仍缺少在空間上有機組織酶分子的手段。如果能夠人為地構建空間有序的多酶復合體將使人工合成反應體系更快速、有效。蛋白納米纖維和病毒顆粒及其包裝蛋白區別于其它納米材料，具有天然的蛋白質特性、穩定規整的結構和自組裝特性，可用構建人工蛋白“腳手架”來“鉚定”多酶反應體的功能元件，以保護多酶耦合體系并實現其順序裝配。該項目一方面通過基因克隆構建了帶有功能元件的自組裝元件，發展了一種自組裝功能納米纖維的制備方法，并利用不同熒光信號分子標記代表不同蛋白組分，實現多種不同標記的Sup35有序組裝。同時，發展了一種基于磁珠技術控制的催化納米線的制備方法，該方法不同于傳統的吸附、包埋和共價交聯等方法，其偶聯過程依賴于自組裝，不會產生形變、變性或取向性明顯改變，因此對酶分子本身的活性少有直接的影響。對酶分子的固定化和多酶催化體系的模擬提供了新的技術路線和思路。另一方面以SV40病毒衣殼蛋白和HBV衣殼蛋白為研究對象，獲得了兩種體外組裝的類病毒樣顆粒，這兩種類病毒樣顆粒都具有籠型蛋白結構。這種籠型蛋白結構是空心的核殼性結構，一般具有內腔，這些特點為制造多層級、復雜、順序的多酶體系提供了優良的性質。該研究利用這個特性，在不影響類病毒樣顆粒組裝結構的前題下，在結構表面插入了一些功能基團，實現了這種三維蛋白骨架的多功能化。

關鍵詞：人造功能器件 蛋白納米纖維 類病毒樣顆粒 組裝 多功能化

Abstract：Using of synthetic biology to establish artificial synthetic biological systems and bio manufacturing is a systematic engineering，in which artificial function device plays a very important role. With Present synthetic biology technique scientists can to make use of sequential catalytic enzymes from different sources to build new synthetic pathways in cells， but scientists still lack the means to narrow the distance between the enzyme molecules to each other in space. Synthetic reaction system will become more rapid and more effective if we can artificially build space orderly multienzyme system. Different from other nano materials， protein nanofiber and virus-like particle， which have characteristics of natural protein， regular and stable structure and self-assembly properties， can be used to construct artificial protein “scaffolding” for “riveting” multienzyme system. This artificial protein "scaffolding" protect enzyme coupling system and its sequence assembly. In this project， on one hand， we construct the self-assembling component with functional components by gene cloning， develop the method for preparation of self-assembling functional nanofiber， and make use of different fluorescence signal molecular markers to represent different protein components for achieving ordered assembly of different labeled sup35 protein. At same time， we develop the method for preparation of the catlytic nanowires based on control of magnetic bead technique.Different from the traditional method of adsorption， entrapment and covalent cross-linking， the coupling process in this method relies on self-assembly， and deformation， degeneration or orientation change will not generate， so that only little direct influence to the activity of enzyme molecule will appear. This method provides a new technology roadmap and ideas for immobilization of enzyme molecules and simulation of multienzyme system. On the other hand， through the study of SV40 capsid protein and HBV capsid protein， we obtain two self-assembly of virus-like particles in vitro. Virus-like particles have cage-type protein structure， which is hollow core-shell structure and has cavity generally. These characteristics of virus-like particles provide excellent properties for manufacturing multi-level， complex， sequential multienzyme system and achieving multi-functional three-dimensional protein.

Key Words：Artificial function device；Protein nanofiber；Virus-like particles；Assembly；Multi-functional

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