蛋白质元器件的合成与组装

冷艳　门冬

摘 要：利用合成生物学建立人工合成生物体系与生物制造是一个系统工程，人造功能器件在其中发挥着重要作用。当前的合成生物学能够组织不同来源的顺序催化酶分子在细胞内制造全新的合成途径，但仍缺少在空间上有机组织酶分子的手段。如果能够人为地构建空间有序的多酶复合体将使人工合成反应体系更快速、有效。蛋白纳米纤维和病毒颗粒及其包装蛋白区别于其它纳米材料，具有天然的蛋白质特性、稳定规整的结构和自组装特性，可用构建人工蛋白“脚手架”来“铆定”多酶反应体的功能元件，以保护多酶耦合体系并实现其顺序装配。该项目一方面通过基因克隆构建了带有功能元件的自组装元件，发展了一种自组装功能纳米纤维的制备方法，并利用不同荧光信号分子标记代表不同蛋白组分，实现多种不同标记的Sup35有序组装。同时，发展了一种基于磁珠技术控制的催化纳米线的制备方法，该方法不同于传统的吸附、包埋和共价交联等方法，其偶联过程依赖于自组装，不会产生形变、变性或取向性明显改变，因此对酶分子本身的活性少有直接的影响。对酶分子的固定化和多酶催化体系的模拟提供了新的技术路线和思路。另一方面以SV40病毒衣壳蛋白和HBV衣壳蛋白为研究对象，获得了两种体外组装的类病毒样颗粒，这两种类病毒样颗粒都具有笼型蛋白结构。这种笼型蛋白结构是空心的核壳性结构，一般具有内腔，这些特点为制造多层级、复杂、顺序的多酶体系提供了优良的性质。该研究利用这个特性，在不影响类病毒样颗粒组装结构的前题下，在结构表面插入了一些功能基团，实现了这种三维蛋白骨架的多功能化。

关键词：人造功能器件 蛋白纳米纤维 类病毒样颗粒 组装 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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