State Key Laboratory of Chemical Resources Engineering, Beijing University of Chemical Technology, 100029, Beijing, People's Republic of China.
Microb Cell Fact. 2018 Feb 20;17(1):26. doi: 10.1186/s12934-018-0873-3.
When using the microbial cell factories for green manufacturing, several important issues need to be addressed such as how to maintain the stability of biocatalysts used in the bioprocess and how to improve the synthetic efficiency of the biological system. One strategy widely used during natural evolution is the creation of organelles which can be used for regional control. This kind of compartmentalization strategy has inspired the design of artificial organelle-like nanodevice for synthetic biology and "green chemistry".
Mimicking the natural concept of functional compartments, here we show that the engineered thermostable ketohydroxyglutarate aldolase from Thermotoga maritima could be developed as a general platform for nanoreactor design via supramolecular self-assembly. An industrial biocatalyst-(+)-γ-lactamase was selected as a model catalyst and successful encapsulated in the nanoreactor with high copies. These nanomaterials could easily be synthesized by Escherichia coli by heterologous expression and subsequently self-assembles into the target organelle-like nanoreactors both in vivo and in vitro. By probing their structural characteristics via transmission electronic microscopy and their catalytic activity under diverse conditions, we proved that these nanoreactors could confer a significant benefit to the cargo proteins. The encapsulated protein exhibits significantly improved stability under conditions such as in the presence of organic solvent or proteases, and shows better substrate tolerance than free enzyme.
Our biodesign strategy provides new methods to develop new catalytically active protein-nanoreactors and could easily be applied into other biocatalysts. These artificial organelles could have widely application in sustainable catalysis, synthetic biology and could significantly improve the performance of microbial cell factories.
在利用微生物细胞工厂进行绿色制造时，需要解决几个重要问题，例如如何维持生物过程中使用的生物催化剂的稳定性，以及如何提高生物系统的合成效率。在自然进化过程中广泛使用的一种策略是创建细胞器，可用于区域控制。这种分隔化策略启发了用于合成生物学和“绿色化学”的人工细胞器样纳米器件的设计。
模仿功能隔室的自然概念，我们展示了来自 Thermotoga maritima 的工程化耐热酮羟戊二酸醛缩酶可以通过超分子自组装开发为纳米反应器设计的通用平台。选择工业生物催化剂(+)-γ-内酰胺酶作为模型催化剂，并成功地以高拷贝数封装在纳米反应器中。这些纳米材料可以通过大肠杆菌的异源表达容易地合成，并在体内和体外都能自发组装成目标细胞器样纳米反应器。通过透射电子显微镜探测它们的结构特征和在不同条件下的催化活性，我们证明这些纳米反应器可以为货物蛋白带来显著的益处。封装的蛋白质在存在有机溶剂或蛋白酶等条件下表现出显著提高的稳定性，并且比游离酶具有更好的底物耐受性。
我们的生物设计策略为开发新的催化活性蛋白-纳米反应器提供了新方法，并且可以轻松应用于其他生物催化剂。这些人工细胞器在可持续催化、合成生物学中具有广泛的应用前景，并可以显著提高微生物细胞工厂的性能。
