Engineered catalytic biofilms: Site-specific enzyme immobilization onto E-coli curli nanofibers
Engineered catalytic biofilms: Site-specific enzyme immobilization onto E-coli curli nanofibers
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DOI：
10.1002/bit.25638
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发表时间：
2015-10-01
影响因子：
3.8
通讯作者：
Joshi, Neel S.
中科院分区：
文献类型：
--
作者：
Botyanszki, Zsofia;Tay, Pei Kun R.;Joshi, Neel S.
生物催化转化通常依赖于纯化的酶或全细胞来进行复杂的转化，其在工业规模上用于化学、药物和生物燃料合成、农药去污和水净化。然而，这两种系统都具有与酶纯化相关的成本、固定化酶的长期稳定性、催化剂回收和与苛刻反应条件的相容性相关的固有缺点。我们开发了一种基于生物膜集成纳米纤维显示（BIND）的合理设计的生物催化表面的新策略，该策略利用了大肠杆菌的卷曲系统。大肠杆菌，以创建能够共价固定酶的功能性生物素网络。这种方法是有吸引力的，因为它是可扩展的，代表了一个模块化的策略，位点特异性酶固定化，并有可能在变性环境条件下稳定酶。我们将与SpyCatcher连接结构域融合的重组β-淀粉酶定点固定到E.显示互补SpyTag捕获结构域的大肠杆菌卷曲纤维。我们表征了这种固定化技术在生物膜上的有效性，并测试了固定化淀粉酶在不利条件下的稳定性。这种酶修饰的生物膜在暴露于广泛的pH和有机溶剂条件下时保持其活性。与依赖于高细胞代谢的其他基于生物膜的催化剂相比，改性的基于卷曲的生物膜即使在由于有机溶剂暴露而导致的细胞死亡后仍保持活性。这项工作为利用大肠杆菌胞外聚合物基质的新方法奠定了基础。大肠杆菌用于创造新的生物催化表面。Biotechnol. Bioeng. 2015;112：2016-2024。(c)2015 Wiley Periodicals，Inc.
Biocatalytic transformations generally rely on purified enzymes or whole cells to perform complex transformations that are used on industrial scale for chemical, drug, and biofuel synthesis, pesticide decontamination, and water purification. However, both of these systems have inherent disadvantages related to the costs associated with enzyme purification, the long-term stability of immobilized enzymes, catalyst recovery, and compatibility with harsh reaction conditions. We developed a novel strategy for producing rationally designed biocatalytic surfaces based on Biofilm Integrated Nanofiber Display (BIND), which exploits the curli system of E. coli to create a functional nanofiber network capable of covalent immobilization of enzymes. This approach is attractive because it is scalable, represents a modular strategy for site-specific enzyme immobilization, and has the potential to stabilize enzymes under denaturing environmental conditions. We site-specifically immobilized a recombinant -amylase, fused to the SpyCatcher attachment domain, onto E. coli curli fibers displaying complementary SpyTag capture domains. We characterized the effectiveness of this immobilization technique on the biofilms and tested the stability of immobilized -amylase in unfavorable conditions. This enzyme-modified biofilm maintained its activity when exposed to a wide range of pH and organic solvent conditions. In contrast to other biofilm-based catalysts, which rely on high cellular metabolism, the modified curli-based biofilm remained active even after cell death due to organic solvent exposure. This work lays the foundation for a new and versatile method of using the extracellular polymeric matrix of E. coli for creating novel biocatalytic surfaces. Biotechnol. Bioeng. 2015;112: 2016-2024. (c) 2015 Wiley Periodicals, Inc.