Park Miso, Sun Qing, Liu Fang, DeLisa Matthew P, Chen Wilfred
Department of Chemical and Biomolecular Engineering, University of Delaware, Newark, Delaware, United States of America.
School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York, United States of America.
PLoS One. 2014 May 12;9(5):e97103. doi: 10.1371/journal.pone.0097103. eCollection 2014.
The systematic organization of enzymes is a key feature for the efficient operation of cascade reactions in nature. Here, we demonstrate a facile method to create nanoscale enzyme cascades by using engineered bacterial outer membrane vesicles (OMVs) that are spheroid nanoparticles (roughly 50 nm in diameter) produced by Gram-negative bacteria during all phases of growth. By taking advantage of the fact that OMVs naturally contain proteins found in the outer cell membrane, we displayed a trivalent protein scaffold containing three divergent cohesin domains for the position-specific presentation of a three-enzyme cascade on OMVs through a truncated ice nucleation protein anchoring motif (INP). The positional assembly of three enzymes for cellulose hydrolysis was demonstrated. The enzyme-decorated OMVs provided synergistic cellulose hydrolysis resulting in 23-fold enhancement in glucose production than free enzymes.
酶的系统组织是自然界中级联反应高效运行的关键特征。在此，我们展示了一种简便方法，通过使用工程化细菌外膜囊泡（OMV）来创建纳米级酶级联反应，OMV是革兰氏阴性细菌在生长各阶段产生的球形纳米颗粒（直径约50纳米）。利用OMV天然含有外细胞膜中发现的蛋白质这一事实，我们展示了一种三价蛋白质支架，其包含三个不同的黏连蛋白结构域，通过截短的冰核蛋白锚定基序（INP）在OMV上进行位置特异性展示三酶级联反应。证明了三种用于纤维素水解的酶的位置组装。酶修饰的OMV提供协同纤维素水解，导致葡萄糖产量比游离酶提高23倍。
