Spore-displayed enzyme cascade with tunable stoichiometry
Spore-displayed enzyme cascade with tunable stoichiometry
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DOI：
10.1002/btpr.2416
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发表时间：
2017-03-01
影响因子：
2.9
通讯作者：
Ge, Xin
中科院分区：
文献类型：
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作者：
Chen, Long;Mulchandani, Ashok;Ge, Xin
利用内生孢子惰性和稳定的特性，我们开发了一种在枯草芽孢杆菌孢子表面固定化多种酶的生物催化平台。在枯草芽孢杆菌外衣蛋白中，CotG 介导热纤梭菌粘连蛋白 (CtCoh) 的高表达水平，其功能展示能力约为每个孢子 10(4) 个木糖还原酶-C 分子。热纤细胞泊素融合蛋白 (XR-CtDoc)。通过黄色瘤胃球菌粘连蛋白-dockerin模块将亚磷酸脱氢酶（PTDH）共固定在孢子表面，实现了NADPH的再生。木糖还原酶 (XR) 和 PTDH 在孢子表面展示时都表现出增强的稳定性。更重要的是，通过改变与CotG融合的CtCoh和RfCoh的拷贝数，以可控的方式调节固定化酶之间的摩尔比。孢子展示 XR/PTDH 化学计量的优化导致木糖醇产量增加。总之，内生孢子表面展示提供了一种酶级联固定的新方法，具有改进的稳定性和可调的化学计量。 (c) 2016 年美国化学工程师生物技术研究所。编, 33:383-389, 2017
Taking the advantages of inert and stable nature of endospores, we developed a biocatalysis platform for multiple enzyme immobilization on Bacillus subtilis spore surface. Among B. subtilis outer coat proteins, CotG mediated a high expression level of Clostridium thermocellum cohesin (CtCoh) with a functional display capability of approximate to 10(4) molecules per spore of xylose reductase-C. thermocellum dockerin fusion protein (XR-CtDoc). By co-immobilization of phosphite dehydrogenase (PTDH) on spore surface via Ruminococcus flavefaciens cohesin-dockerin modules, regeneration of NADPH was achieved. Both xylose reductase (XR) and PTDH exhibited enhanced stability upon spore surface display. More importantly, by altering the copy numbers of CtCoh and RfCoh fused with CotG, the molar ratio between immobilized enzymes was adjusted in a controllable manner. Optimization of spore-displayed XR/PTDH stoichiometry resulted in increased yields of xylitol. In conclusion, endospore surface display presents a novel approach for enzyme cascade immobilization with improved stability and tunable stoichiometry. (c) 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:383-389, 2017