Engineered protein nano-compartments for targeted enzyme localization.
Engineered protein nano-compartments for targeted enzyme localization.
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用于靶向酶定位的工程蛋白质纳米室。
DOI：
10.1371/journal.pone.0033342
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Schmidt-Dannert C
中科院分区：
文献类型：
--
作者：
Choudhary S;Quin MB;Sanders MA;Johnson ET;Schmidt-Dannert C
酶及其底物的区室化共定位代表了工程细胞中多酶合成和生物催化的有吸引力的方法。酶和底物的螯合将大大提高反应效率，同时还保护工程化的宿主细胞免受潜在毒性反应中间体的影响。几种细菌形成基于蛋白质的多面体微区室，其隔离功能相关的酶并调节它们对底物和其他小代谢物的获取。这样的细菌微区室可以被工程化到基于蛋白质的纳米生物反应器中，条件是它们可以在非天然宿主细胞中组装，并且异源酶和底物可以靶向到工程化的区室中。在这里，我们报告了重组表达沙门氏菌乙醇胺利用（eut）细菌微室壳蛋白在E。大肠杆菌导致多面体蛋白质壳的形成。纯化的重组壳在形态上类似于从S.肠令人惊讶的是，重组表达的外壳蛋白（EutS）中的仅一个是足够的和必要的创建适当界定的区室。与EutS的共表达也促进了与推定的Eut壳靶向信号序列融合的EGFP的包封。我们还证明了针对重组壳的异源酶（β-半乳糖苷酶）的功能定位。我们的研究结果为生物合成和生物催化的蛋白质纳米区室的工程设计提供了概念验证。
Compartmentalized co-localization of enzymes and their substrates represents an attractive approach for multi-enzymatic synthesis in engineered cells and biocatalysis. Sequestration of enzymes and substrates would greatly increase reaction efficiency while also protecting engineered host cells from potentially toxic reaction intermediates. Several bacteria form protein-based polyhedral microcompartments which sequester functionally related enzymes and regulate their access to substrates and other small metabolites. Such bacterial microcompartments may be engineered into protein-based nano-bioreactors, provided that they can be assembled in a non-native host cell, and that heterologous enzymes and substrates can be targeted into the engineered compartments. Here, we report that recombinant expression of Salmonella enterica ethanolamine utilization (eut) bacterial microcompartment shell proteins in E. coli results in the formation of polyhedral protein shells. Purified recombinant shells are morphologically similar to the native Eut microcompartments purified from S. enterica. Surprisingly, recombinant expression of only one of the shell proteins (EutS) is sufficient and necessary for creating properly delimited compartments. Co-expression with EutS also facilitates the encapsulation of EGFP fused with a putative Eut shell-targeting signal sequence. We also demonstrate the functional localization of a heterologous enzyme (β-galactosidase) targeted to the recombinant shells. Together our results provide proof-of-concept for the engineering of protein nano-compartments for biosynthesis and biocatalysis.
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影响因子：
5.1
作者：
Bertin, Yolande;Girardeau, J. P.;Martin, Christine
通讯作者：
Martin, Christine
影响因子：
3.2
作者：
Kofoid, E;Rappleye, C;Roth, J
通讯作者：
Roth, J
DOI：
10.1073/pnas.0913199107
发表时间：
2010-04-20
影响因子：
11.1
作者：
Fan, Chenguang;Cheng, Shouqiang;Bobik, Thomas A.
通讯作者：
Bobik, Thomas A.
影响因子：
3.2
作者：
Bobik, TA;Havemann, GD;Aldrich, HC
通讯作者：
Aldrich, HC
DOI：
10.1073/pnas.96.22.12929
发表时间：
1999-10-26
影响因子：
11.1
作者：
Burbulis, IE;Winkel-Shirley, B
通讯作者：
Winkel-Shirley, B