Engineering Functional Membrane-Membrane Interfaces by InterSpy.
Engineering Functional Membrane-Membrane Interfaces by InterSpy.
复制标题
DOI：
10.1002/smll.202202104
复制
发表时间：
2023-03
期刊：
影响因子：
13.3
通讯作者：
Ha, Taekjip
中科院分区：
文献类型：
--
作者：
Moghimianavval, Hossein;Patel, Chintan;Mohapatra, Sonisilpa;Hwang, Sung-Won;Kayikcioglu, Tunc;Bashirzadeh, Yashar;Liu, Allen P.;Ha, Taekjip
关键词：
膜之间的工程合成界面在设计非天然细胞通信途径和创建合成组织中具有潜在的应用。在这里，InterSpy作为一种合成生物学工具被引入，该工具由异二聚体蛋白组成，该异二聚体蛋白被工程化以通过SpyTag/SpyCatcher相互作用形成和维持并置合成膜以及细胞膜之间的膜-膜界面。在设计的InterSpy工具包中包含分裂的荧光蛋白片段，可以跟踪膜-膜界面的形成和功能性荧光蛋白在贴壁膜之间的空间中的重建。我们首先证明InterSpy通过使用哺乳动物无细胞表达系统测试分裂蛋白质设计。通过利用共翻译螺旋插入，将无细胞合成的InterSpy片段掺入脂质体的膜中，并支持具有所需拓扑结构的脂质双层。膜之间的分裂荧光蛋白的功能重建严格依赖于SpyTag/SpyCatcher。最后，由于InterSpy是完全遗传编码的，因此它对细胞的适应性已经建立。InterSpy展示了无细胞表达系统通过邻近诱导蛋白在合成膜界面功能重建中的能力。这项技术也可能被证明对合成生物学有用，在合成生物学中，使用最少的组件以受控的方式重建细胞间的接触和通信。
Engineering synthetic interfaces between membranes has potential applications in designing non-native cellular communication pathways and creating synthetic tissues. Here, InterSpy is introduced as a synthetic biology tool consisting of a heterodimeric protein engineered to form and maintain membrane-membrane interfaces between apposing synthetic as well as cell membranes through SpyTag/SpyCatcher interaction. Inclusion of split fluorescent protein fragments in the designed InterSpy toolkit allows tracking the formation of membrane-membrane interface and reconstitution of functional fluorescent protein in the space between apposing membranes. We first demonstrate InterSpy by testing split protein designs using a mammalian cell-free expression system. By utilizing co-translational helix insertion, cell-free synthesized InterSpy fragments are incorporated into the membrane of liposomes and supported lipid bilayers with a desired topology. Functional reconstitution of split fluorescent protein between the membranes is strictly dependent on SpyTag/SpyCatcher. Finally, since InterSpy is fully genetically encoded, its adaptation to cells is established. InterSpy demonstrates the power of cell-free expression systems in functional reconstitution of synthetic membrane interfaces via proximity-inducing proteins. This technology may also prove useful for synthetic biology where cell-cell contacts and communication are recreated in a controlled manner using minimal components.
登录
查看更多内容
DOI：
10.1083/jcb.200911024
发表时间：
2010-08-09
期刊：
The Journal of cell biology
影响因子：
--
作者：
Friedman JR;Webster BM;Mastronarde DN;Verhey KJ;Voeltz GK
通讯作者：
Voeltz GK
改进的荧光蛋白用于内源性蛋白质标记。
影响因子：
16.6
作者：
Feng S;Sekine S;Pessino V;Li H;Leonetti MD;Huang B
通讯作者：
Huang B
影响因子：
16.1
作者：
Chen, Xiaoying;Zaro, Jennica L.;Shen, Wei-Chiang
通讯作者：
Shen, Wei-Chiang
DOI：
10.1083/jcb.201009141
发表时间：
2011-03-21
期刊：
The Journal of cell biology
影响因子：
--
作者：
Baum B;Georgiou M
通讯作者：
Georgiou M
影响因子：
15
作者：
Kim CH;Axup JY;Dubrovska A;Kazane SA;Hutchins BA;Wold ED;Smider VV;Schultz PG
通讯作者：
Schultz PG