诺如病毒多功能蛋白NS3用于人工细胞物质交换模块的构建

孙雪倩; 尚坤; 李明; 杨晨光; 陆颖

doi:10.12211/2096-8280.2026-067

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诺如病毒多功能蛋白NS3用于人工细胞物质交换模块的构建

研究论文 | 更新时间：2026-10-15

- 诺如病毒多功能蛋白NS3用于人工细胞物质交换模块的构建
- Construction of artificial cell material exchange module using norovirus multifunctional protein NS3
- 合成生物学 2026年页码：1-17
- 作者机构：
  
  1.中国科学院物理研究所，北京 100190
  2.延安大学延安医学院，陕西延安 716000
  3.天津医科大学肿瘤医院，天津 300060
- 作者简介：
  
  孙雪倩（1999—），女，硕士研究生。研究方向为病原生物学。
  杨晨光（1994—），男，博士后，研究方向为单分子生物物理；膜成孔蛋白的动力学过程。
  陆颖（1981—），女，研究员，博士生导师。研究方向为生物膜系统的复杂动力学；人工细胞膜的形变及输运。
- 基金信息：
  
  中国科学院战略性先导科技专项(XDB0480000);国家重点研发计划(2019YFA0709304);国家自然科学基金(T2221001;32171228;32471278)
- DOI：10.12211/2096-8280.2026-067
  中图分类号： Q789;Q939.4
- 收稿：2026-07-29，
  
  修回：2026-10-12，
  
  网络首发：2026-10-15，
- 稿件说明：
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孙雪倩，尚坤，李明，杨晨光，陆颖. 诺如病毒多功能蛋白 NS3 用于人工细胞物质交换模块的构建［J］. 合成生物学， 2026， 7. DOI： 10.12211/2096-8280.2026-067

SUN Xueqian， SHANG Kun， LI Ming， YANG Chenguang， LU Ying. Construction of artificial cell material exchange module using norovirus multifunctional protein NS3［J］. Synthetic Biology Journal， 2026， 7.DOI： 10.12211/2096-8280.2026-067
孙雪倩，尚坤，李明，杨晨光，陆颖. 诺如病毒多功能蛋白 NS3 用于人工细胞物质交换模块的构建［J］. 合成生物学， 2026， 7. DOI： 10.12211/2096-8280.2026-067 DOI：

SUN Xueqian， SHANG Kun， LI Ming， YANG Chenguang， LU Ying. Construction of artificial cell material exchange module using norovirus multifunctional protein NS3［J］. Synthetic Biology Journal， 2026， 7.DOI： 10.12211/2096-8280.2026-067 DOI：

摘要

在自下而上构建人工细胞的研究中，理性构建具备多重生物活性的功能模块，是实现复杂类生命行为的重要基础。病毒多功能蛋白因其结构紧凑、功能多样，具有作为人工细胞功能元件的应用潜力。本研究以诺如病毒多功能蛋白NS3为例，利用无细胞蛋白质合成系统（CFPS）实现其体外合成，并证实所得NS3同时保留RNA解旋酶活性和磷脂膜透化活性。进一步研究表明，Mg

对NS3的两类功能具有差异性调控作用：Mg

促进NS3介导的RNA解旋过程，却显著抑制其诱导的脂质体透化活性。通过单分子荧光分析，我们发现Mg

可诱导NS3由开放构象向紧凑构象转变，并降低其在磷脂膜界面的插入深度，从而削弱膜透化能力。最后，我们利用NS3实现了对细胞尺寸巨型单层囊泡（GUV）的膜透化，并观察到Mg

能够降低其对GUV的膜透化活性，从概念验证层面初步表明了NS3作为人工细胞可控物质交换模块的应用潜力。本研究为探索病毒多功能蛋白在人工细胞功能模块构建中的应用提供了初步实验依据。

Abstract

In bottom-up artificial cell construction

building life-like functions such as material exchange

information transmission

and environmental responsiveness from a limited set of molecular components is an important foundation toward more complex behaviors. Multifunctional viral proteins have undergone extensive evolution and often exhibit compact structures

diverse functions

and high conformational plasticity

making them promising components for multifunctional artificial cell modules. In this study

we investigated NS3

a multifunctional protein from human norovirus. NS3 was expressed in vitro using a cell-free protein synthesis (CFPS) system and purified by Ni-NTA affinity chromatography. SDS-PAGE and Western blot analyses confirmed that the CFPS system produced soluble NS3. The purified protein retained both NTPase and RNA helicase activities and also exhibited substantial phospholipid membrane-permeabilizing activity. Further experiments revealed that Mg

differentially regulates these functions: it promotes NS3-mediated RNA unwinding while markedly inhibiting NS3-induced liposome permeabilization. To examine this mechanism in greater detail

we combined sin

gle-molecule fluorescence resonance energy transfer (smFRET) with graphene oxide-based single-molecule surface-induced fluorescence attenuation (smSIFA). These measurements showed that Mg

induces a transition from a relatively open

extended NS3 conformation to a more compact one. Mg

also decreases the depth of NS3 insertion into the membrane without significantly changing its steady-state membrane adsorption density. Together

these findings suggest that conformational rearrangement and altered membrane insertion

rather than reduced membrane adsorption

may underlie the inhibitory effect of Mg

on NS3-mediated membrane permeabilization. Finally

we reconstituted this function in giant unilamellar vesicles (GUVs)

which serve as cell-scale artificial membrane compartments. NS3 promoted the entry of CF into GUVs

whereas Mg

significantly reduced the proportion of GUVs permeabilized by NS3. These results further establish the membrane-permeabilizing function of NS3 and demonstrate that it can be regulated by Mg

in an artificial cell setting. Overall

NS3 combines membrane permeabilization with RNA helicase activity

and these functions can be differentially regulated by Mg

. This study provides preliminary experimental support for using NS3 as a controllable material-exchange module in artificial cells and offers a basis for exploring viral multifunctional proteins as components for artificial cell engineering.

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