The selective detection of specific cells of interest and their effective visualization is important but challenging, and fluorescent cell imaging with target-specific probes is commonly used to visualize cell morphology and components and to track cellular processes. Multiple displays of two or more targeting ligands on a polyvalent single template would make it possible to construct versatile multiplex fluorescent cell imaging probes that can visualize two or more target cells individually without the need for a set of individual probes. To achieve this goal, we used encapsulin, a new class of protein cage nanoparticles, as a template and implanted dual targeting capability by presenting two different affibody molecules on a single encapsulin protein cage nanoparticle post-translationally. Encapsulin was self-assembled from 60 identical subunits to form a hollow and symmetric spherical structure with a uniform size. We genetically inserted SpyTag peptides onto the encapsulin surface and prepared various SpyCatcher-fused proteins, such as fluorescent proteins and targeting affibody molecules. We successfully displayed fluorescent proteins and affibody molecules together on a single encapsulin in a mix-and-match manner post-translationally using bacterial superglue, the SpyTag/SpyCatcher ligation system, and demonstrated that these dual functional encapsulins can be used as target-specific fluorescent cell imaging probes. Dual targeting protein cage nanoparticles were further constructed by ligating two different affibody molecules onto the encapsulin surface with fluorescent dyes, and they effectively recognized and bound to two individual targeting cells independently, which could be visualized by selective colors on demand.

中文翻译：

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使用细菌超级胶水设计可调节的双功能蛋白笼纳米颗粒。

选择性检测感兴趣的特定细胞及其有效的可视化很重要，但是具有挑战性，通常使用带有靶标特异性探针的荧光细胞成像来可视化细胞形态和成分并跟踪细胞过程。在多价单一模板上多次显示两个或多个靶向配体将使构建通用的多重荧光细胞成像探针成为可能，该探针可单独显示两个或多个靶细胞，而无需一组单独的探针。为了实现这一目标，我们使用新型新型蛋白笼纳米颗粒衣壳蛋白作为模板，并通过在翻译后的单个衣壳蛋白笼子纳米颗粒上呈现两个不同的亲和分子，从而植入了双重靶向能力。荚膜蛋白由60个相同的亚基自组装而成，形成大小均匀的中空对称球形结构。我们将SpyTag肽基因插入胶囊蛋白表面，并制备了各种SpyCatcher融合蛋白，例如荧光蛋白和靶向亲和分子。我们使用细菌强力胶SpyTag / SpyCatcher连接系统在翻译后通过混合匹配的方式成功地在单个胶囊上展示了荧光蛋白和亲和分子，并证明了这些双重功能的胶囊可以用作目标特异性荧光细胞成像探针。通过使用荧光染料将两个不同的亲和分子连接到胶囊蛋白表面上，进一步构建了双重靶向蛋白笼纳米颗粒，