Department of Biomedical Engineering, Oregon Health & Science University, Portland, Oregon 97239, United States.
Biochemistry. 2020 Aug 25;59(33):3051-3059. doi: 10.1021/acs.biochem.0c00526. Epub 2020 Aug 17.
Microscopy allows researchers to interrogate proteins within a cellular context. To deliver protein-specific contrast, we developed a new class of genetically encoded peptide tags called versatile interacting peptide (VIP) tags. VIP tags deliver a reporter to a target protein via the formation of a heterodimer between the peptide tag and an exogenously added probe peptide. We report herein a new VIP tag named MiniVIPER, which is comprised of a MiniE-MiniR heterodimer. We first demonstrated the selectivity of MiniVIPER by labeling three cellular targets: transferrin receptor 1 (TfR1), histone protein H2B, and the mitochondrial protein TOMM20. We showed that either MiniE or MiniR could serve as the genetically encoded tag. Next, we demonstrated MiniVIPER's versatility by generating five spectrally distinct probe peptides to label tagged TfR1 on live cells. Lastly, we demonstrated two new applications for VIP tags. First, we used MiniVIPER in combination with another VIP tag, VIPER, to selectively label two different proteins in a single cell (e.g., TfR1 with H2B or TOMM20). Second, we used MiniVIPER to translocate a fluorescent protein to the nucleus through in situ dimerization of mCherry with H2B-mEmerald. In summary, MiniVIPER is a new peptide tag that enables multitarget imaging and artificial dimerization of proteins in cells.
显微镜使研究人员能够在细胞环境中检测蛋白质。为了提供蛋白质特异性对比,我们开发了一类新的称为多功能相互作用肽(VIP)标签的基因编码肽标签。VIP 标签通过肽标签和外源性添加的探针肽之间形成异二聚体将报告蛋白递送到靶蛋白。我们在此报告了一种新的 VIP 标签,称为 MiniVIPER,它由 MiniE-MiniR 异二聚体组成。我们首先通过标记三种细胞靶标:转铁蛋白受体 1(TfR1)、组蛋白蛋白 H2B 和线粒体蛋白 TOMM20,证明了 MiniVIPER 的选择性。我们表明,MiniE 或 MiniR 都可以作为基因编码标签。接下来,我们通过生成五个光谱上不同的探针肽来标记活细胞上标记的 TfR1,展示了 MiniVIPER 的多功能性。最后,我们展示了 VIP 标签的两个新应用。首先,我们将 MiniVIPER 与另一个 VIP 标签 VIPER 结合使用,以在单个细胞中选择性标记两种不同的蛋白质(例如,TfR1 与 H2B 或 TOMM20)。其次,我们使用 MiniVIPER 通过 mCherry 与 H2B-mEmerald 的原位二聚化将荧光蛋白转位到细胞核中。总之,MiniVIPER 是一种新的肽标签,可实现细胞中多靶标成像和蛋白质的人工二聚化。