Tumorigenic risk of undifferentiated human induced pluripotent stem cells (iPSCs), being a major obstacle for clinical application of iPSCs, requires novel approaches for selectively eliminating undifferentiated iPSCs. Here, we show that an l-phosphopentapeptide, upon the dephosphorylation catalyzed by alkaline phosphatase (ALP) overexpressed by iPSCs, rapidly forms intranuclear peptide assemblies made of α-helices to selectively kill iPSCs. The phosphopentapeptide, consisting of four l-leucine residues and a C-terminal l-phosphotyrosine, self-assembles to form micelles/nanoparticles, which transform into peptide nanofibers/nanoribbons after enzymatic dephosphorylation removes the phosphate group from the l-phosphotyrosine. The concentration of ALP and incubation time dictates the morphology of the peptide assemblies. Circular dichroism and FTIR indicate that the l-pentapeptide in the assemblies contains a mixture of an α-helix and aggregated strands. Incubating the l-phosphopentapeptide with human iPSCs results in rapid killing of the iPSCs (=<2 h) due to the significant accumulation of the peptide assemblies in the nuclei of iPSCs. The phosphopentapeptide is innocuous to normal cells (e.g., HEK293 and hematopoietic progenitor cell (HPC)) because normal cells hardly overexpress ALP. Inhibiting ALP, mutating the l-phosphotyrosine from the C-terminal to the middle of the phosphopentapeptides, or replacing l-leucine to d-leucine in the phosphopentapeptide abolishes the intranuclear assemblies of the pentapeptides. Treating the l-phosphopentapeptide with cell lysate of normal cells (e.g., HS-5) confirms the proteolysis of the l-pentapeptide. This work, as the first case of intranuclear assemblies of peptides, not only illustrates the application of enzymatic noncovalent synthesis for selectively targeting nuclei of cells but also may lead to a new way to eliminate other pathological cells that express a high level of certain enzymes.

中文翻译：

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酶促形成核内肽组装体以选择性杀死人类诱导的多能干细胞

未分化的人诱导多能干细胞 (iPSCs) 的致瘤风险是 iPSCs 临床应用的主要障碍，需要新的方法来选择性地消除未分化的 iPSCs。在这里，我们展示了一种l-磷酸五肽，在 iPSC 过表达的碱性磷酸酶 (ALP) 催化的去磷酸化作用下，迅速形成由 α-螺旋构成的核内肽组装体，以选择性地杀死 iPSC。由四个l-亮氨酸残基和一个 C-末端l-磷酸酪氨酸组成的磷酸五肽自组装形成胶束/纳米颗粒，在酶促去磷酸化后将磷酸基团从l--磷酸酪氨酸。ALP 的浓度和孵育时间决定了肽组件的形态。圆二色性和 FTIR 表明组件中的l-五肽包含 α-螺旋和聚集链的混合物。由于肽组装体在 iPSCs 细胞核中的显着积累，将l-磷酸五肽与人 iPSCs 一起温育导致 iPSCs 的快速杀死（=<2 小时）。磷酸五肽对正常细胞（例如，HEK293 和造血祖细胞 (HPC)）是无害的，因为正常细胞几乎不会过度表达 ALP。抑制 ALP，将l-磷酸酪氨酸从 C 端突变到磷酸五肽的中间，或将l-亮氨酸替换为d磷酸五肽中的-亮氨酸消除了五肽的核内组装。用正常细胞的细胞裂解物（例如，HS-5）处理L-磷酸五肽证实了L-五肽的蛋白水解。这项工作作为第一个肽核内组装的案例，不仅说明了酶促非共价合成在选择性靶向细胞核中的应用，而且可能导致一种新的方法来消除其他表达高水平某些酶的病理细胞。