Protein transduction domains (PTDs) are versatile peptide sequences that facilitate cell delivery of several cargo molecules including proteins. PTDs usually consist of short stretches of basic amino acids that can cross the plasma membrane and gain entry into cells. Traditionally, to assess PTD mediated protein delivery, PTD-fusion proteins have been used as purified proteins. To overcome the requirement for a protein purification step, we used a secretory signal peptide to allow PTD-CRE fusion proteins to be exported from transfected mammalian cells. PTD induced protein transduction into cells was assessed by a CRE-mediated recombination event that resulted in beta-galactosidase expression. Several PTDs were tested including the prototypic TAT, different TAT variants, Antp, MTS and polyarginine. A negative correlation was observed between the cationic charge on the PTD and the extent of secretion. Poor secretion was found when the PTD charge was greater than +5. One TAT-CRE protein variant had a 14-fold enhancement above CRE alone when added to cells in the presence of chloroquine. This PTD domain also enhanced gene expression after plasmid delivery. These data illustrate that some secreted PTD proteins may be useful reagents to improve protein delivery in mammalian systems and a novel approach to enhancing the response to DNA transfections.

中文翻译：

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介导分泌的CRE蛋白的细胞传递中蛋白质转导域的比较。

蛋白质转导域（PTD）是通用的肽序列，可促进包括蛋白质在内的几种货物分子的细胞递送。PTD通常由一小段碱性氨基酸组成，可以穿过质膜并进入细胞。传统上，为了评估PTD介导的蛋白传递，已将PTD融合蛋白用作纯化蛋白。为了克服蛋白质纯化步骤的要求，我们使用了分泌信号肽以允许PTD-CRE融合蛋白从转染的哺乳动物细胞中输出。通过CRE介导的重组事件（导致β-半乳糖苷酶表达）评估了PTD诱导的蛋白向细胞的转导。测试了几个PTD，包括原型TAT，不同的TAT变体，Antp，MTS和聚精氨酸。在PTD上的阳离子电荷和分泌程度之间观察到负相关。当PTD电荷大于+5时，发现分泌不良。当在存在氯喹的情况下添加到细胞中时，一种TAT-CRE蛋白变体比单独的CRE具有14倍的增强。该PTD结构域还增强了质粒递送后的基因表达。这些数据表明，某些分泌的PTD蛋白可能是改善哺乳动物系统中蛋白传递的有用试剂，并且是增强对DNA转染反应的新颖方法。该PTD结构域还增强了质粒递送后的基因表达。这些数据表明，某些分泌的PTD蛋白可能是改善哺乳动物系统中蛋白传递的有用试剂，并且是增强对DNA转染反应的新颖方法。该PTD结构域还增强了质粒递送后的基因表达。这些数据表明，某些分泌的PTD蛋白可能是改善哺乳动物系统中蛋白传递的有用试剂，并且是增强对DNA转染反应的新颖方法。