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Cell penetrating protein adaptor molecules and their application in research and medicine

机译:细胞穿透蛋白衔接子分子及其在研究和医学中的应用

摘要

Cell penetrating peptides (CPPs) are established as a strategy to move cargoes into the interior of eukaryotic cells by engaging import machinery on the cell surface. In most cases the CPP is covalently linked to the cargo; it is common to express cargo proteins with a CPP extension. In some experiments a non-specific interactions (e.g., hydrophobic interactions) have been used to form CPP-cargo complexes, but this has numerous drawbacks. Transport is less efficient and the lack of specificity means that other macromolecules in the medium will also be internalized. This application describes the use of specific CPP labeled adaptor proteins that can be used to move a wide variety of cargoes into the cell interior. The prototype adaptor protein is calmodulin; it is small, stable and easily produced, and binds short (17 amino acid) targets with high affinity in the presence of calcium. A cargo produced with a calmodulin binding tag can be internalized efficiently by CPP tagged calmodulin. Because calcium in the cell interior is typically kept low, CPP-calmodulin releases cargoes inside the cell. The calmodulin binding tag can be used to efficiently purify target proteins by affinity chromatography.
机译:建立细胞穿透肽(CPPs)作为通过在细胞表面上结合导入机制将货物移动到真核细胞内部的策略。在大多数情况下,CPP与货物共价连接。通常会表达具有CPP延伸的货物蛋白。在一些实验中,非特异性相互作用(例如疏水相互作用)已被用于形成CPP-货物复合物,但这具有许多缺点。运输效率较低,缺乏特异性意味着培养基中的其他大分子也将被内化。该申请描述了使用特定的CPP标记的衔接蛋白,其可用于将各种各样的货物移入细胞内部。原型衔接子蛋白是钙调蛋白。它小巧,稳定且易于生产,并且在钙存在下以高亲和力结合短(17个氨基酸)的靶标。带有钙调蛋白结合标签的货物可以通过CPP标记的钙调蛋白有效地内化。由于通常将细胞内部的钙保持在较低水平,因此CPP-钙调蛋白会释放细胞内的货物。钙调蛋白结合标签可用于通过亲和层析有效地纯化靶蛋白。

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