Greenwood Kathryn P, Daly Norelle L, Brown Darren L, Stow Jennifer L, Craik David J
Institute for Molecular Bioscience and Australian Research Council Special Research Centre for Functional and Applied Genomics, The University of Queensland, Brisbane, Queensland 4072, Australia.
Int J Biochem Cell Biol. 2007;39(12):2252-64. doi: 10.1016/j.biocel.2007.06.016. Epub 2007 Jul 7.
The cyclotides are macrocyclic knotted proteins characterized by a compact topology and exceptional stability. Accordingly it has been hypothesized that they may be useful as protein engineering frameworks for the stabilization and delivery of bioactive peptide sequences. This study examined the internalization of cyclotides into mammalian cells, a vital step for the delivery of bioactive peptide sequences to intracellular targets. Although the entry of various linear peptides into cells has been reported previously, this is the first report of internalization of a macrocyclic peptide. Cell uptake was examined for representatives of two cyclotide subfamilies; the first was MCoTI-II, a member of the trypsin inhibitor subfamily, which was internalized by a macrophage and breast cancer cell line and the second, the prototypic cyclotide kalata B1 from the Möbius subfamily, which remained extracellular. Biotin labeled MCoTI-II entered macrophages by macropinocytosis, resulting in vesicular encapsulation without trafficking to lysosomes for degradation. The ready uptake, coupled with low cytotoxicity, indicates that MCoTI-II has the potential to transport grafted bioactivities to intracellular targets, making it a potentially valuable framework in drug design applications.
环肽是一种大环打结蛋白，其特点是拓扑结构紧凑且稳定性极高。因此，有人推测它们可能作为蛋白质工程框架，用于稳定和递送生物活性肽序列。本研究检测了环肽进入哺乳动物细胞的情况，这是将生物活性肽序列递送至细胞内靶点的关键步骤。虽然此前已有各种线性肽进入细胞的报道，但这是大环肽内化的首次报道。研究了两个环肽亚家族代表的细胞摄取情况；第一个是胰蛋白酶抑制剂亚家族的成员MCoTI-II，它可被巨噬细胞和乳腺癌细胞系内化，第二个是来自莫比乌斯亚家族的原型环肽kalata B1，它停留在细胞外。生物素标记的MCoTI-II通过巨胞饮作用进入巨噬细胞，导致其被囊泡包裹，且不会转运至溶酶体进行降解。易于摄取，再加上低细胞毒性，表明MCoTI-II有潜力将嫁接的生物活性物质转运至细胞内靶点，使其成为药物设计应用中一个潜在有价值的框架。
