基于碱基切除修复的 DNA 马达,由细胞内脱嘌呤/脱嘧啶核酸内切酶驱动。

Base excision repair-inspired DNA motor powered by intracellular apurinic/apyrimidinic endonuclease.

机构信息

College of Life Science and Technology, Beijing University of Chemical Technology, Beijing 100029, China.

出版信息

Nanoscale. 2019 Jan 17;11(3):1343-1350. doi: 10.1039/c8nr07813k.

Abstract

The transition of DNA nanomachines from test tubes to living cells would realize the ultimate goal of smart therapeutic dynamic DNA nanotechnology. The operation of DNA nanomachines in living cells remains challenging because it is difficult to utilize an endogenous driving force. Inspired by the base excision repair (BER) process, we demonstrate a 'burnt-bridge' DNA motor system powered by intracellular apurinic/apyrimidinic (AP) endonuclease APE1. The high specificity of APE1 to the AP site in double-stranded DNA permits directional and autonomous movement. The advanced single-molecule fluorescence technique was utilized to directly monitor the stepwise movement of the motor strand, confirming the excellent controllability and processivity of this system. The speed of this DNA motor relies highly on APE1 concentration, allowing discrimination by APE1 level against cancer cells and normal cells. Western blot was used to confirm APE1 expression level. Successful operation of the DNA motor in living cells demonstrates that an endogenous enzyme can operate the rationally designed DNA nanostructures in a programmable way, rather than digesting simple molecular probes. This is useful and practicable for broad application, such as for cellular diagnostic tools, gene regulators for DNA repair, and enzyme-mediated drug delivery.

摘要

DNA 纳米机器从试管到活细胞的转变将实现智能治疗动态 DNA 纳米技术的最终目标。由于难以利用内源性驱动力,因此 DNA 纳米机器在活细胞中的操作仍然具有挑战性。受碱基切除修复 (BER) 过程的启发,我们展示了一种由细胞内脱嘌呤/脱嘧啶内切酶 APE1 驱动的“烧桥” DNA 马达系统。APE1 对双链 DNA 中 AP 位点的高特异性允许定向和自主运动。先进的单分子荧光技术被用于直接监测马达链的逐步运动,证实了该系统具有出色的可控性和连续性。该 DNA 马达的速度高度依赖于 APE1 浓度,允许通过 APE1 水平区分癌细胞和正常细胞。Western blot 用于确认 APE1 的表达水平。该 DNA 马达在活细胞中的成功运行表明,内源性酶可以以可编程的方式操作合理设计的 DNA 纳米结构,而不是消化简单的分子探针。这对于广泛的应用是有用且可行的,例如用于细胞诊断工具、用于 DNA 修复的基因调节剂和酶介导的药物输送。

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