Department of Chemistry & Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, MA 02138, USA; Howard Hughes Medical Institute, Harvard University, 12 Oxford Street, Cambridge, MA 02138, USA.
Department of Chemistry & Chemical Biology, Harvard University, 12 Oxford Street, Cambridge, MA 02138, USA; Howard Hughes Medical Institute, Harvard University, 12 Oxford Street, Cambridge, MA 02138, USA.
Cell Chem Biol. 2016 Jan 21;23(1):57-73. doi: 10.1016/j.chembiol.2015.12.009.
Programmable DNA nucleases have provided scientists with the unprecedented ability to probe, regulate, and manipulate the human genome. Zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and the clustered regularly interspaced short palindromic repeat-Cas9 system (CRISPR-Cas9) represent a powerful array of tools that can bind to and cleave a specified DNA sequence. In their canonical forms, these nucleases induce double-strand breaks at a DNA locus of interest that can trigger cellular DNA repair processes that disrupt or replace genes. The fusion of these programmable nucleases with a variety of other protein domains has led to a rapidly growing suite of tools for activating, repressing, visualizing, and modifying loci of interest. Maximizing the usefulness and therapeutic relevance of these tools, however, requires precisely controlling their activity and specificity to minimize potentially toxic side effects arising from off-target activities. This need has motivated the application of chemical biology principles and methods to genome-editing proteins, including the engineering of variants of these proteins with improved or altered specificities, and the development of genetic, chemical, optical, and protein delivery methods that control the activity of these agents in cells. Advancing the capabilities, safety, effectiveness, and therapeutic relevance of genome-engineering proteins will continue to rely on chemical biology strategies that manipulate their activity, specificity, and localization.
可编程 DNA 核酸酶为科学家提供了前所未有的能力，可用于探测、调节和操纵人类基因组。锌指核酸酶 (ZFNs)、转录激活因子样效应核酸酶 (TALENs) 和成簇规律间隔短回文重复系统-Cas9（CRISPR-Cas9）代表了一系列强大的工具，可以结合并切割特定的 DNA 序列。在其典型形式中，这些核酸酶在感兴趣的 DNA 基因座上诱导双链断裂，从而触发细胞 DNA 修复过程，破坏或替换基因。这些可编程核酸酶与各种其他蛋白结构域的融合导致了一系列用于激活、抑制、可视化和修饰感兴趣基因座的工具迅速发展。然而，为了最大限度地提高这些工具的有用性和治疗相关性，需要精确控制其活性和特异性，以最小化潜在的脱靶活性引起的毒性副作用。这种需求促使人们将化学生物学原理和方法应用于基因组编辑蛋白，包括对这些蛋白进行工程改造，以提高或改变其特异性，并开发遗传、化学、光学和蛋白递送方法，以控制这些试剂在细胞中的活性。推进基因组工程蛋白的功能、安全性、有效性和治疗相关性将继续依赖于操纵其活性、特异性和定位的化学生物学策略。
