CRISPR loci and Cas proteins provide adaptive immunity in prokaryotes against invading bacteriophages and plasmids. In response, bacteriophages have evolved a broad spectrum of anti-CRISPR proteins (anti-CRISPRs) to counteract and overcome this immunity pathway. Numerous anti-CRISPRs have been identified to date, which suppress single-subunit Cas effectors (in CRISPR class 2, type II, V and VI systems) and multisubunit Cascade effectors (in CRISPR class 1, type I and III systems). Crystallography and cryo-electron microscopy structural studies of anti-CRISPRs bound to effector complexes, complemented by functional experiments in vitro and in vivo, have identified four major CRISPR–Cas suppression mechanisms: inhibition of CRISPR–Cas complex assembly, blocking of target binding, prevention of target cleavage, and degradation of cyclic oligonucleotide signalling molecules. In this Review, we discuss novel mechanistic insights into anti-CRISPR function that have emerged from X-ray crystallography and cryo-electron microscopy studies, and how these structures in combination with function studies provide valuable tools for the ever-growing CRISPR–Cas biotechnology toolbox, to be used for precise and robust genome editing and other applications.

CRISPR 基因座和 Cas 蛋白在原核生物中提供针对入侵的噬菌体和质粒的适应性免疫。作为回应，噬菌体已经进化出广谱的抗 CRISPR 蛋白（抗 CRISPR）来抵消和克服这种免疫途径。迄今为止，已经鉴定出许多抗 CRISPR，它们抑制单亚基 Cas 效应器（在 CRISPR 2 类、II、V 和 VI 型系统中）和多亚基级联效应器（在 CRISPR 1 类、I 型和 III 型系统中）。结合效应复合物的抗 CRISPR 的晶体学和冷冻电子显微镜结构研究，辅以体外和体内的功能实验，确定了四种主要的 CRISPR-Cas 抑制机制：抑制 CRISPR-Cas 复合物组装、阻断靶标结合、防止目标裂解，和环状寡核苷酸信号分子的降解。在这篇综述中，我们讨论了从 X 射线晶体学和冷冻电子显微镜研究中出现的关于抗 CRISPR 功能的新机制见解，以及这些结构与功能研究相结合如何为不断发展的 CRISPR-Cas 生物技术提供有价值的工具工具箱，用于精确和强大的基因组编辑和其他应用程序。