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Nucleic Acids Res., Published: 16 June 2020, gkaa512 | https://doi.org/10.1093/nar/gkaa512 Intrinsic disorder is essential for Cas9 inhibition of anti-CRISPR AcrIIA5

So Young An¹, Donghyun Ka¹, Iktae Kim¹, Eun-Hee Kim², Nak-Kyoon Kim³, Euiyoung Bae¹ and Jeong-Yong Suh^1,4,*

¹Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul 08826, South Korea
²Protein Structure Research Team, Korea Basic Science Institute, 162 Yeongudanji-Ro, Ochang, Chungbuk 28119, South Korea
³Advanced Analysis Center, Korea Institute of Science and Technology, Seoul 02792, South Korea
⁴Institute for Biomedical Sciences, Shinshu University, Minamiminowa, Nagano 399-4598, Japan

^*Corresponding author

Abstract

Clustered regularly interspaced short palindromic repeats (CRISPRs) and CRISPR-associated (Cas) proteins provide adaptive immunity to prokaryotes against invading phages and plasmids. As a countermeasure, phages have evolved anti-CRISPR (Acr) proteins that neutralize the CRISPR immunity. AcrIIA5, isolated from a virulent phage of Streptococcus thermophilus, strongly inhibits diverse Cas9 homologs, but the molecular mechanism underlying the Cas9 inhibition remains unknown. Here, we report the solution structure of AcrIIA5, which features a novel α/β fold connected to an N-terminal intrinsically disordered region (IDR). Remarkably, truncation of the N-terminal IDR abrogates the inhibitory activity against Cas9, revealing that the IDR is essential for Cas9 inhibition by AcrIIA5. Progressive truncations and mutations of the IDR illustrate that the disordered region not only modulates the association between AcrIIA5 and Cas9–sgRNA, but also alters the catalytic efficiency of the inhibitory complex. The length of IDR is critical for the Cas9–sgRNA recognition by AcrIIA5, whereas the charge content of IDR dictates the inhibitory activity. Conformational plasticity of IDR may be linked to the broad-spectrum inhibition of Cas9 homologs by AcrIIA5. Identification of the IDR as the main determinant for Cas9 inhibition expands the inventory of phage anti-CRISPR mechanisms.

논문정보

형식Research article
게재일2020년 06월 (BRIC 등록일 2020-06-26)
연구진국내(교신)+국외 연구진
분야 생명과학 > 구조생물 및 생물물리학

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