An, So Young Ka, Donghyun Kim, Iktae Kim, Eun-Hee Kim, Nak-Kyoon Bae, Euiyoung Suh, Jeong-Yong 2024-01-19T17:02:50Z 2024-01-19T17:02:50Z 2021-09-04 2020-07-27 0305-1048 https://pubs.kist.re.kr/handle/201004/118362 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 Strepto-coccus 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 alpha/beta 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. English OXFORD UNIV PRESS PROTEIN CLASSIFICATION SYSTEM MECHANISMS EVOLUTION DISCOVERY BACTERIA IMPACT ANGLES GENES Intrinsic disorder is essential for Cas9 inhibition of anti-CRISPR AcrIIA5 Article 10.1093/nar/gkaa512 1 NUCLEIC ACIDS RESEARCH, v.48, no.13, pp.7584 - 7594 NUCLEIC ACIDS RESEARCH 48 13 7584 7594 scie scopus 000574293400046 2-s2.0-85088495716 Biochemistry & Molecular Biology Biochemistry & Molecular Biology Article PROTEIN CLASSIFICATION SYSTEM MECHANISMS EVOLUTION DISCOVERY BACTERIA IMPACT ANGLES GENES