10VC

Crystal structure of GeoCas9 HNH domain bound to anti-CRISPR AcrIIC1

Summary for 10VC

• Entry DOI: 10.2210/pdb10vc/pdb
• Descriptor: CRISPR-associated endonuclease Cas9, Anti-CRISPR protein (AcrIIC1) (3 entities in total)
• Functional Keywords: nuclease, cas9, inhibitor, anti-crispr, crispr, dna binding protein
• Biological source: Geobacillus stearothermophilus; More
• Total number of polymer chains: 12
• Total formula weight: 136299.88

Authors

Knight, A.L.,D'Ordine, A.M.,Lisi, G.P.,Jogl, G. (deposition date: 2026-02-10, release date: 2026-04-29)

Primary citation

Knight, A.L.,Belato, H.B.,Dresser, C.S.,Pindi, C.,Mercado, B.J.,Lasekan, P.,Luo, J.,Arantes, P.R.,Jogl, G.,Palermo, G.,Lisi, G.P.
Orthosteric and allosteric effects of anti-CRISPR II-C1 inhibition on Geo Cas9 from integrated structural biophysics.
Biorxiv, 2026

PubMed Abstract: Anti-CRISPRs (Acrs) are small protein inhibitors of CRISPR-Cas effectors that originate from the translated genetic material of bacteriophage. Harnessing the natural ability of Acrs to bind and disrupt CRISPR-Cas editing can provide enhanced spatiotemporal control of gene editing. Recent studies have revealed diverse structures and functions of Acrs, however, atomistic studies of the specific molecular mechanisms behind Acr inhibition are lacking. Here, we reveal how structure, function, and dynamics govern AcrIIC1 inhibition of Cas9 from ( Cas9) via its HNH nuclease domain. An X-ray crystal structure of the HNH-AcrIIC1complex reveals a conserved binding interface at the catalytic site and disruption of crucial electrostatic contacts known to modulate the thermostability of Cas9. AcrIIC1 binding also rewires the intrinsic dynamics of the HNH domain, stimulates millisecond motions that are absent from the unliganded nuclease, and attenuates the guide RNA affinity of Cas9. Subsequent AcrIIC1 mutations in residues at its crystallographic binding interface uncouple Acr binding from inhibition, providing new insight into mechanism by which AcrIIC1 acts on Cas9.

PubMed: 41993526
DOI: 10.64898/2026.04.08.717222

Experimental method

X-RAY DIFFRACTION (1.79 Å)