5XN4

Anti-CRISPR protein AcrIIA4

Summary for 5XN4

• Entry DOI: 10.2210/pdb5xn4/pdb
• NMR Information: BMRB: 36085
• Descriptor: Anti-CRISPR AcrIIA4 (1 entity in total)
• Functional Keywords: cas9 inhibitor, hydrolase inhibitor
• Biological source: Listeria monocytogenes
• Total number of polymer chains: 1
• Total formula weight: 10182.07

Authors

Suh, J.-Y.,Kim, I. (deposition date: 2017-05-17, release date: 2018-03-28, Last modification date: 2024-05-15)

Primary citation

Kim, I.,Jeong, M.,Ka, D.,Han, M.,Kim, N.K.,Bae, E.,Suh, J.Y.
Solution structure and dynamics of anti-CRISPR AcrIIA4, the Cas9 inhibitor.
Sci Rep, 8:3883-3883, 2018

PubMed Abstract: The bacterial CRISPR-Cas system provides adaptive immunity against invading phages. Cas9, an RNA-guided endonuclease, specifically cleaves target DNA substrates and constitutes a well-established platform for genome editing. Recently, anti-CRISPR (Acr) proteins that inhibit Cas9 have been discovered, promising a useful off-switch for Cas9 to avoid undesirable off-target effects. Here, we report the solution structure and dynamics of Listeria monocytogenes AcrIIA4 that inhibits Streptococcus pyogenes Cas9 (SpyCas9). AcrIIA4 forms a compact monomeric αβββαα fold comprising three antiparallel β strands flanked by three α-helices and a short 3-helix. AcrIIA4 exhibits distinct backbone dynamics in fast and slow timescales at loop regions that form interaction surfaces for SpyCas9. In particular, the β1-β2 loop that binds to the RuvC domain of SpyCas9 is highly mobile, and the β1-β2 and α2-α3 loops that bind to the RuvC and C-terminal domains of SpyCas9, respectively, undergoes conformational exchanges in microsecond-to-millisecond time scales. AcrIIA4 binds to apo-SpyCas9 with K ~4.8 μM, which compares to K ~0.6 nM for AcrIIA4 binding to sgRNA-bound SpyCas9. Since the binary complex between AcrIIA4 and SpyCas9 does not compete with the target DNA binding, it can effectively disable the Cas9 nuclease activity by forming a tight ternary complex in the presence of sgRNA.

PubMed: 29497118
DOI: 10.1038/s41598-018-22177-0

Experimental method

SOLUTION NMR