Dynamics and mechanisms of CRISPR-Cas9 through the lens of computational methods

• Published in: Current opinion in structural biology 2022-08, Vol.75, p.102400-102400, Article 102400
• Main Authors: Saha, Aakash, Arantes, Pablo R., Palermo, Giulia
• Format: Article
• Language: English
• Subjects: CRISPR-Associated Protein 9; CRISPR-Cas Systems; DNA Cleavage; Gene Editing - methods; Molecular Dynamics Simulation
• ISSN: 0959-440X; 1879-033X; 1879-033X
• DOI: 10.1016/j.sbi.2022.102400

The clustered regularly interspaced short palindromic repeat (CRISPR) genome-editing revolution established the beginning of a new era in life sciences. Here, we review the role of state-of-the-art computations in the CRISPR-Cas9 revolution, from the early refinement of cryo-EM data to enhanced simulations of large-scale conformational transitions. Molecular simulations reported a mechanism for RNA binding and the formation of a catalytically competent Cas9 enzyme, in agreement with subsequent structural studies. Inspired by single-molecule experiments, molecular dynamics offered a rationale for the onset of off-target effects, while graph theory unveiled the allosteric regulation. Finally, the use of a mixed quantum-classical approach established the catalytic mechanism of DNA cleavage. Overall, molecular simulations have been instrumental in understanding the dynamics and mechanism of CRISPR-Cas9, contributing to understanding function, catalysis, allostery, and specificity. [Display omitted] •Molecular dynamics reveals the conformational activation of CRISPR-Cas9.•Simulations predict an activated state confirmed by cryo-EM structures.•Enhanced simulations establish a mechanistic rationale for off-target DNA binding.•Graph theory decrypts the allosteric modulation.•Ab-initio methods unveil the catalytic mechanism of DNA cleavage.