CRISPR-Cas9 Structures and Mechanisms.
Level 5 - mechanism / opinion, no new human data
Narrative review of biochemical and structural mechanisms (by design analogy, not clinical CEBM)
PubMed 28375731 · doi:10.1146/annurev-biophys-062215-010822
What was done
This narrative review synthesizes biochemical and structural studies to detail the mechanisms of CRISPR-Cas9-mediated, RNA-guided DNA targeting and cleavage, as well as rational engineering strategies.
What was found
The abstract outlines the sequence of Cas9 action: target recognition strictly requires a flanking protospacer adjacent motif (PAM), followed by R-loop formation and site-specific double-stranded DNA cleavage driven by guide RNA-DNA base pairing, Cas9-DNA interactions, and associated conformational changes. It also notes that synthetic single-guide RNA (sgRNA) simplifies the natural dual tracrRNA-crRNA architecture for genome editing. The abstract provides no quantitative metrics or comparative data.
Why it matters
Elucidating Cas9 conformational changes and DNA-cleavage biochemistry provides a structural blueprint to rationally engineer enzymes with modified PAM requirements, higher guide RNA specificity, and reduced off-target cleavage for genetic therapies.
Limits
As a narrative review, it does not provide original experimental data or systematic search criteria. It focuses on basic structural biology without reporting quantitative error rates, off-target frequencies, or clinical therapeutic outcomes.
Cited by
- supports In the CRISPR-Cas9 system, target DNA sequence specificity is determined by complementary base pairing with a guide RNA molecule.