AI Article Synopsis
- CRISPR-Cas12a is an RNA-guided enzyme that creates double-strand breaks in DNA by first cutting a non-target strand and then the target strand.
- Research using single-molecule FRET revealed that the 3' overhangs of both the non-target strand and the guide RNA (crRNA) block the target strand from entering the enzyme's active site.
- The study provides valuable insights into how these overhangs affect the cleavage process of Cas12a, which could inform future strategies for regulating its activity.
Article Abstract
CRISPR-Cas12a, an RNA-guided DNA endonuclease, induces double-strand breaks by cleaving the non-target strand (NTS) first, followed by the target strand (TS). Using single-molecule FRET with alternating-laser excitation, we found that steric hindrance from the 3' overhangs of both the cleaved NTS and crRNA impedes TS loading into the catalytic core. Our study highlights the direct involvement of both 3' NTS and crRNA overhangs in TS cleavage, offering insights into regulatory strategies for Cas12a cleavage reactions.
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| http://dx.doi.org/10.1039/d4cc04716h | DOI Listing |
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