Mechanism of RNA-independent cleavage of CRISPR-Cas9
| dc.contributor.author | Liu, Jin | |
| dc.contributor.author | Chen, Xiongping | |
| dc.contributor.author | Hayatshahi, Sayyed | |
| dc.creator | Wang, Duen-Shian | |
| dc.date.accessioned | 2020-12-13T17:16:28Z | |
| dc.date.available | 2020-12-13T17:16:28Z | |
| dc.date.issued | 2020 | |
| dc.description.abstract | Purpose: CRISPR(Clustered-Regularly-interspaced-short-palindromic-repeats)-Cas9 (CRISPR-associated-protein-9) uncovered a new path toward gene therapy. However, non-specific cleavage of Cas9 raises concerns on human therapeutic applications so that it is critical to understand and minimize those non-specifics cuttings. Recent in vitro studies showed that Cas9 cleavage occurred even without the guidance from the guide RNA (gRNA) in the presence of Mn2+, implying the serious issue of off-target effect of Cas9. The purpose of this study is to elucidate the mechanism of the RNA-independent cleavage of CRISPR-Cas9, which may provide insights for the improvement of Cas9 specificity. Method: Based on our previously captured structure of catalytically-active Cas9-gRNA-dsDNA complex, we performed molecular dynamic (MD) simulations on Cas9 complexed with and without gRNA, respectively. We also compared the simulations in the presence of Mn2+vs. Mg2+. All MD simulations were performed using AMBER package with GPU acceleration. Result: We expect our MD simulations to demonstrate the different coordination environments of Mn2+ and Mg2+ in the presence or absence of gRNA, elucidating a novel mechanism for Cas9 off-target effects. Conclusion: In this study, we expect to identify the mechanism of RNA-independent cleavage of CRISPR-Cas9, shedding light on the development of new Cas9 variants to reduce off-target effects. | |
| dc.identifier.uri | https://hdl.handle.net/20.500.12503/30335 | |
| dc.language.iso | en | |
| dc.title | Mechanism of RNA-independent cleavage of CRISPR-Cas9 | |
| dc.type | poster | |
| dc.type.material | text |