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Structural Plasticity of PAM Recognition by Engineered Variants of the RNA-Guided Endonuclease Cas9
Changes in DNA Target Recognition by Modified Versions of the Gene-Editing Protein Cas9
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Abstract
Crystal structures of SpCas9 variants reveal an induced fit mechanism for recognizing non-canonical PAMs.
- DNA cleavage by SpCas9 requires a specific 5'-NGG-3' protospacer adjacent motif (PAM), limiting target sequences.
- Artificial SpCas9 variants with altered PAM specificities have been developed to overcome these limitations.
- The structures of VQR, EQR, and VRER variants bound to their preferred PAM sequences show conformational changes in the PAM region of the DNA.
- Amino acid substitutions in these SpCas9 variants enhance sequence-specific base recognition and conformational remodeling.
- A new SpCas9 variant was engineered to specifically recognize NAAG PAMs based on structural insights.
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