Fig. 2

Structural analysis of inactivated, activated and activated cysteine-to-serine Cas12a mutant RNPs on the basis of AlphaFold3. A Structural comparison between the simulation model based on AlphaFold3 via inactivated Cas12a RNP (deepteal) and activated Cas12a RNP (purple) (RMSD = 3.133). B Molecular simulation estimates of distance changes between Val377 and Gln1136 due to changes in the Cas12a conformation before and after activation by AlphaFold3. The Val377–Gln1136 distance in inactivated RNPs is 24.22 Å, and in activated RNPs, it is 32.55 Å. Green: Cas12a protein; blue: nuclease domain (NUC) in Cas12a; brown: gRNA; slate: ssDNA trigger. C Structural comparison between the simulation model based on AlphaFold3 using the activated Cas12a RNP (purple) and the cysteine-to-serine Cas12a mutant RNP (smudge) (RMSD = 2.082). D Molecular simulation estimates of distance changes between Val377 and Gln1136 due to the Cas12a conformation changes resulting from cysteine to serine mutations by AlphaFold3. The Val377–Gln1136 distance from the activated cysteine-to-serine Cas12a mutant RNP is 42.87 Å, which is greater than the 32.55 Å distance in the activated Cas12a RNP. Green: Cas12a protein; blue: nuclease domain (NUC) in Cas12a; brown: gRNA; slate: ssDNA trigger