The three-dimensional structure 

Cas9 is the most powerful gene editing enzymy that have been widely used around the world since its first usage report published in 2012. Its x-ray crystallography showed that there are two function domains that responsible for DNA cleavage which locate in its conserved structure. Besides, the structurally divergent regions, including the PAM recognition loops, are likely responsible for distinct guide RNA and PAM specificities. Cas9 enzymes adopt a catalytically inactive conformation in the apo state, neces- sitating structural activation for DNA recognition and cleavage. Our EM analysis shows that by triggering a conformational rearrangement in Cas9, the guide RNA acts as a critical determinant of target DNA binding.