CAS9: INSIDE THE MAGIC BOX

1

Proteins such as Cas9 are made up of strings of amino acids which form certain structures – alpha helices (coils), beta sheets (flat ribbons) and joining loops. These structures combine to form domains, each of which have a particular function. Cas proteins are able to cut DNA at a specific sequence. In Cas9 the process begins when the largest domain, REC I (beige), binds to a guide RNA.

2

When the RNA binds to Cas9 it changes the protein’s shape into an “active” form. Next

Cas9 goes looking for a key section of DNA: the protospacer adjacent motif PAM (yellow stars). The PAM is a short sequence of DNA just two or three bases long near the target section. Another domain, the Pam-interacting domain (orange) binds to the PAM.

3

Once bound to the PAM, Cas9 will separate the strands of DNA near the PAM, and check if the DNA sequence matches the complementary region of the guide RNA. If the DNA is a good match, the HNH (blue) and Ruvc (yellow) will cut or cleave the target DNA.

The bridge helix domain (magenta) also helps with the processes of binding the guide RNA and the target DNA and cleaving the target DNA. The function of some domains is better understood than others; the role of RECII (grey) in this process is not yet clear.