Holding something real
There’s nothing like hands-on experience to figure out how something works. In that spirit, I worked with 3D Molecular Designs to get a deconstructable, 3D printed model of Cas9 doing its thing. The model is based on the Jinek lab’s recent structure of Cas9 bound to PAM-containing DNA, with some artistic license to build in the full non-complementary strand and the upstream and downstream duplexes. I think the model tuned out extremely well and beautifully illustrates how Cas9 tightly binds DNA by using the sgRNA protospacer. It also very concretely highlights some interesting mysteries about the structure, such as the HNH active site pointing towards the wrong DNA strand. Just the few hours I’ve been playing with this model have led to some neat insights.
Edit: correctly linked to 3D Molecular Designs instead of MSOE (sister organizations).
Cas9 bound to sgRNA and DNA target
HnH domain removed (sgRNA in orange, DNA in blue)
|The DNA and sgRNA can be removed from the model (PAM in green)||The DNA strands can even be separated at the actual Cas9 cleavage site|