Rice University Scientists Engineer Small and Precise CRISPR-Cas13 System

Rice University scientists completed the biochemical and structural characterization one of the smallest CRISPR-Cas13 systems, CRISPR-Cas13bt3, and then used their findings to enhance the precision of the system.

A cryo-electron microscope was used to map the CRISPR system's structure and create a three-dimensional model. Among the findings was that this system deploys a mechanism distinct from other proteins in the Cas13 family. While other proteins in the family initially have two separate domains that come together like scissor blades to perform a cut, CRISPR-Cas13bt3 employs a different strategy. It uses a binding element on two unique loops to connect different parts of the protein and hook onto the RNA strand at the right target site.

The team first worked to determine the structure of the protein and RNA complex, stabilizing it for mapping. Then once they figured out how the system worked, they tweaked the system, increasing its precision and testing its activity and specificity in living cells.

“We found that in cell cultures these systems were able to hone in on a target much easier,” said Sherry Gao, co-author on the study published in Nature Communications. “What is really remarkable about this work is that the detailed structural biology insights enabled a rational determination of the engineering efforts needed to improve the tool’s specificity while still maintaining high on-target RNA editing activity.