News Office, MIT, December 1, 2015. http://news.mit.edu/2015/overcome-crispr-cas9-genome-editing-hurdle-1201
One of the major obstacles facing wider adoption of CRISPR/Cas9 systems as a gene editing and gene therapy technique is the off target cleavage of DNA. In a report published in Science (http://www.sciencemag.org/content/early/2015/11/30/science.aad5227.abstract) MIT researchers used rational protein engineering to improve CRISPR/Cas9 specificity. Using the crystal structure of the CRISPR/Cas9 complex researchers identified a positively charged groove that is able to stabilize binding of negatively charged DNA. By changing three of the positive amino acids found in this groove to neutral amino acids the researchers reasoned that DNA binding would become more dependent on proper Watson-Crick base pairing between the CRISPR guide RNA and the targeted DNA. This improved Cas9, coined “enhanced specificity” SpCas9 or eSpCas9, eliminated 22 of the 24 off-target events observed when compared to wild-type Cas9 treatment.