The introduction of the tool for gene editing, the so-called gene scissors CRISPR, in 2007 was a revolution within medical science and cell biology. But even though the perspectives are great, the launch of CRISPR has been followed by debate, especially focussing on ethical issues and the technology's degree of accuracy and side effects.
However, in a new study published in the scientific journal Cell researchers from the Novo Nordisk Foundation Center for Protein Research have described how one of the CRISPR technologies, the so-called Cas12a, works - all the way down to the molecular level. This makes it possible to fine-tune the gene-editing process to only achieve the desired effects.
'If we compare CRISPR to a car engine, what we have done is make a complete 3D map of the engine and thus gained an understanding of how it works. This knowledge will enable us to fine-tune the CRISPR engine and make it work in various ways - as a Formula 1 racer as well as an off-road truck', says Professor Guillermo Montoya from the Novo Nordisk Foundation Center for Protein Research.
Molecular Film
The researchers have used a so-called cryo-electron microscope to map the technology. The recently inaugurated cryoEM facility at the University of Copenhagen has established the state-of-the-art technology enabling the researchers to take photographs of the different shapes of the molecule when CRISPR-Cas12a cuts up the DNA strand.
They combined it with a fluorescent microscopy technique called 'single molecule FRET' that directly observes the motions of the molecules and the sequence of events for each individual protein.
Among other things, this sequence of events revealed to the researchers that three "pieces" of the CRISPR tools must change form for the DNA to be cut properly.
'Our new study shows the precise series of events in the genome leading to gene editing. These three "pieces" that change, work like airport security checks. You have to complete all checks and in the right order to proceed', says Associate Professor Nikos Hatzakis from the Department of Chemistry and the Nano-Science center.
More Editing than Desired According to the researchers, their new findings can explain why CRISPR technology can have side effects on the genome. Once the DNA strand has been cut, the three 'security checks' remain open.
This can cause the process to last longer than wanted, because the machinery behind gene editing continues to run and can cause genetic changes.
However, now the researchers expect their new knowledge to put an end to this. They believe it can be used to fine-tune the gene-editing technology right away.
###
The study is funded by the Novo Nordisk Foundation, the Carlsberg Foundation and The Villum Foundation.
Both research groups are members of the Integrative Structural Biology Cluster (ISBUC) at University of Copenhagen.
Read the study: 'Conformational Activation Promotes CRISPR-Cas12a Catalysis and Resetting of the Endonuclease Activity' at Cell.
Journal
Cell