Groundbreaking study seeks to illuminate how DNA-editing protein Cas9 interacts with genetic material

Opening the door to new ways of studying gene editing proteins and potential biomedical applications like improved diagnostic technologies, Professor Nicole Weckman (ISTEP, ChemE) has co-authored a new study entitled, Sensing the DNA-mismatch tolerance of catalytically inactive Cas9 via barcoded DNA nanostructures in solid-state nanopores. The research, now published in the prestigious Nature Biomedical Engineering, sheds light on the intricate interactions between proteins and DNA, particularly focusing on the CRISPR associated protein, Cas9, famous for its role in DNA editing. In this case, the catalytically inactive form of Cas9 is used, which allows the team to study how Cas9 binds to a specific DNA sequence without cutting it.

The central aim of the project is to decipher the sensitivity and specificity of these binding interactions between Cas9 and DNA molecules, which are essential for understanding genetic processes and developing advanced genetic engineering and diagnostic techniques. In simpler terms, the researchers aimed to comprehend how a specific protein, Cas9, interacts with DNA and how these interactions change if the DNA sequence changes.

“CRISPR-based technologies have immense potential for revolutionizing medicine and biotechnology, but to harness this potential, we need to deeply understand the intricacies of protein-DNA interactions. Our study develops new technologies that take us a step closer to achieving this understanding,” says Professor Weckman.

The team devised an ingenious approach, using minute structures made of DNA, referred to as “barcoded DNA nanostructures.” These nanostructures were meticulously designed to have predictable changes in size and shape when Cas9 proteins attach to specific segments of DNA. These changes in size and shape can then be observed as the barcoded DNA nanostructure passes through a small sensor called a nanopore, enabling the researchers to study the protein-DNA interactions in real-time.

“This new method we have developed, allows us to get a snapshot of each DNA nanostructure as it passes through the nanopore. By zooming in on individual protein-DNA interactions, we gain unprecedented insights into the binding strength, sequence specificity, and tolerance to DNA sequence variations including mutations,” explains Professor Weckman.

The results of the study have far-reaching implications. By analyzing how Cas9 proteins recognize and bind to different DNA sequences, the team hopes to unlock the key to enhancing the precision of DNA editing techniques and developing tailor-made diagnostic sensors to rapidly diagnose disease and detect mutations in DNA sequences. “One example of where this could be particularly important is in detecting mutations that make infections resistant to antibiotics so that we can match patients to the best treatments,” elaborates Professor Weckman. “Moreover, the innovative new tools for measuring protein-DNA interactions could be adapted to create ultrasensitive assays for a myriad of scientific and medical purposes.”

As Professor Weckman and her team continue to explore the implications of their findings, the scientific community and medical field eagerly anticipate the potential breakthroughs that may follow.