DNA origami tiles as well as extended DNA grids provide an excellent platform to orient proteins with high precision. We are using both templates to study the influence of protein orientation on their structure and function.
The advent of the DNA origami technology has certainly transformed DNA bio-nanotechnology. Precision made DNA nanostructures have become available, where 2D and 3D structures can be designed and created almost at will. We are using both discrete DNA origami tiles and extended DNA arrays for precision templating of proteins. Together with Dafydd Jones in Cardiff we make use of site-specifically modified proteins with azides, which are coupled to alkyne-DNA using copper-free click chemistry (ACS Nano 2017). This allows to make proteins where exactly one DNA strand is attached to the protein at very precise locations. This is in stark contrast to the general procedure of cross-linking to surface lysins. With this we have controlled the orientation and activity of proteins on DNA origami surfaces and noticed a substantial increase in activity compared to the modified protein in solution.
These results have opened very important questions in protein analysis. Clearly, the activity is governed by the orientation on the surface and the attachment point of the DNA, whereas the relative distance to the origami surface did not seem to make much of a difference. However, this system can now be used to further structurally characterise proteins bound to surfaces, and here we are using SRCD to obtain information on specific spectroscopic characteristic of oriented proteins. This is research that is in part sponsored by DLS B23; first results are due to come out shortly.
Funding Provider:
Diamond Light Source
https://www.diamond.ac.uk/Home.html
BBSRC
