It’s good to be single in 2015

Single molecule work has really stood out to me at this meeting, as researchers are developing all sorts of interesting methods to study molecules both in vitro and in live cells. As someone who doesn’t do single molecule work, I found the Biophysics 101: Super-Resolution Microscopy session on Monday tremendously insightful. I think for the first time I truly understood what goes into improving resolution for optical microscopy techniques. In particular, I found Dr. Keith Lidke’s presentation very thorough and easy to understand, as he went through theory and then stepped through each and every acronym (fPALM, STORM, etc) and what was different about each one. I understand that the presentations will be posted online somewhere; does anyone have a link to this? Please share it in the comments or send it and I will be happy to post it here as well.

Having absorbed some basic background knowledge, I wandered over to the Single-Molecule Platform session on Tuesday morning. In particular, I was curious about a presentation on the development of single-molecule electron paramagnetic resonance (EPR) by Richelle M. Teeling-Smith from Ohio State. I use EPR spectroscopy in my own work to study protein-protein interactions and conformational changes. There are a variety of EPR methods, but the one I utilize employs the placement of thiol-reactive spin labels to introduce paramagnetic centers into proteins. The technique relies on the sensitivity of the label to its environment, and is a useful probe for protein dynamics on timescales that cannot be achieved by optical microscopy.

In the traditional continuous-wave EPR experiment, this label (or the corresponding protein it is attached to) needs to be enriched to a high enough concentration (typically 50-200 uM) to achieve a robust signal in the experiment. This is definitely a few more than a single molecule, and this the measurement is heavily averaged. So, I was quite curious about how this group had managed to develop a strong enough signal from one molecule to make a measurement.

Teeling-Smith and colleagues employed the covalent tethering of a nanodiamond label, which I’d never heard of previously but apparently is enriched with paramagnetic centers, to a single molecule of DNA. On its own, the nanodiamond achieves a robust signal that is sensitive to whether it is free in solution or attached to something bigger (much like a nitroxide spin label). It’s of course much larger than a nitroxide label (which is about the size of an amino acid side-chain), which I suspect makes it impractical for work such as my own. However, it can clearly be a sensitive reporter in larger systems, and may offer information at a level of detail that is not possible with single-molecule optical techniques. I found the work very creative and fascinating, and I will be curious to see how it progresses in the coming years.