Clare Stevenson runs the Biophysical Analysis facility at the John Innes Centre.
Recently, she explained why she is passionate about increasing the use of these Biophysical techniques and making the science more accessible to a wider range of users.
“The Biophysical Analysis (BA) facility has state of the art instrumentation for looking at structures of molecules and enables scientists to observe and measure the strength of the interactions between biomolecules.
Biomolecular interactions are central to biochemistry and an understanding of these interactions can aid our knowledge across a broad range of science, so the biomolecules we look at may be proteins, DNA, RNA, small molecules or drugs.
We recently added a third technique to our biophysical service; a Circular Dichroism Spectroscopy (CD), maintained and run by Julia Mundy.
The CD spectrophotometer uses a Xenon light source to collect CD data on biological macromolecules in solution at near and far UV wavelengths. This technique is commonly used to look at the folding of proteins, provide information on the secondary structure of proteins and can detect changes in structure during protein-protein or protein-ligand binding events. Julia runs samples for scientists as a service or will train users how to use the equipment themselves.
In addition to the CD, the BA facility has two complementary techniques for studying biomolecular interactions; Surface Plasmon Resonance (SPR) and Isothermal Titration Calorimetry (ITC).
In Surface Plasmon Resonance (SPR), one of the biomolecules you are investigating is attached to a surface and the other biomolecule is flowed over the surface. If there is an interaction a response is observed. That interaction can then be observed, on the computer, in real time and you see the biomolecules binding and unbinding. Once you know that an interaction occurs you can do some further measurements to determine how strong or weak the interaction is.
The SPR is highly automated and once an experiment is optimised it can even be left testing multiple interactions while the scientist is at home sleeping. Recently I have been working with Dr Tung Le, looking at protein-DNA interactions. Once the experiment was planned, it can be run quickly and within 20 minutes we can see if binding is happening. Tung’s group can easily screen many different protein samples and DNA sequences quickly in an automated manner getting the results the next day.
We’ve also recently been successful in obtaining some funding which will allow us to purchase a new instrument that will enhance our SPR capability even further.
Isothermal Titration Calorimetry (ITC) is a complimentary technique to SPR but rather than requiring one biomolecule to be immobilised, it has one in a cell and one in a syringe. The molecule in the syringe is injected into the one in the cell and if binding occurs the heat given out (exothermic) or taken in (endothermic) can be measured.
Changes in heat occur when molecules bind, and the ITC can measure these tiny changes in heat with high sensitivity. This information can then tell us whether the biomolecules interact and, if they do, how strong the interactions is. Plus, knowing the heat changes in the interaction can also give us a clue to how the molecules are binding.
Both techniques are complimentary but have different advantages and disadvantages. As a facility we speak to the scientists and recommend the best technique for the desired experiment and spend time training, troubleshooting projects and analysing results.
We also run regular training courses where people interested in all three techniques can come and learn what is involved and what they need to understand before putting on their lab coat.
Our BA facility is available for anyone to book and use.
Predominantly we work with John Innes Centre scientists, but we have also worked with The Sainsbury Laboratory, University of East Anglia and external companies like Leaf Expression Systems. If you want to measure any interactions, we can help, wherever you are.
Our facility can run 24-hours-a-day, 365-days-a-year, thanks to being able to automate much of the process, which in turn means we have the instrument capacity to take on more than we are currently doing.
I enjoy my job and my career has had lots of highlights.
I am particularly proud of a method that I developed to study protein DNA interactions by SPR. This is our Reusable DNA Capture Technique (ReDCaT) and enables DNA to be attached to a surface and easily removed. This means that the SPR instrument can be used to measure the interaction of multiple proteins with many DNA sequences in a high throughput and automated manner.
I was always good at science and maths at school, but I think my parents expected me to become a doctor, rather than a scientist. I come from a medical family, full of doctors and nurses, so it was sort of assumed I would follow the family trade.
However, I was bit of a rebellious teenager, so I found myself applying to do Biochemistry at Liverpool Polytechnic, now Liverpool John Moores University, and realised that biochemistry was a fascinating subject and I wanted to learn more.
My degree included a year in industry and I really enjoyed working in the lab. After graduation I found a job with a rival company and I had a lot of fun and learnt a lot working in the area of drug metabolism.
After a new challenge in 1996 I applied for a job as a Research Technician for Professor David Lawson. His passion for solving the 3-dimensional structures of proteins using crystals was inspiring and it encouraged me to take a step sideways. I must admit, I am hugely grateful to David, because he took a bit of a chance on me, as when I first came, I had only done a little bit of protein work and knew very little about protein crystallography. However, I learnt quickly and had excellent technical skills so was able to become competent at growing protein crystals and solving structures.
I realised that in academia having a PhD is important (although believe it is your competency and skills rather than the qualification that is what is really the important thing…) but I was lucky to have the support from the John Innes Centre to complete my PhD in 2007. It took over seven years studying part-time, while I was working full time and I also had a baby.
I solved several structures as part of my PhD but also had to study a protein-DNA interactions and had to learn how to use SPR. The more I did it the more I enjoyed it and over time I became the most experienced person at the John Innes Centre for using this technique and ended up training and advising others.
Over time SPR became the first technique in the facility and then it expanded to include ITC and CD. I now manage the facility but also continue working for David on the Protein Crystallography Facility.
There was, and I think remains, a feeling that biophysics is quite pedantic and difficult, but the machines are actually relatively easy to use with the right training and support. I love working with students and post docs showing them the techniques and designing their experiments with them.
Running a facility means every day is different and sometime hectic but feel lucky to work with so many great scientists and make their experiments happen.”