This summer in Flagstaff, Arizona at Northern Arizona University (NAU) I worked with Dr. Samrat Dutta in the Nishikawa Lab for biomechanics performing atomic force microscopy (AFM) on the N2A region of titin molecules at different pulling speeds in both the presence and absence of calcium. Titin is found in vertebrate muscles and is the largest known protein molecule (Nishikawa et. al, 2011). Currently, we know that titin functions in passive muscle movement. However, it may provide an important addition to our current understanding of both active and passive muscle function (Nishikawa et. al, 2011). Understanding titin isn’t just revolutionary for muscle theory, Nishikawa’s lab is applying this new information to improve prosthetics. AFM is a non-conventional type of microscope (shown below) that allows us to record the stiffness and stability of biomolecules such as titin by pulling on its spring-like domains. The titin is chemically attached to a surface and the AFM traverses that surface and records changes in its topography using a laser. This experiment allows us to predict the behavior of titin and its contribution to muscle force under different conditions.

Working with AFM has been a steep learning curve for me. AFM wasn’t a process that I was at all familiar with before this summer. With guidance from Dr. Dutta over the course of these 10 weeks I’ve learned about the chemistry, function, and potential of AFM. Unfortunately, we received low usable data yields and this may have been a result of the protein unfolding before the experiment began. As a result, there was a lot of tweaking of our methods to gather a larger set of more accurate data. The analysis of our data afforded me an opportunity to learn physics and chemistry beyond the scope of my university classes. However, we have not yet completed the analysis of our data. I look forward to seeing the outcome of our experiment and contribute to the ever-growing data on titin. Hopefully, my research will answer how much force titin can contribute and in comparison to previous works, does the N2A region of titin react differently than other regions.

What was it like working in the lab?

Dr. Kiisa Nishikawa’s lab group is filled with scientists doing various projects in all different disciplines of the muscle physiology field. Throughout my time at NAU, I had the opportunity to network and learn from all different kinds of people such as postdocs, graduate students, and full professors. They all guided me through my new environment at NAU and supplied me with both professional and scientific knowledge from their different disciplines. Working in a collaborative group meant having support during disappointing moments and always having someone to run ideas by. When the experiment wasn’t producing the amount of data we expected, a team of graduate students and postdocs helped my mentor and I brainstorm possible causes and solutions. This brainstorming session was how we determined that dialysis may be a useful alternative to our previous protein purification method. Working with a large team also means that you must share resources and space which, can make things more difficult. Overall, this summer was an invaluable experience in many aspects and it wouldn’t have been possible without the American Physiological Society (APS). I want to thank APS for allowing me and so many other undergraduates the opportunity to contribute to different fields of research.

References

1. Nishikawa, K. C., et al. Is Titin a ‘Winding Filament’? A New Twist on Muscle Contraction. Proceedings of the Royal Society B: Biological Sciences 279(1730), 981–90, 2011.

Blair Thompson is studying biology at Scripps College in Claremont, California. She is a 2017 fellowship recipient of Integrative Organismal Systems Physiology (IOSP) funded by the APS and a grant from the National Science Foundation Integrative Organismal Systems (IOS). She worked with Dr. Samrat Dutta and Dr. Jenna Monroy at Northern Arizona University this summer. After graduation, she plans to attend medical school and become a physician.