Summer Summary

While the summer research has come to an end, my passion and eagerness for more research has only been enhanced.

My original goal of my summer research was to write a paper about my AzoBen unnatural amino acid (UAA), and I can happily say that it is 90% complete! Before the summer began, I had already synthesized the UAA, but I hadn’t characterized its affects and utility. While working with AzoBen, most of my time was spent on a fluorimeter (named Florence), analyzing emission spectra. I inserted the UAA into the fluorescent protein GFP at various residues. The residues I used were 3, 133, 66, and 151, where I would test fluorescence of GFP with each of residues, individually. After testing preliminary fluorescence, I would irradiate the protein in order to cause the photoisomerization of AzoBen. Also, I grew several culture plates of bacteria with GFP and my UAA. Half of the plates were irradiated and compared to the other half, as well as a control plate. From my results, I was able to conclude that AzoBen is able to alter the fluorescence of GFP, especially at GFP 66 (which makes sense because the 66th residue is the fluorophore. The bacteria plates were especially exciting, as there was a clear change in fluorescence on of the sides of the plates. Florence was trickier, as I had to measure emission spectrum at different wavelengths. I ended up doing hundreds of scans, for various the various residues, and I did see a clear change in fluorescence for all of the residues. While the idea that AzoBen is able to alter the fluorescence of GFP is exciting, the results show a much deeper conclusion. AzoBen is able to be inserted into an active biological system, and be manipulated externally to alter the functionality of a protein. ¬†Proteins are the workhorses of the cell, and being able to change their functionality or deactivate them means enormous opportunities. For example, say an ion channel was misregulating sodium flow outside of the cell. If AzoBen was inserted into that ion channel, it could be photoisomerized, and deactivate the ion channel. The medical implications of this ability are staggering, as many diseases are caused not only by faulty ion channels, but malfunctioning proteins in general.

While my research with AzoBen is most likely finished (due to technological restrictions), I have gained an immense understanding  and knowledge of the biochemical world.

While my summer’s main focus was on AzoBen, I was also given the opportunity to research another UAA: ProTyro. This UAA has a free alkyne group that allows it to perform dynamic reactions. One such reaction is the “click” reaction, in which two alkynes are reacted together and form a bond. Click reactions are also research in Dr. Young’s lab, and it provides the ability for my follow labmates and I to work together. Before I can actually perform the click reaction, I need to synthesize the molecule. If you read my previous posts, the synthesis has been difficult. Throughout the summer, I attempted the synthesis four separate times (my fellow organic chemists would laugh at this), and on the last attempt, I got product that was confirmed to be ProTyro. Throughout each of the six steps to from ProTyro, I really focused on perfecting technique to allow for the highest yield as I could muster. While I was able to perform the first four steps rather successfully, the last two steps (which involved microwaves) were difficult. On my last synthesis I realized I needed to use different microwave settings, and this allowed the reaction to be successful.

My goal throughout Junior year is to be able to perform click reactions on ProTyro, both isolated, and in a protein.

Summer Research 2014 has been a great blend of friendships, progress, and fun. I might be difficult, but I hope that next year’s session will compare to this one!

-Marshall