Today was national doughnut day. Per the laws of office jobs, someone brought in two dozen doughnuts and they all disappeared within a matter of minutes. You could probably model the amount of doughnuts in the break room with an exponential decay graph. I barely got half of a doughnut, but Dr. Masoudi and I had to carry on with our forlorn bellies. Our empty stomachs pretty much paralleled our experiment (which was fruitless). The electrophoresis analysis that we ran yesterday told us that we needed to start the entire protein-harvesting process all over again. Dr. Masoudi got out a new batch of E. Coli cells, introducing a bacterium to the flasks and keeping the containers in incubators. We also set up a protein-identifying process called a “Western” that creates a protein imprint on a membrane using an electric current – I have yet to see the results of this experiment. We ran a second electrophoresis experiment today as well, and we received a new and unexpected result. The gel that holds our data works by showing us the distribution of peptide chains based on molar mass. Yesterday, we assumed that the thickest, boldest line indicated our target protein, Nb6B9, even though its molar mass was a bit off. The new setup tells us something completely different; that thick, bold line was actually a contaminant that attached itself to our nickel resin during filtration and was never successfully washed out. In our new analysis, a faint second line pops up below a bolder line and is between the molar masses of 5,000 and 17,000 kiloDaltons (the first and second notches) – this line better matches Nb6B9 which is approximately 12,000 kDa. The new trial has pretty much dispelled our previous preconception that the bold line was our target protein, supporting the idea that our filtration methods resulted in an unsatisfactory percent yield.
Besides this bad news, the rest of the day went pretty well. I’m getting to know everyone in the lab a bit better, and I’ve even observed some other chemists setting up X-ray crystallography. In this practice, proteins are crystallized for stability and X-rays are shot at the configurations to create images of the crystallized structures. Hopefully Dr. Masoudi and I will soon get the chance to crystallize and examine our stingy Nb6B9 protein when its attached to a cell receptor (if the protein ever decides to cooperate). Fingers crossed!
Here’s an end-of-the-week haiku:
Insect receptors
Patiently wait for stingy
Nb6B9