Speaker: Derek Stein
Department: Department of Physics, Brown University
Location: TU Delft, Bionanoscience, Kavli Institute of Nanoscience
Author: Mirte Golverdingen
Derek Stein gave his lecture to show what the new frontiers are in the Nanofluidics research. He started with the research to use nanoscience for protein sequencing. The general idea was to combine mass spectrometry and nanotubes. The techniques are combined so they overcome the limitations of each technique. The nanotubes were already known by the public. Mass spectrometry, however, was a more obscure technique for us. Derek therefore gave some background to give more understanding of this technique.
Mass Spectrometry is a powerful tool; it can tell the weight of every single molecule. Therefore, it is an extremely well established technique. The tool separates ions by their mass to charge ratio (m/z) in vacuum. Nowadays it has such a high resolution that it can resolve ions differing in mass by a single Dalton.
Another new development is electrospray ionization. This is a soft ionization technique that can be used for proteins without destroying the conformation of the molecule. It works as follows; you take a little tube with a diameter of several dozens of nanometers. Through the tube you pass a fluid with the analyzing molecules. Then you put a strong voltage between the liquid and a metal part at the end of the evaporation chamber in the vacuum chamber. Small nanodroplets that leave the tip of the tube come into an evaporation chamber where there is a gas that dries them out. This results in forces that become stronger than the forces in the liquid droplets, so the droplets blow up and this results in smaller droplets. At the end of this process the droplets are shrinking into ions.
Stein used both techniques to take a mass spectrum of adenosine monophosphate. The resolution of this technique is very precise and it showed a peak at 346,4 FW. For Single-Ion Detection Stein used an electron multiplier, the Channeltron. In a process called secondary emission, a single ion can induce the emission of roughly 1 to 3 electrons. In the Channeltron a chain reaction of emissions is created, so one ion sprays out a lot of electrons. In this way, the signal of one ion is amplified. Therefore, Stein could detect the single ions that resulted from the electrospray ionization.
However, if you use mass spectrometry for sequencing proteins, peptide fragmentation is necessary. The polypeptide will break on the backbone, between the O and the NH atoms. This is ideal because the peptides will break into amino acids, however, you do not know how the protein breaks apart. More tests are therefore needed to discover the sequence of the protein, and, how longer the protein, how more tests are needed.
Stein’s lab wants to combine mass spectrometry with a source of single ions detection. If you can start your measurement by passing a single polymer across a nanotube, then you will break the polymer apart and use the mass spectrometer to measure which amino acid passes. The order of arrival will tell the exact sequence of the protein, without using further tests. Because this will take place in a vacuum chamber, no random motion will occur. In his lab, they have built a system to start test this idea. (See figure 1)
Figure 1: Schematic representation of the setup Stein has proposed. The protein gains access to the vacuum chamber by passing a nanotube. At the end of the tube fragmentation takes place. The amino acids then are recognized by mass spectrometry. The sequence of the amino acids is determined by the sequence of the protein that entered the nanotube. Liszweski, K. (2016). The Next Next Thing in Sequencing. GEN. 36.Retrieved from: http://www.genengnews.com/gen-articles/the-next-next-thing-in sequencing/5653?kwrd=Oxford%20Nanopore
Stein showed me a very exciting application in biology of nanotechnology. He also showed how important knowledge of biology, chemistry and physics is in the nanobiology field. I am very excited about new ways of thinking about protein sequencing and adapting nanotechnology in the biochemistry field. This is also something I want to learn more about. By adapting the protein fragmentation into the mass spectroscopy, you can determine the sequence of the protein more easily. This idea uses nanotechnology in the form of nanotubes and mass spectroscopy. The idea of Stein is very exciting and could be a revolution in the protein sequence world.