First, some background
Scientists estimate that our cells contain more than a million structurally unique proteins, collectively known as the human proteome. Called proteoforms, each of these proteins has the potential to impact health and disease, so scientists are very interested in learning what they look like and being able to identify them. However, commercial mass spectrometers (MS) are limiting factors in this research: It takes hundreds of experiments to characterize only a fraction of the human proteome. And commercial MS can characterize proteins only up to about 30 daltons (or kDa, a unit of molecular mass), which only accounts for half the human proteome.
What did scientists discover?
Using a powerful, sensitive instrument designed and built for the MagLab, scientists identified far more proteoforms in human colorectal cancer cells per experiment (using a technique called liquid chromatography mass spectrometry/mass spectrometry) than previous studies.
Specifically, the largest intact protein identification study prior to this work focused on proteins with a molecular mass of less than 30 kDa, and required 423 separate experiments. By contrast, scientists at the MagLab identified a similar number of proteins (1,820) during just eight experiments — in less than 2 percent of the time required in the older set of experiments. This sets a new efficiency standard for intact protein analysis. In addition, the MagLab experiments identified ranged proteins in size from 5 to 55 kDa, which extends the mass range of intact protein analysis by a factor of two.
Why is this important?
Improving the throughput, sequence coverage, and molecular weight range of intact protein analysis will facilitate discovery of thousands of new proteoforms, many of which will have direct clinical relevance to human disease.
Who did the research?
L.C. Anderson1, C.J. DeHart1,2, N.K. Kaiser1, R.T. Fellers2, D.F. Smith1, J.B. Greer2, R.D. LeDuc2, G.T. Blakney1, P.M. Thomas2, N.L. Kelleher2,3, C.L. Hendrickson1,4
1National High Magnetic Field Laboratory; 2Proteomics Center of Excellence, Northwestern University; 3Department of Chemistry and Molecular Biosciences, Northwestern University; 4Department of Chemistry and Biochemistry, Florida State University
Why did they need the MagLab?
Successful high-throughput identification of intact proteins from complex biological samples requires instrumentation capable of high mass resolving power, mass accuracy, sensitivity, and spectral acquisition rate. The MagLab’s 21-tesla Fourier Transform–Ion Cyclotron Resonance (FT-ICR) mass spectrometer is state-of-the-art in each and every one of these critically important specifications. The instrument also allows scientists to get more information out of a small amount of sample, which is important when the material is limited and/or costly.
Details for scientists
- View or download the expert-level Science Highlight, Human Protein Identification by 21 T FT-ICR Mass Spectrometry
- Read the full-length publication, Identification and Characterization of Human Proteoforms by Top-Down LC-21 Tesla FT-ICR Mass Spectrometry , in Journal of Proteome Research
This research was funded by the following grants: G.S. Boebinger (NSF DMR-1157490); N.K. Kelleher (NIH P41GM108569)
For more information, contact Chris Hendrickson.