<p> Electrospray ionization (ESI) is a premier method for volatilizing and ionizing biological analytes for mass spectrometry. In conventional mass spectrometry (MS), the spectrum of mass-to-charge ratio (<i>m/z</i>) for an ensemble of ions is measured. ESI produces a distribution of charges for each ionized species, and the mass of each species is determined by assigning a charge state to each peak in the <i>m/z</i> spectrum. These peaks are difficult to resolve for species above the 100-kDa range because of peak broadening and shifting due to salt adducts, incomplete desolvation, and intrinsic heterogeneity. Without resolved charge states, the mass cannot be determined. Charge detection mass spectrometry (CDMS) offers a solution to this problem. </p><p> In CDMS, both the <i>m/z</i> and the charge are measured simultaneously for individual ions. Multiplying those measurements for each ion yields the mass. Thus, there is no need for charge state resolution in an m/z spectrum. CDMS can therefore be used to measure the masses of extremely heavy and heterogeneous analytes far beyond the capabilities of conventional MS. This comes at the cost of efficiency, since single ions are measured serially, and resolution, since the charge measurement historically has been imprecise in CDMS. </p><p> Here we report a nearly perfect charge measurement in CDMS by analyzing each ion for 3 s in an electrostatic ion trap and implementing a novel analysis method. Then we discuss spontaneous mass and charge losses of trapped ions. Finally, we discuss multiple applications of CDMS to bacteriophage P22. P22 capsids assemble into <i>T = 7</i> ‘procapsids’ with the assistance of a distribution of scaffolding proteins; we report the typical width of that distribution. Next we report our observation of mass loss in P22 procapsids over the course of weeks due to precipitation of scaffolding proteins. Then we discuss how the charge on electrosprayed P22 capsids allows us to distinguish morphologies of P22 capsids. Finally, we report an accurate mass measurement of the infectious P22 phage, a >50 MDa particle containing nucleic acid and nine kinds of protein.</p>
| Identifer | oai:union.ndltd.org:PROQUEST/oai:pqdtoai.proquest.com:10129671 |
| Date | 26 August 2016 |
| Creators | Keifer, David Z. |
| Publisher | Indiana University |
| Source Sets | ProQuest.com |
| Language | English |
| Detected Language | English |
| Type | thesis |
Page generated in 0.0015 seconds