Analysis of Native-Like Proteins and Protein Complexes Using Cation to Anion Proton Transfer Reactions (CAPTR).
Mass spectra of native-like protein complexes often exhibit narrow charge-state distributions, broad peaks, and contributions from multiple, coexisting species. These factors can make it challenging to interpret those spectra, particularly for mixtures with significant heterogeneity. Here we demonstrate the use of ion/ion proton transfer reactions to reduce the charge states of m/z-selected, native-like ions of proteins and protein complexes, a technique that we refer to as cation to anion proton transfer reactions (CAPTR). We then demonstrate that CAPTR can increase the accuracy of charge state assignments and the resolution of interfering species in native mass spectrometry. The CAPTR product ion spectra for pyruvate kinase exhibit ~30 peaks and enable unambiguous determination of the charge state of each peak, whereas the corresponding precursor spectra exhibit ~6 peaks and the assigned charge states have an uncertainty of ±3%. 15+ bovine serum albumin and 21+ yeast enolase dimer both appear near m/z 4450 and are completely unresolved in a mixture. After a single CAPTR event, the resulting product ions are baseline resolved. The separation of the product ions increases dramatically after each subsequent CAPTR event; 12 events resulted in a 3000-fold improvement in separation relative to the precursor ions. Finally, we introduce a framework for interpreting and predicting the figures of merit for CAPTR experiments. More generally, these results suggest that CAPTR strongly complements other mass spectrometry tools for analyzing proteins and protein complexes, particularly those in mixtures. Graphical Abstract ᅟ.
Department of Chemistry, University of Washington, Seattle, WA, 98195-1700, USA.
Kenneth J
KJ
Laszlo
Department of Chemistry, University of Washington, Seattle, WA, 98195-1700, USA. mattbush@uw.edu.
Matthew F
MF
Bush
D000818:Animals; D000838:Anions / Q000737:chemistry; D002412:Cations / Q000737:chemistry; D002417:Cattle; D004867:Equipment Design; D013058:Mass Spectrometry / Q000295:instrumentation / Q000379:methods*; D010751:Phosphopyruvate Hydratase / Q000032:analysis; D011506:Proteins / Q000032:analysis*; D011522:Protons; D012710:Serum Albumin, Bovine / Q000032:analysis; D015003:Yeasts / Q000201:enzymology
D000838:Anions; D002412:Cations; D011506:Proteins; D011522:Protons; D012710:Serum Albumin, Bovine; D010751:Phosphopyruvate Hydratase
Ion chemistry; Ion/ion chemistry; Native mass spectrometry; Protein complexes; Proton transfer reactions