Submicrometer Emitter ESI Tips for Native Mass Spectrometry of Membrane Proteins in Ionic and Nonionic Detergents

  • Anna C. Susa
  • Jennifer L. Lippens
  • Zijie Xia
  • Joseph A. Loo
  • Iain D. G. Campuzano
  • Evan R. Williams
Application Note

Abstract

Native mass spectrometry (native-MS) of membrane proteins typically requires a detergent screening protocol, protein solubilization in the preferred detergent, followed by protein liberation from the micelle by collisional activation. Here, submicrometer nano-ESI emitter tips are used for native-MS of membrane proteins solubilized in both nonionic and ionic detergent solutions. With the submicrometer nano-ESI emitter tips, resolved charge-state distributions of membrane protein ions are obtained from a 150 mM NaCl, 25 mM Tris-HCl with 1.1% octyl glucoside solution. The relative abundances of NaCl and detergent cluster ions at high m /z are significantly reduced with the submicrometer emitters compared with larger nano-ESI emitters that are commonly used. This technique is beneficial for significantly decreasing the abundances (by two to three orders of magnitude compared with the larger tip size: 1.6 μm) of detergent cluster ions formed from aqueous ammonium acetate solutions containing detergents that can overlap with the membrane protein ion signal. Resolved charge-state distributions of membrane protein ions from aqueous ammonium acetate solutions containing ionic detergents were obtained with the submicrometer nano-ESI emitters; this is the first report of native-MS of membrane proteins solubilized by ionic detergents.

Graphical Abstract

Keywords

Membrane protein Native MS Detergents Desalting Ionic detergents Submicron Tips ESI Bacteriorhodopsin Aquaporin Z 

Notes

Acknowledgments

The authors are grateful for financial support from the National Institutes of Health (R01GM097357 to E.R.W. and R01GM103479 to J.A.L.) and for funds to acquire the Synapt G2Si (S10OD020062) used in these experiments. The authors thank Drs. Ryan Leib and Anthony Iavarone for helpful discussions, Drs. Nicholas Woodall and James U. Bowie (UCLA) for the generous gift of the bacteriorhodopsin T47A mutant, and Dr. Pascal Egea (UCLA) for preparation and gift of Aquaporin Z.

Supplementary material

13361_2017_1793_MOESM1_ESM.docx (18.7 mb)
ESM 1 (DOCX 19142 kb)

References

  1. 1.
    Hopkins, A.L., Groom, C.R.: The druggable genome. Nat. Rev. Drug Discov. 1, 727–730 (2002)CrossRefGoogle Scholar
  2. 2.
    Arinaminpathy, Y., Khurana, E., Engelman, D.M., Gerstein, M.B.: Computational analysis of membrane proteins: the largest class of drug targets. Drug Discov. Today. 14, 1130–1135 (2009)CrossRefGoogle Scholar
  3. 3.
    Hopper, J.T.S., Yu, Y.T.-C., Li, D., Raymond, A., Bostock, M., Liko, I., Mikhailov, V., Laganowsky, A., Benesch, J.L.P., Caffrey, M., Nietlispach, D., Robinson, C.V.: Detergent-free mass spectrometry of membrane protein complexes. Nat. Methods. 10, 1206–1208 (2013)CrossRefGoogle Scholar
  4. 4.
    Laganowsky, A., Reading, E., Hopper, J.T.S., Robinson, C.V.: Mass spectrometry of intact membrane protein complexes. Nat. Protoc. 8, 639–651 (2013)CrossRefGoogle Scholar
  5. 5.
    Harvey, S.R., Liu, Y., Liu, W., Wysocki, V.H., Laganowsky, A.: Surface induced dissociation as a tool to study membrane protein complexes. Chem. Commun. 53, 3106–3109 (2017)CrossRefGoogle Scholar
  6. 6.
    Cong, X., Liu, Y., Liu, W., Liang, X., Russell, D.H., Laganowsky, A.: Determining membrane protein-lipid binding thermodynamics using native mass spectrometry. J. Am. Chem. Soc. 138, 4346–4349 (2016)CrossRefGoogle Scholar
  7. 7.
    Campuzano, I.D.G., Li, H., Bagal, D., Lippens, J.L., Svitel, J., Kurzeja, R.J.M., Xu, H., Schnier, P.D., Loo, J.A.: Native MS analysis of bacteriorhodopsin and an empty nanodisc by orthogonal acceleration time-of-flight, Orbitrap, and ion cyclotron resonance. Anal. Chem. 88, 12427–12436 (2016)CrossRefGoogle Scholar
  8. 8.
    Kintzer, A.F., Sterling, H.J., Tang, I.I., Abdul-Gader, A., Miles, A.J., Wallace, B.A., Williams, E.R., Krantz, B.A.: Role of the protective antigen octamer in the molecular mechanism of anthrax lethal toxin stabilization in plasma. J. Mol. Biol. 399, 741–758 (2010)CrossRefGoogle Scholar
  9. 9.
    Reading, E., Liko, I., Allison, T.M., Benesch, J.L.P., Laganowsky, A., Robinson, C.V.: The role of the detergent micelle in preserving the structure of membrane proteins in the gas phase. Angew. Chem. Int. Ed. Eng. 54, 4577–4581 (2015)CrossRefGoogle Scholar
  10. 10.
    Barrera, N.P., Robinson, C.V.: Advances in the mass spectrometry of membrane proteins: from individual proteins to intact complexes. Annu. Rev. Biochem. 80, 247–271 (2011)CrossRefGoogle Scholar
  11. 11.
    Hernández, H., Robinson, C.V.: Determining the stoichiometry and interactions of macromolecular assemblies from mass spectrometry. Nat. Protoc. 2, 715–726 (2007)CrossRefGoogle Scholar
  12. 12.
    Iavarone, A.T., Udekwu, O.A., Williams, E.R.: Buffer loading for counteracting metal salt-induced signal suppression in electrospray ionization. Anal. Chem. 76, 3944–3950 (2004)CrossRefGoogle Scholar
  13. 13.
    Clarke, D.J., Campopiano, D.J.: Desalting large protein complexes during native electrospray mass spectrometry by addition of amino acids to the working solution. Analyst. 140, 2679–2686 (2015)CrossRefGoogle Scholar
  14. 14.
    Cassou, C.A., Williams, E.R.: Desalting protein ions in native mass spectrometry using supercharging reagents. Analyst. 139, 4810–4819 (2014)CrossRefGoogle Scholar
  15. 15.
    Flick, T.G., Cassou, C.A., Chang, T.M., Williams, E.R.: Solution additives that desalt protein ions in native mass spectrometry. Anal. Chem. 84, 7511–7517 (2012)CrossRefGoogle Scholar
  16. 16.
    DeMuth, J.C., McLuckey, S.A.: Electrospray droplet exposure to organic vapors: metal ion removal from proteins and protein complexes. Anal. Chem. 87, 1210–1218 (2015)CrossRefGoogle Scholar
  17. 17.
    Hu, J., Guan, Q.-Y., Wang, J., Jiang, X.-X., Wu, Z.-Q., Xia, X.-H., Xu, J.-J., Chen, H.-Y.: Effect of nanoemitters on suppressing the formation of metal adduct ions in electrospray ionization mass spectrometry. Anal. Chem. 89, 1838–1845 (2017)CrossRefGoogle Scholar
  18. 18.
    Schmidt, A., Karas, M., Dülcks, T.: Effect of different solution flow rates on analyte ion signals in nano-ESI MS, or: when does ESI turn into nano-ESI? J. Am. Soc. Mass Spectrom. 14, 492–500 (2003)CrossRefGoogle Scholar
  19. 19.
    Susa, A.C., Xia, Z., Williams, E.R.: Native mass spectrometry from common buffers with salts that mimic the extracellular environment. Angew. Chem. Int. Ed. Eng. 56, 7912–7915 (2017)CrossRefGoogle Scholar
  20. 20.
    Susa, A.C., Xia, Z., Williams, E.R.: Small emitter tips for native mass spectrometry of proteins and protein complexes from nonvolatile buffers that mimic the intracellular environment. Anal. Chem. 89, 3116–3122 (2017)CrossRefGoogle Scholar
  21. 21.
    Mortensen, D.N., Williams, E.R.: Electrothermal supercharging of proteins in native MS: effects of protein isoelectric point, buffer, and nanoESI-emitter tip size. Analyst. 141, 5598–5606 (2016)CrossRefGoogle Scholar
  22. 22.
    Seddon, A.M., Curnow, P., Booth, P.J.: Membrane proteins, lipids, and detergents: not just a soap opera. Biochim. Biophys. Acta. 1666, 105–117 (2004)CrossRefGoogle Scholar

Copyright information

© American Society for Mass Spectrometry 2017

Authors and Affiliations

  • Anna C. Susa
    • 1
  • Jennifer L. Lippens
    • 2
  • Zijie Xia
    • 1
  • Joseph A. Loo
    • 3
  • Iain D. G. Campuzano
    • 2
  • Evan R. Williams
    • 1
  1. 1.Department of ChemistryUniversity of CaliforniaBerkeleyUSA
  2. 2.Discovery Analytical Sciences, AmgenThousand OaksUSA
  3. 3.Department of Chemistry and Biochemistry, and Department of Biological ChemistryUniversity of California-Los AngelesLos AngelesUSA

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