Native Mass Spectrometry as a Tool in Structural Biology

Kristina Lorenzen1, Esther van Duijn1

1 Utrecht University, Utrecht, The Netherlands
Publication Name:  Current Protocols in Protein Science
Unit Number:  Unit 17.12
DOI:  10.1002/0471140864.ps1712s62
Online Posting Date:  November, 2010
GO TO THE FULL TEXT: PDF or HTML at Wiley Online Library

Abstract

Native mass spectrometry (native MS) gives information about the composition, topological arrangements, dynamics, and structural properties of protein complexes. The mass range is principally unlimited and highly dynamic, allowing the detection of small subunits and large complexes within the same measurement. The amount of protein needed for an analysis is, compared to most other structural biology methods, very low. This unit provides an introduction to native MS. It starts with an explanation of the basic method and details on how to measure intact proteins and protein complexes, and continues with the study of dynamics and complex stability in the gas phase. The final section discusses the most recent extension to the native MS field, ion mobility, which allows the direct assessment of the structural properties of the complexes of interest. Curr. Protoc. Protein Sci. 62:17.12.1‐17.12.17. © 2010 by John Wiley & Sons, Inc.

Keywords: native mass spectrometry; protein complex topology; structural biology; ion mobility–mass spectrometry; tandem mass spectrometry; sample preparation and conditions; protein ion charge states

     
 
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library

Table of Contents

  • Introduction
  • Generation of Ions
  • Mass Analyzers
  • Sample Requirements
  • Protein Ligand Binding
  • Specific or Non‐Specific Protein Complexes
  • Tandem Mass Spectrometry Experiments
  • Tandem MS to Study Membrane Proteins
  • In‐Solution Dissociation Experiments
  • Influencing Protein Charge States
  • Ion Mobility–Mass Spectrometry
  • Processing Structural Information
  • Summary
  • Acknowledgements
  • Literature Cited
  • Figures
  • Tables
     
 
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library

Materials

GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library

Figures

Videos

Literature Cited

Literature Cited
   Abzalimov, R., Wehri, S.C., Lorimer, G.H., and Kaltashov, I.A. 2009. Enhanced stoichiometry measurements of heterogeneous sub‐MDa protein assemblies by tandem MS: Elucidation of subunit exchange mechanism in GroEL In 57th ASMS 2009. Philadelphia.
   Andersen, J.S., Svensson, B., and Roepstorff, P. 1996. Electrospray ionization and matrix assisted laser desorption/ionization mass spectrometry: Powerful analytical tools in recombinant protein chemistry. Nat. Biotechnol. 14:449‐457.
   Bagal, D., Kitova, E.N., Liu, L., El‐Hawiet, A., Schnier, P.D., and Klassen, J.S. 2009. Gas Phase stabilization of noncovalent protein complexes formed by electrospray ionization. Anal. Chem. 81:7801‐7806.
   Barrera, N.P., Di Bartolo, N., Booth, P.J., and Robinson, C.V. 2008. Micelles protect membrane complexes from solution to vacuum. Science 321:243‐246.
   Barrera, N.P., Isaacson, S.C., Zhou, M., Bavro, V.N., Welch, A., Schaedler, T.A., Seeger, M.A., Miguel, R.N., Korkhov, V.M., van Veen, H.W., Venter, H., Walmsley, A.R., Tate, C.G., and Robinson, C.V. 2009a. Mass spectrometry of membrane transporters reveals subunit stoichiometry and interactions. Nat. Methods 6:585‐587.
   Barrera, N.P., Isaacson, S.C., Zhou, M., Bavro, V.N., Welch, A., Schaedler, T.A., Seeger, M.A., Miguel, R.N., Korkhov, V.M., van Veen, H.W., Venter, H., Walmsley, A.R., Tate, C.G., and Robinson, C.V. 2009b. Mass spectrometry of membrane transporters reveals subunit stoichiometry and interactions. Nat. Methods 6:585‐587.
   Benesch, J.L., Ruotolo, B.T., Simmons, D.A., and Robinson, C.V. 2007. Protein Complexes in the gas phase: Technology for structural genomics and proteomics. Chem. Rev. 107:3544‐3567.
   Breuker, K. and McLafferty, F.W. 2008. Stepwise evolution of protein native structure with electrospray into the gas phase, 10−12 to 102 s. Proc. Natl. Acad. Sci. U.S.A. 105:18145‐18152.
   Chaurand, P., Luetzenkirchen, F., and Spengler, B. 1999. Peptide and protein identification by matrix‐assisted laser desorption ionization (MALDI) and MALDI‐post‐source decay time‐of‐flight mass spectrometry. J. Am. Soc. Mass Spectrom. 10:91‐103.
   Chernushevich, I.V. and Thomson, B.A. 2004. Collisional cooling of large ions in electrospray mass spectrometry. Anal. Chem. 76:1754‐1760.
   de la Mora, J.F. 2000. Electrospray ionization of large multiply charged species proceeds via Dole's charged residue mechanism. Analytica Chimica Acta 406:93‐104.
   Elizabeth, D.‐N., Monique, B.K., Rolf, B., Arjan, B., Albert, J.H., Robert, H.v.d.H. and Ramón, D.‐O. 2009. A mutagenic analysis of the RNase mechanism of the bacterial Kid toxin by mass spectrometry. FEBS J. 276:4973‐4986.
   Ganem, B. and Henion, J.D. 1991a. Observation of noncovalent enzyme‐substrate and enzyme‐product complexes by ion‐spray mass spectrometry. J. Am. Chem. Soc. 113:7818‐7819.
   Ganem, B., Li, Y.T., and Henion, J.D. 1991b. Detection of noncovalent receptor ligand complexes by mass‐spectrometry. J. Am. Chem. Soc. 113:6294‐6296.
   Gavin, A.C., Aloy, P., Grandi, P., Krause, R., Boesche, M., Marzioch, M., Rau, C., Jensen, L.J., Bastuck, S., Dümpelfeld, B., Edelmann, A., Heurtier, M.A., Hoffman, V., Hoefert, C., Klein, K., Hudak, M., Michon, A.M., Schelder, M., Schirle, M., Remor, M., Rudi, T., Hooper, S., Bauer, A., Bouwmeester, T., Casari, G., Drewes, G., Neubauer, G., Rick, J.M., Kuster, B., Bork, P., Russell, R.B., and Superti‐Furga, G. 2006. Proteome survey reveals modularity of the yeast cell machinery. Nature 440:631‐636.
   Guilhaus, M., Selby, D., and Mlynski, V. 2000. Orthogonal acceleration time‐of‐flight mass spectrometry. Mass Spectrom. Rev. 19:65‐107.
   Heck, A.J. 2008. Native mass spectrometry: A bridge between interactomics and structural biology. Nat. Methods 5:927‐933.
   Hernandez, H., Dziembowski, A., Taverner, T., Seraphin, B., and Robinson, C.V. 2006. Subunit architecture of multimeric complexes isolated directly from cells. EMBO Rep. 7:605‐610.
   Ilag, L.L., Ubarretxena‐Belandia, I., Tate, C.G., and Robinson, C.V. 2004. Drug binding revealed by tandem mass spectrometry of a protein‐micelle complex. J. Am. Chem. Soc. 126:14362‐14363.
   Jecklin, M.C., Schauer, S., Dumelin, C.E., and Zenobi, R. 2009. Label‐free determination of protein‐ligand binding constants using mass spectrometry and validation using surface plasmon resonance and isothermal titration calorimetry. J. Mol. Recognit. 22:319‐329.
   Jorgensen, T.J.D., Roepstorff, P., and Heck, A.J.R. 1998. Direct determination of solution binding constants for noncovalent complexes between bacterial cell wall peptide analogues and vancomycin group antibiotics by electrospray ionization mass spectrometry. Anal. Chem. 70:4427‐4432.
   Kaltashov, I.A. and Mohimen, A. 2005. Estimates of protein surface areas in solution by electrospray ionization mass spectrometry. Anal. Chem. 77:5370‐5379.
   Kebarle, P. 2000. A brief overview of the present status of the mechanisms involved in electrospray mass spectrometry. J. Mass Spectrom. 35:804‐817.
   Kebarle, P. and Peschke, M. 2000. On the mechanisms by which the charged droplets produced by electrospray lead to gas phase ions. Anal. Chim. Acta 406:11‐35.
   Kenny, D.J., Brown, J.M., Palmer, M.E., Snel, M.F., and Bateman, R.H. 2006. A parallel approach to post source decay MALDI‐TOF analysis. J. Am. Soc. Mass Spectrom. 17:60‐66.
   Konermann, L. and Douglas, D.J. 1998. Unfolding of proteins monitored by electrospray ionization mass spectrometry: A comparison of positive and negative ion modes. J. Am. Soc. Mass Spectrom. 9:1248‐1254.
   Lane, L.A., Ruotolo, B.T., Robinson, C.V., Favrin, G., and Benesch, J.L.P. 2009. A Monte Carlo approach for assessing the specificity of protein oligomers observed in nano‐electrospray mass spectra. Int. J. Mass Spectrom. 283:169‐177.
   Leary, J.A., Schenauer, M.R., Stefanescu, R., Andaya, A., Ruotolo, B.T., Robinson, C.V., Thalassinos, K., Scrivens, J.H., Sokabe, M., and Hershey, J.W. 2009. Methodology for measuring conformation of solvent‐disrupted protein subunits using T‐WAVE ion mobility MS: an investigation into eukaryotic initiation factors. J. Am. Soc. Mass Spectrom. 20:1699‐1706.
   Lomeli, S.H., Yin, S., Ogorzalek Loo, R.R., and Loo, J.A. 2009. Increasing charge while preserving noncovalent protein complexes for ESI‐MS. J. Am. Soc. Mass Spectrom. 20:593‐596.
   Loo, J.A. 1997. Studying noncovalent protein complexes by electrospray ionization mass spectrometry. Mass Spectrom. Rev. 16:1‐23.
   Loo, J.A., Berhane, B., Kaddis, C.S., Wooding, K.M., Xie, Y., Kaufman, S.L., and Chernushevich, I.V. 2005. Electrospray ionization mass spectrometry and ion mobility analysis of the 20S proteasome complex. J. Am. Soc. Mass Spectrom. 16:998‐1008.
   Lorenzen, K., Vannini, A., Cramer, P., and Heck, A.J.R. 2007a. Structural biology of RNA polymerase III: Mass spectrometry elucidates subcomplex architecture. Structure 15:1237‐1245.
   Lorenzen, K., Versluis, C., van Duijn, E., van den Heuvel, R.H.H., and Heck, A.J.R. 2007b. Optimizing macromolecular tandem mass spectrometry of large non‐covalent complexes using heavy collision gases. Int. J. Mass Spectrom. 268:198‐206.
   Lorenzen, K., Olia, A.S., Uetrecht, C., Cingolani, G., and Heck, A.J. 2008. Determination of stoichiometry and conformational changes in the first step of the P22 tail assembly. J. Mol. Biol. 379:385‐396.
   Mamyrin, B.A. 2001. Time‐of‐flight mass spectrometry (concepts, achievements, and prospects). Int. J. Mass Spectrom. 206:251‐266.
   McKay, A.R., Ruotolo, B.T., Ilag, L.L., and Robinson, C.V. 2006. Mass measurements of increased accuracy resolve heterogeneous populations of intact ribosomes. J. Am. Chem. Soc. 128:11433‐11442.
   Ruotolo, B.T., Giles, K., Campuzano, I., Sandercock, A.M., Bateman, R.H., and Robinson, C.V. 2005. Evidence for macromolecular protein rings in the absence of bulk water. Science 310:1658‐1661.
   Ruotolo, B.T., Benesch, J.L., Sandercock, A.M., Hyung, S.J., and Robinson, C.V. 2008. Ion mobility‐mass spectrometry analysis of large protein complexes. Nat. Protoc. 3:1139‐1152.
   Smith, D.P., Knapman, T.W., Campuzano, I., Malham, R.W., Berryman, J.T., Radford, S.E., and Ashcroft, A.E. 2009. Deciphering drift time measurements from travelling wave ion mobility spectrometry‐mass spectrometry studies. Eur. J. Mass Spectrom. 15:113‐130.
   Sobott, F., Hernandez, H., McCammon, M.G., Tito, M.A., and Robinson, C.V. 2002. A tandem mass spectrometer for improved transmission and analysis of large macromolecular assemblies. Anal. Chem. 74:1402‐1407.
   Sun, J., Kitova, E.N., and Klassen, J.S. 2007. Method for stabilizing protein‐ligand complexes in nanoelectrospray ionization mass spectrometry. Anal. Chem. 79:416‐425.
   Sun, N., Sun, J., Kitova, E., and Klassen, J.S. 2009. Identifying nonspecific ligand binding in electrospray ionization mass spectrometry using the reporter molecule method. J. Am. Soc. Mass Spectrom. 20:1242‐1250.
   Synowsky, S.A., van den Heuvel, R.H., Mohammed, S., Pijnappel, P.W., and Heck, A.J. 2006. Probing genuine strong interactions and post‐translational modifications in the heterogeneous yeast exosome protein complex. Mol. Cell. Proteomics 5:1581‐1592.
   Tahallah, N., Pinkse, M., Maier, C.S., and Heck, A.J. 2001. The effect of the source pressure on the abundance of ions of noncovalent protein assemblies in an electrospray ionization orthogonal time‐of‐flight instrument. Rapid Commun. Mass Spectrom. 15:596‐601.
   Taverner, T., Hernandez, H., Sharon, M., Ruotolo, B.T., Matak‐Vinkovic, D., Devos, D., Russell, R.B., and Robinson, C.V. 2008. Subunit architecture of intact protein complexes from mass spectrometry and homology modeling. Acc. Chem. Res. 41:617‐627.
   Uetrecht, C., Versluis, C., Watts, N.R., Roos, W.H., Wuite, G.J., Wingfield, P.T., Steven, A.C., and Heck, A.J. 2008a. High‐resolution mass spectrometry of viral assemblies: Molecular composition and stability of dimorphic hepatitis B virus capsids. Proc. Natl. Acad. Sci. U.S.A. 105:9216‐9220.
   Uetrecht, C., Versluis, C., Watts, N.R., Wingfield, P.T., Steven, A.C., and Heck, A.J. 2008b. Stability and shape of hepatitis B virus capsids in vacuo. Angew Chem. Int. Ed. Engl. 47:6247‐6251.
   Uetrecht, C., Rose, R.J., Van Duijn, E., Lorenzen, K., and Heck, A.J. 2010. Ion mobility mass spectrometry of proteins and protein assemblies. Chemical Society Reviews. 39:1633‐1655.
   van den Heuvel, R.H.H., van Duijn, E., Mazon, H., Synowsky, S.A., Lorenzen, K., Versluis, C., Brouns, S.J., Langridge, D., van der Oost, J., Hoyes, J., and Heck, A.J. 2006. Improving the performance of a quadrupole time‐of‐flight instrument for macromolecular mass spectrometry. Anal. Chem. 78:7473‐7483.
   Van Duijn, E., Barendregt, A., Synowsky, S.A., Versluis, C., and Heck, A.J. 2009. Chaperonin complexes monitored by ion mobility mass spectrometry. J. Am. Chem. Soc. 131:1452‐1459.
   Videler, H., Ilag, L.L., McKay, A.R.C., Hanson, C.L., and Robinson, C.V. 2005. Mass spectrometry of intact ribosomes. FEBS Lett. 579:943‐947.
   Wisztorsk, M., Croix, D., Macagno, E., Fournier, I., and Salzet, M. 2008. Molecular MALDI imaging: An emerging technology for neuroscience studies. Dev. Neurobiol. 68:845‐858.
GO TO THE FULL PROTOCOL:
PDF or HTML at Wiley Online Library