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Title: Native top-down mass spectrometry for the structural characterization of human hemoglobin

Abstract

Native mass spectrometry (MS) has become an invaluable tool for the characterization of proteins and non-covalent protein complexes under near physiological solution conditions. Here we report the structural characterization of human hemoglobin (Hb), a 64 kDa oxygen-transporting protein complex, by high resolution native top-down mass spectrometry using electrospray ionization (ESI) and a 15-Tesla Fourier transform ion cyclotron resonance (FTICR) mass spectrometer. Native MS preserves the non-covalent interactions between the globin subunits, and electron capture dissociation (ECD) produces fragments directly from the intact Hb complex without dissociating the subunits. Using activated ion ECD, we observe the gradual unfolding process of the Hb complex in the gas phase. Without protein ion activation, the native Hb shows very limited ECD fragmentation from the N-termini, suggesting a tightly packed structure of the native complex and therefore low fragmentation efficiency. Precursor ion activation allows steady increase of N-terminal fragment ions, while the C-terminal fragments remain limited (38 c ions and 4 z ions on the α chain; 36 c ions and 2 z ions on the β chain). This ECD fragmentation pattern suggests that upon activation, the Hb complex starts to unfold from the N-termini of both subunits, whereas the C-terminal regions and therefore themore » potential regions involved in the subunit binding interactions remain intact. ECD-MS of the Hb dimer show similar fragmentation patterns as the Hb tetramer, providing further evidence for the hypothesized unfolding process of the Hb complex in the gas phase. Native top-down ECD-MS allows efficient probing of the Hb complex structure and the subunit binding interactions in the gas phase. Finally, it may provide a fast and effective means to probe the structure of novel protein complexes that are intractable to traditional structural characterization tools.« less

Authors:
 [1];  [1];  [1];  [1]
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  1. Univ. of California, Los Angeles, CA (United States)
Publication Date:
Research Org.:
Univ. of California, Los Angeles, CA (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1345518
Grant/Contract Number:  
FC03-02ER63421
Resource Type:
Accepted Manuscript
Journal Name:
European Journal of Mass Spectrometry
Additional Journal Information:
Journal Volume: 21; Journal Issue: 3; Journal ID: ISSN 1469-0667
Publisher:
IM Publications
Country of Publication:
United States
Language:
English
Subject:
47 OTHER INSTRUMENTATION; native mass spectrometry; electron capture dissociation; protein complex; gas phase structure; binding interface; hemoglobin

Citation Formats

Zhang, Jiang, Malmirchegini, G. Reza, Clubb, Robert T., and Loo, Joseph A. Native top-down mass spectrometry for the structural characterization of human hemoglobin. United States: N. p., 2015. Web. doi:10.1255/ejms.1340.
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Zhang, Jiang, Malmirchegini, G. Reza, Clubb, Robert T., & Loo, Joseph A. Native top-down mass spectrometry for the structural characterization of human hemoglobin. United States. https://doi.org/10.1255/ejms.1340
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Zhang, Jiang, Malmirchegini, G. Reza, Clubb, Robert T., and Loo, Joseph A. Tue . "Native top-down mass spectrometry for the structural characterization of human hemoglobin". United States. https://doi.org/10.1255/ejms.1340. https://www.osti.gov/servlets/purl/1345518.
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@article{osti_1345518,
title = {Native top-down mass spectrometry for the structural characterization of human hemoglobin},
author = {Zhang, Jiang and Malmirchegini, G. Reza and Clubb, Robert T. and Loo, Joseph A.},
abstractNote = {Native mass spectrometry (MS) has become an invaluable tool for the characterization of proteins and non-covalent protein complexes under near physiological solution conditions. Here we report the structural characterization of human hemoglobin (Hb), a 64 kDa oxygen-transporting protein complex, by high resolution native top-down mass spectrometry using electrospray ionization (ESI) and a 15-Tesla Fourier transform ion cyclotron resonance (FTICR) mass spectrometer. Native MS preserves the non-covalent interactions between the globin subunits, and electron capture dissociation (ECD) produces fragments directly from the intact Hb complex without dissociating the subunits. Using activated ion ECD, we observe the gradual unfolding process of the Hb complex in the gas phase. Without protein ion activation, the native Hb shows very limited ECD fragmentation from the N-termini, suggesting a tightly packed structure of the native complex and therefore low fragmentation efficiency. Precursor ion activation allows steady increase of N-terminal fragment ions, while the C-terminal fragments remain limited (38 c ions and 4 z ions on the α chain; 36 c ions and 2 z ions on the β chain). This ECD fragmentation pattern suggests that upon activation, the Hb complex starts to unfold from the N-termini of both subunits, whereas the C-terminal regions and therefore the potential regions involved in the subunit binding interactions remain intact. ECD-MS of the Hb dimer show similar fragmentation patterns as the Hb tetramer, providing further evidence for the hypothesized unfolding process of the Hb complex in the gas phase. Native top-down ECD-MS allows efficient probing of the Hb complex structure and the subunit binding interactions in the gas phase. Finally, it may provide a fast and effective means to probe the structure of novel protein complexes that are intractable to traditional structural characterization tools.},
doi = {10.1255/ejms.1340},
journal = {European Journal of Mass Spectrometry},
number = 3,
volume = 21,
place = {United States},
year = {Tue Jun 09 00:00:00 EDT 2015},
month = {Tue Jun 09 00:00:00 EDT 2015}
}
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Works referenced in this record:

Proteome survey reveals modularity of the yeast cell machinery
journal, January 2006

  • Gavin, Anne-Claude; Aloy, Patrick; Grandi, Paola
  • Nature, Vol. 440, Issue 7084
  • DOI: 10.1038/nature04532

High-Quality Binary Protein Interaction Map of the Yeast Interactome Network
journal, October 2008

Bioinformatics
book, January 2008

Combined Electron Capture and Infrared Multiphoton Dissociation for Multistage MS/MS in a Fourier Transform Ion Cyclotron Resonance Mass Spectrometer
journal, July 2003

  • Håkansson, Kristina; Chalmers, Michael J.; Quinn, John P.
  • Analytical Chemistry, Vol. 75, Issue 13
  • DOI: 10.1021/ac030015q

Folding and assembly of hemoglobin monitored by electrospray mass spectrometry using an on-line dialysis system
journal, January 2007

  • Boys, Brian L.; Konermann, Lars
  • Journal of the American Society for Mass Spectrometry, Vol. 18, Issue 1
  • DOI: 10.1016/j.jasms.2006.08.013

Top-Down Quantitative Proteomics Identified Phosphorylation of Cardiac Troponin I as a Candidate Biomarker for Chronic Heart Failure
journal, September 2011

  • Zhang, Jiang; Guy, Moltu J.; Norman, Holly S.
  • Journal of Proteome Research, Vol. 10, Issue 9
  • DOI: 10.1021/pr200258m

Elucidating the site of protein-ATP binding by top-down mass spectrometry
journal, June 2010

A Protein Interaction Map of Drosophila melanogaster
journal, December 2003

A Human Protein-Protein Interaction Network: A Resource for Annotating the Proteome
journal, September 2005

Protein–Protein Binding Affinities in Solution Determined by Electrospray Mass Spectrometry
journal, February 2011

  • Liu, Jiangjiang; Konermann, Lars
  • Journal of The American Society for Mass Spectrometry, Vol. 22, Issue 3
  • DOI: 10.1007/s13361-010-0052-1

Top-Down ESI-ECD-FT-ICR Mass Spectrometry Localizes Noncovalent Protein-Ligand Binding Sites
journal, November 2006

  • Xie, Yongming; Zhang, Jennifer; Yin, Sheng
  • Journal of the American Chemical Society, Vol. 128, Issue 45
  • DOI: 10.1021/ja063197p

Atomic resolution cryo electron microscopy of macromolecular complexes
book, April 2011

Top-down mass spectrometry of supercharged native protein–ligand complexes
journal, March 2011

Huntingtin–protein interactions and the pathogenesis of Huntington's disease
journal, March 2004

Protein–protein interactions in human disease
journal, August 2005

Systematic Interactome Mapping and Genetic Perturbation Analysis of a C. elegans TGF-β Signaling Network
journal, February 2004

Native Electrospray and Electron-Capture Dissociation FTICR Mass Spectrometry for Top-Down Studies of Protein Assemblies
journal, July 2011

  • Zhang, Hao; Cui, Weidong; Wen, Jianzhong
  • Analytical Chemistry, Vol. 83, Issue 14
  • DOI: 10.1021/ac200695d

Mass Spectrometry in the Study of Hemoglobin: from Covalent Structure to Higher Order Assembly
journal, March 2006

The Nitty-Gritty of Protein Interactions
journal, December 2005

Highly Asymmetric Interactions between Globin Chains during Hemoglobin Assembly Revealed by Electrospray Ionization Mass Spectrometry †
journal, August 2003

  • Griffith, Wendell P.; Kaltashov, Igor A.
  • Biochemistry, Vol. 42, Issue 33
  • DOI: 10.1021/bi034035y

Global landscape of protein complexes in the yeast Saccharomyces cerevisiae
journal, March 2006

  • Krogan, Nevan J.; Cagney, Gerard; Yu, Haiyuan
  • Nature, Vol. 440, Issue 7084
  • DOI: 10.1038/nature04670

Gas-phase dissociation of hemoglobin
journal, September 2001

Unfolding of proteins monitored by electrospray ionization mass spectrometry: a comparison of positive and negative ion modes
journal, December 1998

Structure of Hæmoglobin: A Three-Dimensional Fourier Synthesis at 5.5-Å. Resolution, Obtained by X-Ray Analysis
journal, February 1960

  • Perutz, M. F.; Rossmann, M. G.; Cullis, Ann F.
  • Nature, Vol. 185, Issue 4711
  • DOI: 10.1038/185416a0

Electrostatic Stabilization of a Native Protein Structure in the Gas Phase
journal, November 2010

  • Breuker, Kathrin; Brüschweiler, Sven; Tollinger, Martin
  • Angewandte Chemie International Edition, Vol. 50, Issue 4
  • DOI: 10.1002/anie.201005112

Protein interaction mapping: A Drosophila case study
journal, February 2005

A Protein Interaction Network Links GIT1, an Enhancer of Huntingtin Aggregation, to Huntington's Disease
journal, September 2004

Systematic identification of protein complexes in Saccharomyces cerevisiae by mass spectrometry
journal, January 2002

  • Ho, Yuen; Gruhler, Albrecht; Heilbut, Adrian
  • Nature, Vol. 415, Issue 6868
  • DOI: 10.1038/415180a

Observation of the heme-globin complex in native myoglobin by electrospray-ionization mass spectrometry
journal, October 1991

  • Katta, Viswanatham; Chait, Brian T.
  • Journal of the American Chemical Society, Vol. 113, Issue 22
  • DOI: 10.1021/ja00022a058

Native electrospray ionization and electron-capture dissociation for comparison of protein structure in solution and the gas phase
journal, November 2013

  • Zhang, Hao; Cui, Weidong; Gross, Michael L.
  • International Journal of Mass Spectrometry, Vol. 354-355
  • DOI: 10.1016/j.ijms.2013.06.019

Interaction between amyloid precursor protein and presenilins in mammalian cells: Implications for the pathogenesis of Alzheimer disease
journal, July 1997

  • Xia, W.; Zhang, J.; Perez, R.
  • Proceedings of the National Academy of Sciences, Vol. 94, Issue 15
  • DOI: 10.1073/pnas.94.15.8208

The Role of Mass Spectrometry in Structure Elucidation of Dynamic Protein Complexes
journal, June 2007

Aberrant Interactions of Transcriptional Repressor Proteins with the Huntington's Disease Gene Product, Huntingtin
journal, September 1999

  • Boutell, J. M.; Thomas, P.; Neal, J. W.
  • Human Molecular Genetics, Vol. 8, Issue 9
  • DOI: 10.1093/hmg/8.9.1647

Native Electrospray and Electron-Capture Dissociation in FTICR Mass Spectrometry Provide Top-Down Sequencing of a Protein Component in an Intact Protein Assembly
journal, December 2010

  • Zhang, Hao; Cui, Weidong; Wen, Jianzhong
  • Journal of the American Society for Mass Spectrometry, Vol. 21, Issue 12
  • DOI: 10.1016/j.jasms.2010.08.006

Revealing Ligand Binding Sites and Quantifying Subunit Variants of Noncovalent Protein Complexes in a Single Native Top-Down FTICR MS Experiment
journal, June 2014

  • Li, Huilin; Wongkongkathep, Piriya; Van Orden, Steve L.
  • Journal of The American Society for Mass Spectrometry, Vol. 25, Issue 12
  • DOI: 10.1007/s13361-014-0928-6

Native Top-Down Electrospray Ionization-Mass Spectrometry of 158 kDa Protein Complex by High-Resolution Fourier Transform Ion Cyclotron Resonance Mass Spectrometry
journal, December 2013

  • Li, Huilin; Wolff, Jeremy J.; Van Orden, Steve L.
  • Analytical Chemistry, Vol. 86, Issue 1
  • DOI: 10.1021/ac4033214

Collisional activation of protein complexes: Picking up the pieces
journal, March 2009

Towards a proteome-scale map of the human protein–protein interaction network
journal, September 2005

  • Rual, Jean-François; Venkatesan, Kavitha; Hao, Tong
  • Nature, Vol. 437, Issue 7062
  • DOI: 10.1038/nature04209

Analysis of protein mixtures by electrospray mass spectrometry: Effects of conformation and desolvation behavior on the signal intensities of hemoglobin subunits
journal, July 2007

  • Kuprowski, Mark C.; Boys, Brian L.; Konermann, Lars
  • Journal of the American Society for Mass Spectrometry, Vol. 18, Issue 7
  • DOI: 10.1016/j.jasms.2007.04.002

Analysis of the human protein interactome and comparison with yeast, worm and fly interaction datasets
journal, February 2006

  • Gandhi, T. K. B.; Zhong, Jun; Mathivanan, Suresh
  • Nature Genetics, Vol. 38, Issue 3
  • DOI: 10.1038/ng1747

Electrospray ionization mass spectrometry: a technology for studying noncovalent macromolecular complexes
journal, December 2000

Studying 18 MDa Virus Assemblies with Native Mass Spectrometry
journal, February 2013

  • Snijder, Joost; Rose, Rebecca J.; Veesler, David
  • Angewandte Chemie International Edition, Vol. 52, Issue 14
  • DOI: 10.1002/anie.201210197

Activated Ion Electron Capture Dissociation for Mass Spectral Sequencing of Larger (42 kDa) Proteins
journal, October 2000

  • Horn, David M.; Ge, Ying; McLafferty, Fred W.
  • Analytical Chemistry, Vol. 72, Issue 20
  • DOI: 10.1021/ac000494i

Molecular dynamics of protein complexes from four-dimensional cryo-electron microscopy
journal, September 2004

  • Heymann, J. Bernard; Conway, James F.; Steven, Alasdair C.
  • Journal of Structural Biology, Vol. 147, Issue 3
  • DOI: 10.1016/j.jsb.2004.02.006

The way to NMR structures of proteins
journal, November 2001

  • Wüthrich, Kurt
  • Nature Structural Biology, Vol. 8, Issue 11, p. 923-925
  • DOI: 10.1038/nsb1101-923

A Map of the Interactome Network of the Metazoan C. elegans
journal, January 2004

Staphylococcus aureus Uses a Novel Multidomain Receptor to Break Apart Human Hemoglobin and Steal Its Heme
journal, November 2012

  • Spirig, Thomas; Malmirchegini, G. Reza; Zhang, Jiang
  • Journal of Biological Chemistry, Vol. 288, Issue 2
  • DOI: 10.1074/jbc.M112.419119

Deciphering modifications in swine cardiac troponin I by top-down high-resolution tandem mass spectrometry
journal, June 2010

  • Zhang, Jiang; Dong, Xintong; Hacker, Timothy A.
  • Journal of the American Society for Mass Spectrometry, Vol. 21, Issue 6
  • DOI: 10.1016/j.jasms.2010.02.005

Mapping a Noncovalent Protein–Peptide Interface by Top-Down FTICR Mass Spectrometry Using Electron Capture Dissociation
journal, May 2011

  • Clarke, David J.; Murray, Euan; Hupp, Ted
  • Journal of The American Society for Mass Spectrometry, Vol. 22, Issue 8
  • DOI: 10.1007/s13361-011-0155-3

Protein Structure Determination by X-Ray Crystallography
book, January 2008

Protein structure determination in solution by NMR spectroscopy.
journal, December 1990

Electrostatic Stabilization of a Native Protein Structure in the Gas Phase
journal, November 2010

  • Breuker, Kathrin; Brüschweiler, Sven; Tollinger, Martin
  • Angewandte Chemie, Vol. 123, Issue 4
  • DOI: 10.1002/ange.201005112

A Protein Interaction Network Links GIT1, an Enhancer of Huntingtin Aggregation, to Huntington’s Disease
journal, July 2005

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