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Title: Fourier Transform-Ion Cyclotron Resonance Mass Spectrometry as a Platform for Characterizing Multimeric Membrane Protein Complexes

Abstract

We report that membrane protein characterization is consistently hampered by challenges with expression, purification and solubilization. Among several biophysical techniques employed for their characterization, native-mass spectrometry (MS) has emerged as a powerful tool for the analysis of membrane proteins and complexes. Here, two MS platforms, the FT-ICR and QToF, have been explored to analyze the homotetrameric water channel protein, AquaporinZ (AqpZ) under non-denaturing conditions. This 97 kDa membrane protein complex can be readily liberated from the octylglucoside (OG) detergent micelle under a range of instrument conditions on both MS platforms. Increasing the applied collision energy of the FT-ICR collision cell yielded varying degrees of tetramer (97 kDa) liberation from the OG micelles, as well as dissociation into the trimeric (72 kDa) and monomeric (24 kDa) substituents. Tandem-MS on the Q-ToF yielded higher intensity tetramer signal and, depending on the m/z region selected, the observed monomer signal varied in intensity. Precursor ion selection of an m/z range above the expected protein signal distribution, followed by mild collisional activation is able to efficiently liberate AqpZ with a high S/N ratio. Lastly, the tetrameric charge state distribution obtained on both instruments demonstrated superpositioning of multiple proteoforms due to varying degrees of N-terminal formylation.

Authors:
 [1];  [2];  [1];  [2];  [2];  [1]
  1. Discovery Attribute Sciences, Amgen, Thousand Oaks, CA (United States)
  2. 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:
1511586
Grant/Contract Number:  
FC03-02ER63421
Resource Type:
Accepted Manuscript
Journal Name:
Journal of the American Society for Mass Spectrometry
Additional Journal Information:
Journal Volume: 29; Journal Issue: 1; Journal ID: ISSN 1044-0305
Publisher:
American Society for Mass Spectrometry
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; Native-mass spectrometry; Membrane protein; Fourier transform ion cyclotron resonance

Citation Formats

Lippens, Jennifer L., Nshanian, Michael, Spahr, Chris, Egea, Pascal F., Loo, Joseph A., and Campuzano, Iain D. G. Fourier Transform-Ion Cyclotron Resonance Mass Spectrometry as a Platform for Characterizing Multimeric Membrane Protein Complexes. United States: N. p., 2017. Web. doi:10.1007/s13361-017-1799-4.
Lippens, Jennifer L., Nshanian, Michael, Spahr, Chris, Egea, Pascal F., Loo, Joseph A., & Campuzano, Iain D. G. Fourier Transform-Ion Cyclotron Resonance Mass Spectrometry as a Platform for Characterizing Multimeric Membrane Protein Complexes. United States. doi:10.1007/s13361-017-1799-4.
Lippens, Jennifer L., Nshanian, Michael, Spahr, Chris, Egea, Pascal F., Loo, Joseph A., and Campuzano, Iain D. G. Mon . "Fourier Transform-Ion Cyclotron Resonance Mass Spectrometry as a Platform for Characterizing Multimeric Membrane Protein Complexes". United States. doi:10.1007/s13361-017-1799-4. https://www.osti.gov/servlets/purl/1511586.
@article{osti_1511586,
title = {Fourier Transform-Ion Cyclotron Resonance Mass Spectrometry as a Platform for Characterizing Multimeric Membrane Protein Complexes},
author = {Lippens, Jennifer L. and Nshanian, Michael and Spahr, Chris and Egea, Pascal F. and Loo, Joseph A. and Campuzano, Iain D. G.},
abstractNote = {We report that membrane protein characterization is consistently hampered by challenges with expression, purification and solubilization. Among several biophysical techniques employed for their characterization, native-mass spectrometry (MS) has emerged as a powerful tool for the analysis of membrane proteins and complexes. Here, two MS platforms, the FT-ICR and QToF, have been explored to analyze the homotetrameric water channel protein, AquaporinZ (AqpZ) under non-denaturing conditions. This 97 kDa membrane protein complex can be readily liberated from the octylglucoside (OG) detergent micelle under a range of instrument conditions on both MS platforms. Increasing the applied collision energy of the FT-ICR collision cell yielded varying degrees of tetramer (97 kDa) liberation from the OG micelles, as well as dissociation into the trimeric (72 kDa) and monomeric (24 kDa) substituents. Tandem-MS on the Q-ToF yielded higher intensity tetramer signal and, depending on the m/z region selected, the observed monomer signal varied in intensity. Precursor ion selection of an m/z range above the expected protein signal distribution, followed by mild collisional activation is able to efficiently liberate AqpZ with a high S/N ratio. Lastly, the tetrameric charge state distribution obtained on both instruments demonstrated superpositioning of multiple proteoforms due to varying degrees of N-terminal formylation.},
doi = {10.1007/s13361-017-1799-4},
journal = {Journal of the American Society for Mass Spectrometry},
number = 1,
volume = 29,
place = {United States},
year = {2017},
month = {10}
}

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