Quantifying Protein–Protein Interactions Within Noncovalent Complexes Using Electrospray Ionization Mass Spectrometry

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Author(s) Erba, Elisabetta Boeri, Barylyuk, Konstantin, Yang, Yang, Zenobi, Renato
Publication Type Journal Items, Publication Status: Published
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Title Quantifying Protein–Protein Interactions Within Noncovalent Complexes Using Electrospray Ionization Mass Spectrometry
Author(s) Erba, Elisabetta Boeri
Barylyuk, Konstantin
Yang, Yang
Zenobi, Renato
Journal or Series Title Analytical chemistry
Volume Number 83
Issue Number 24
Start Page 9251
End Page 9259
ISSN 0003-2700
1520-6882
Publisher American Chemical Society
Publication Place Washington, D.C.
Publication Date 2011-11
Abstract Several electrospray-mass spectrometry (ESI-MS)-based methods are available for determining the constant of association (K(a)) between a protein and a small ligand, but current MS-based strategies are not fully adequate for measuring K(a) of protein-protein interactions accurately. We expanded the application of ESI-MS-based titration to determine the strength of noncovalent interactions between proteins, forming a complex. Taking into account relative response factors (probability of being ionized, transmitted, and detected), we determined K(a) values of an equilibrium between dimers and tetramers at three different pH values (6.8, 3.4, and 8.4). We investigated the association of the lectin concanavalin A, whose dimer-tetramer ratio in the gas phase is affected by solution concentration and by pH. To calculate the constants of association in solution, we also utilized isothermal titration calorimetry (ITC) for a comparison with MS-based titration. At pH 6.8 and pH 8.4, the K(a) values measured by MS and by ITC were in agreement. ITC results allowed us to restrain the response factor to a value close to 4. At pH 3.4, we were able to measure the K(a) only by MS, but not by ITC because of limited sensitivity of calorimetry. Our investigation illustrates the great potential MS for calculating the binding strength of protein-protein interactions within noncovalent complexes. The main advantages of MS over ITC are its sensitivity (i.e., the required amount of sample is >100 times less than the one necessary for ITC), and the possibility to obtain precise information on composition of protein complexes, their stoichiometry, their subunit interactions, and their assembly pathway. Compared to previous investigations, our study shows the strong influence of response factors on determining accurate protein-protein association constants by MS.
DOI 10.1021/ac201576e
Additional Notes Received 21 June 2011, Accepted 02 November 2011, Published 02 November 2011
Document Type Article
Publication Status Published
Language English
Assigned Organisational Unit(s) 03430
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NEBIS System Number 000005465
Source Database ID WOS-000297946900012
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@article{rb2011,
  author = "Erba, Elisabetta Boeri and Barylyuk, Konstantin and Yang, Yang and Zenobi, Renato",
  title = "{Q}uantifying {P}rotein{\textendash}{P}rotein {I}nteractions {W}ithin {N}oncovalent {C}omplexes {U}sing {E}lectrospray {I}onization {M}ass {S}pectrometry",
  journal = "Analytical chemistry",
  year = 2011,
  volume = "83",
  number = "24",
  pages = "9251--9259",
  month = nov,
}


E-Citations record created: Mon, 16 Jan 2012, 08:24:30 CET