Apparent activation energies of protein–protein complex dissociation in the gas–phase determined by electrospray mass spectrometry

Yelena Yefremova, F. Teresa I. Melder, Bright D. Danquah, Kwabena F.M. Opuni, Cornelia Koy, Alexandra Ehrens, David Frommholz, Harald Illges, Knut Koelbel, Frank Sobott, Michael O. Glocker

Research output: Contribution to journalArticlepeer-review

15 Citations (Scopus)

Abstract

We have developed a method to determine apparent activation energies of dissociation for ionized protein–protein complexes in the gas phase using electrospray ionization mass spectrometry following the Rice-Ramsperger-Kassel-Marcus quasi-equilibrium theory. Protein–protein complexes were formed in solution, transferred into the gas phase, and separated from excess free protein by ion mobility filtering. Afterwards, complex disassembly was initiated by collision-induced dissociation with step-wise increasing energies. Relative intensities of ion signals were used to calculate apparent activation energies of dissociation in the gas phase by applying linear free energy relations. The method was developed using streptavidin tetramers. Experimentally determined apparent gas-phase activation energies for dissociation (EAm0g#) of complexes consisting of Fc parts from immunoglobulins (IgG-Fc) and three closely related protein G' variants (IgG-Fc•protein G'e, IgG-Fc•protein G'f, and IgG-Fc•protein G'g) show the same order of stabilities as can be inferred from their in-solution binding constants. Differences in stabilities between the protein–protein complexes correspond to single amino acid residue exchanges in the IgG-binding regions of the protein G' variants. [Figure not available: see fulltext.].

Original languageEnglish
Pages (from-to)6549-6558
Number of pages10
JournalAnalytical and Bioanalytical Chemistry
Volume409
Issue number28
DOIs
Publication statusPublished - 1 Nov 2017

Keywords

  • Collision induced dissociation
  • Ion mobility
  • Native mass spectrometry
  • Protein–protein interaction
  • Quasi equilibrium conditions

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