Mass Spectrometric Analysis of Antibody—Epitope Peptide Complex Dissociation: Theoretical Concept and Practical Procedure of Binding Strength Characterization
Electrospray mass spectrometry is applied to determine apparent binding energies and quasi equilibrium dissociation constants of immune complex dissociation reactions in the gas phase. Myoglobin, a natural protein-ligand complex, has been used to develop the procedure which starts from determining m...
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MDPI AG
2020-10-01
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| Series: | Molecules |
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| Online Access: | https://www.mdpi.com/1420-3049/25/20/4776 |
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| author | Bright D. Danquah Kwabena F. M. Opuni Claudia Roewer Cornelia Koy Michael O. Glocker |
| author_facet | Bright D. Danquah Kwabena F. M. Opuni Claudia Roewer Cornelia Koy Michael O. Glocker |
| author_sort | Bright D. Danquah |
| collection | DOAJ |
| description | Electrospray mass spectrometry is applied to determine apparent binding energies and quasi equilibrium dissociation constants of immune complex dissociation reactions in the gas phase. Myoglobin, a natural protein-ligand complex, has been used to develop the procedure which starts from determining mean charge states and normalized and averaged ion intensities. The apparent dissociation constant <inline-formula><math display="inline"><semantics><mrow><msubsup><mi>K</mi><mrow><mi>D</mi><mo> </mo><mi>m</mi><mn>0</mn><mi>g</mi></mrow><mo>#</mo></msubsup><mo>=</mo></mrow></semantics></math></inline-formula> 3.60 × 10<sup>−12</sup> for the gas phase heme dissociation process was calculated from the mass spectrometry data and by subsequent extrapolation to room temperature to mimic collision conditions for neutral and resting myoglobin. Similarly, for RNAse S dissociation at room temperature a <inline-formula><math display="inline"><semantics><mrow><msubsup><mi>K</mi><mrow><mi>D</mi><mo> </mo><mi>m</mi><mn>0</mn><mi>g</mi></mrow><mo>#</mo></msubsup><mo>=</mo></mrow></semantics></math></inline-formula> 4.03 × 10<sup>−12</sup> was determined. The protocol was tested with two immune complexes consisting of epitope peptides and monoclonal antibodies. For the epitope peptide dissociation reaction of the FLAG peptide from the antiFLAG antibody complex an apparent gas phase dissociation constant <inline-formula><math display="inline"><semantics><mrow><msubsup><mi>K</mi><mrow><mi>D</mi><mo> </mo><mi>m</mi><mn>0</mn><mi>g</mi></mrow><mo>#</mo></msubsup><mo>=</mo></mrow></semantics></math></inline-formula> 4.04 × 10<sup>−12</sup> was calculated. Likewise, an apparent <inline-formula><math display="inline"><semantics><mrow><msubsup><mi>K</mi><mrow><mi>D</mi><mo> </mo><mi>m</mi><mn>0</mn><mi>g</mi></mrow><mo>#</mo></msubsup><mo>=</mo></mrow></semantics></math></inline-formula> 4.58 × 10<sup>−12</sup> was calculated for the troponin I epitope peptide—antiTroponin I antibody immune complex dissociation. Electrospray mass spectrometry is a rapid method, which requires small sample amounts for either identification of protein-bound ligands or for determination of the apparent gas phase protein-ligand complex binding strengths. |
| first_indexed | 2024-03-10T15:32:54Z |
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| spelling | doaj.art-d6bf4e6d86614e5a8e439445d1079b7b2023-11-20T17:32:01ZengMDPI AGMolecules1420-30492020-10-012520477610.3390/molecules25204776Mass Spectrometric Analysis of Antibody—Epitope Peptide Complex Dissociation: Theoretical Concept and Practical Procedure of Binding Strength CharacterizationBright D. Danquah0Kwabena F. M. Opuni1Claudia Roewer2Cornelia Koy3Michael O. Glocker4Proteome Center Rostock, University Medicine Rostock, 18059 Rostock, GermanySchool of Pharmacy, University of Ghana, P. O. Box LG53 Legon, GhanaProteome Center Rostock, University Medicine Rostock, 18059 Rostock, GermanyProteome Center Rostock, University Medicine Rostock, 18059 Rostock, GermanyProteome Center Rostock, University Medicine Rostock, 18059 Rostock, GermanyElectrospray mass spectrometry is applied to determine apparent binding energies and quasi equilibrium dissociation constants of immune complex dissociation reactions in the gas phase. Myoglobin, a natural protein-ligand complex, has been used to develop the procedure which starts from determining mean charge states and normalized and averaged ion intensities. The apparent dissociation constant <inline-formula><math display="inline"><semantics><mrow><msubsup><mi>K</mi><mrow><mi>D</mi><mo> </mo><mi>m</mi><mn>0</mn><mi>g</mi></mrow><mo>#</mo></msubsup><mo>=</mo></mrow></semantics></math></inline-formula> 3.60 × 10<sup>−12</sup> for the gas phase heme dissociation process was calculated from the mass spectrometry data and by subsequent extrapolation to room temperature to mimic collision conditions for neutral and resting myoglobin. Similarly, for RNAse S dissociation at room temperature a <inline-formula><math display="inline"><semantics><mrow><msubsup><mi>K</mi><mrow><mi>D</mi><mo> </mo><mi>m</mi><mn>0</mn><mi>g</mi></mrow><mo>#</mo></msubsup><mo>=</mo></mrow></semantics></math></inline-formula> 4.03 × 10<sup>−12</sup> was determined. The protocol was tested with two immune complexes consisting of epitope peptides and monoclonal antibodies. For the epitope peptide dissociation reaction of the FLAG peptide from the antiFLAG antibody complex an apparent gas phase dissociation constant <inline-formula><math display="inline"><semantics><mrow><msubsup><mi>K</mi><mrow><mi>D</mi><mo> </mo><mi>m</mi><mn>0</mn><mi>g</mi></mrow><mo>#</mo></msubsup><mo>=</mo></mrow></semantics></math></inline-formula> 4.04 × 10<sup>−12</sup> was calculated. Likewise, an apparent <inline-formula><math display="inline"><semantics><mrow><msubsup><mi>K</mi><mrow><mi>D</mi><mo> </mo><mi>m</mi><mn>0</mn><mi>g</mi></mrow><mo>#</mo></msubsup><mo>=</mo></mrow></semantics></math></inline-formula> 4.58 × 10<sup>−12</sup> was calculated for the troponin I epitope peptide—antiTroponin I antibody immune complex dissociation. Electrospray mass spectrometry is a rapid method, which requires small sample amounts for either identification of protein-bound ligands or for determination of the apparent gas phase protein-ligand complex binding strengths.https://www.mdpi.com/1420-3049/25/20/4776mass spectrometric epitope mappinggas phase immune complex dissociationapparent gas phase dissociation constantsapparent gas phase activation energiesITEM-TWOnative mass spectrometry |
| spellingShingle | Bright D. Danquah Kwabena F. M. Opuni Claudia Roewer Cornelia Koy Michael O. Glocker Mass Spectrometric Analysis of Antibody—Epitope Peptide Complex Dissociation: Theoretical Concept and Practical Procedure of Binding Strength Characterization Molecules mass spectrometric epitope mapping gas phase immune complex dissociation apparent gas phase dissociation constants apparent gas phase activation energies ITEM-TWO native mass spectrometry |
| title | Mass Spectrometric Analysis of Antibody—Epitope Peptide Complex Dissociation: Theoretical Concept and Practical Procedure of Binding Strength Characterization |
| title_full | Mass Spectrometric Analysis of Antibody—Epitope Peptide Complex Dissociation: Theoretical Concept and Practical Procedure of Binding Strength Characterization |
| title_fullStr | Mass Spectrometric Analysis of Antibody—Epitope Peptide Complex Dissociation: Theoretical Concept and Practical Procedure of Binding Strength Characterization |
| title_full_unstemmed | Mass Spectrometric Analysis of Antibody—Epitope Peptide Complex Dissociation: Theoretical Concept and Practical Procedure of Binding Strength Characterization |
| title_short | Mass Spectrometric Analysis of Antibody—Epitope Peptide Complex Dissociation: Theoretical Concept and Practical Procedure of Binding Strength Characterization |
| title_sort | mass spectrometric analysis of antibody epitope peptide complex dissociation theoretical concept and practical procedure of binding strength characterization |
| topic | mass spectrometric epitope mapping gas phase immune complex dissociation apparent gas phase dissociation constants apparent gas phase activation energies ITEM-TWO native mass spectrometry |
| url | https://www.mdpi.com/1420-3049/25/20/4776 |
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