Quantitative Protein Topography Measurements by High Resolution Hydroxyl Radical Protein Footprinting Enable Accurate Molecular Model Selection

Abstract We report an integrated workflow that allows mass spectrometry-based high-resolution hydroxyl radical protein footprinting (HR-HRPF) measurements to accurately measure the absolute average solvent accessible surface area (<SASA>) of amino acid side chains. This approach is based on ap...

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Autores principales: Boer Xie, Amika Sood, Robert J. Woods, Joshua S. Sharp
Formato: article
Lenguaje:EN
Publicado: Nature Portfolio 2017
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Acceso en línea:https://doaj.org/article/430818b6609d4251b938e8a82cd1b561
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Sumario:Abstract We report an integrated workflow that allows mass spectrometry-based high-resolution hydroxyl radical protein footprinting (HR-HRPF) measurements to accurately measure the absolute average solvent accessible surface area (<SASA>) of amino acid side chains. This approach is based on application of multi-point HR-HRPF, electron-transfer dissociation (ETD) tandem MS (MS/MS) acquisition, measurement of effective radical doses by radical dosimetry, and proper normalization of the inherent reactivity of the amino acids. The accuracy of the resulting <SASA> measurements was tested by using well-characterized protein models. Moreover, we demonstrated the ability to use <SASA> measurements from HR-HRPF to differentiate molecular models of high accuracy (<3 Å backbone RMSD) from models of lower accuracy (>4 Å backbone RMSD). The ability of <SASA> data from HR-HRPF to differentiate molecular model quality was found to be comparable to that of <SASA> data obtained from X-ray crystal structures, indicating the accuracy and utility of HR-HRPF for evaluating the accuracy of computational models.