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Direct monitoring of protein-chemical reactions utilising nanoelectrospray mass spectrometry

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Journal of the American Society for Mass Spectrometry

Abstract

The feasibility of nanoelectrospray mass spectrometry (nanoESI) for the direct analysis of protein chemical reactions and structural changes of proteins has been evaluated. Taking advantage of the long spraying time and the capability of nanoESI for employing a wide range of solvent conditions such as buffers and detergents, applications of monitoring reaction pathways, and dynamics have been carried out with several peptides and proteins. The time course of proteolytic digestions with trypsin and pepsin was investigated for several model polypeptides, and nanoESI showed to provide an efficient tool for optimising digestion conditions for the mass spectrometric peptide mapping analysis. Examples of specific protein chemical modification reactions at arginine and tyrosine residues illustrate the feasibility of nanoESI to monitoring reaction yields and modification sites for more than 180 min. Furthermore, changes of the pattern of protonated molecules caused by temperature effects and by protein unfolding due to disulfide bond reduction have been studied with the model proteins cytochrome c and hen eggwhite lysozyme. The results indicate that nanoESI is an efficient technique for the direct, molecular characterisation of protein-chemical reactions in solution.

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Authors and Affiliations

  1. Fakultät für Chemie, Universität Konstanz, 78457, Konstanz, Germany

    Thilo A. Fligge, Jürgen Kast, Kai Bruns & Michael Przybylski

Authors
  1. Thilo A. Fligge
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  2. Jürgen Kast
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  3. Kai Bruns
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  4. Michael Przybylski
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Correspondence to Michael Przybylski.

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Fligge, T.A., Kast, J., Bruns, K. et al. Direct monitoring of protein-chemical reactions utilising nanoelectrospray mass spectrometry. J Am Soc Mass Spectrom 10, 112–118 (1999). https://doi.org/10.1016/S1044-0305(98)00131-7

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  • DOI: https://doi.org/10.1016/S1044-0305(98)00131-7

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