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      Recommendations for performing, interpreting and reporting hydrogen deuterium exchange mass spectrometry (HDX-MS) experiments

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      iD , 1 , * , iD , 2 , * , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 , 8 , 18 , 19 , 20 , 21 , 22 , 9 , 23 , 24 , 25 , 14 , 12 , 26 , 27 , 8 , 28 , 29 , 30 , 31 , 32 , 12 , iD , 33 , * , 34 , *
      Nature methods

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          Abstract

          Hydrogen deuterium exchange mass spectrometry (HDX-MS) is a powerful biophysical technique being increasingly applied to a wide variety of problems. As the HDX-MS community continues to grow, adoption of best practices in data collection, analysis, presentation and interpretation will greatly enhance the accessibility of this technique to nonspecialists. Here we provide recommendations arising from community discussions emerging out of the first International Conference on Hydrogen-Exchange Mass Spectrometry (IC-HDX; 2017). It is meant to represent both a consensus viewpoint and an opportunity to stimulate further additions and refinements as the field advances.

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          Most cited references48

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          Hydrogen exchange mass spectrometry for studying protein structure and dynamics.

          Hydrogen/deuterium exchange (HDX) mass spectrometry (MS) has become a key technique for monitoring structural and dynamic aspects of proteins in solution. This approach relies on the fact that exposure of a protein to D(2)O induces rapid amide H → D exchange in disordered regions that lack stable hydrogen-bonding. Tightly folded elements are much more protected from HDX, resulting in slow isotope exchange that is mediated by the structural dynamics ("breathing motions") of the protein. MS-based peptide mapping is a well established technique for measuring the mass shifts of individual protein segments. This tutorial review briefly discusses basic fundamentals of HDX/MS, before highlighting a number of recent developments and applications. Gas phase fragmentation strategies represent a promising alternative to the traditional proteolysis-based approach, but experimentalists have to be aware of scrambling phenomena that can be encountered under certain conditions. Electron-based dissociation methods provide a solution to this problem. We also discuss recent advances that facilitate the applicability of HDX/MS to membrane proteins, and to the characterization of short-lived protein folding intermediates. It is hoped that this review will provide a starting point for novices, as well as a useful reference for practitioners, who require an overview of some recent trends in HDX/MS.
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            Crystal structure of the PTEN tumor suppressor: implications for its phosphoinositide phosphatase activity and membrane association.

            The PTEN tumor suppressor is mutated in diverse human cancers and in hereditary cancer predisposition syndromes. PTEN is a phosphatase that can act on both polypeptide and phosphoinositide substrates in vitro. The PTEN structure reveals a phosphatase domain that is similar to protein phosphatases but has an enlarged active site important for the accommodation of the phosphoinositide substrate. The structure also reveals that PTEN has a C2 domain. The PTEN C2 domain binds phospholipid membranes in vitro, and mutation of basic residues that could mediate this reduces PTEN's membrane affinity and its ability to suppress the growth of glioblastoma tumor cells. The phosphatase and C2 domains associate across an extensive interface, suggesting that the C2 domain may serve to productively position the catalytic domain on the membrane.
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              Hydrogen exchange mass spectrometry for the analysis of protein dynamics.

              Hydrogen exchange coupled to mass spectrometry (MS) has become a valuable analytical tool for the study of protein dynamics. By combining information about protein dynamics with more classical functional data, a more thorough understanding of protein function can be obtained. In many cases, protein dynamics are directly related to specific protein functions such as conformational changes during enzyme activation or protein movements during binding. The method is made possible because labile backbone hydrogens in a protein will exchange with deuterium atoms when the protein is placed in a D2O solution. The subsequent increase in protein mass over time is measured with high-resolution MS. The location of the deuterium incorporation is determined by monitoring deuterium incorporation in peptic fragments that are produced after the labeling reaction. In this review, we will summarize the general principles of the method, discuss the latest variations on the experimental protocol that probe different types of protein movements, and review other recent work and improvements in the field. Copyright 2005 Wiley Periodicals, Inc.
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                Author and article information

                Journal
                101215604
                Nat Methods
                Nat. Methods
                Nature methods
                1548-7091
                1548-7105
                28 June 2019
                27 June 2019
                08 July 2019
                : 16
                : 7
                : 595-602
                Affiliations
                [1 ]MRC Laboratory of Molecular Biology, Cambridge, UK
                [2 ]Department of Biochemistry and Microbiology, University of Victoria, Victoria, BC, Canada
                [3 ]Department of Biochemistry, University of Colorado, Boulder, CO, USA
                [4 ]Department of Biological Science, National University of Singapore, Singapore, Singapore
                [5 ]Genome BC Proteomics Centre, University of Victoria, Victoria, BC, Canada
                [6 ]Institut Pasteur, Chemistry and Structural Biology Department, Paris, France
                [7 ]Biogen Idec, Cambridge, MA, USA
                [8 ]Department of Chemistry and Chemical Biology, Northeastern University, Boston, MA, USA
                [9 ]Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia, PA, USA
                [10 ]Novo Nordisk, Måløv, Denmark
                [11 ]UCB Celltech, Slough, UK
                [12 ]Department of Integrative Structural and Computational Biology, Scripps Florida, The Scripps Research Institute, Jupiter, FL, USA
                [13 ]Department of Chemistry, Washington University in St. Louis, St. Louis, MO, USA
                [14 ]Department of Medicinal Chemistry, School of Pharmacy, University of Washington, Seattle, WA, USA
                [15 ]Johnson & Johnson Pharmaeutical Research and Development, Jersey City, NJ, USA
                [16 ]Bijvoet Center for Biomolecular Research, Utrecht University, Utrecht, the Netherlands
                [17 ]Codiak Biosciences, Cambridge, MA, USA
                [18 ]Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej, Odense, Denmark
                [19 ]Department of Chemistry, University of Massachusetts-Amherst, Amherst, MA, USA
                [20 ]Department of Chemistry, University of California, Berkeley, Berkeley, CA, USA
                [21 ]Department of Chemistry, The University of Western Ontario, London, ON, Canada
                [22 ]Institute of Microbiology, Czech Academy of Sciences, Prague, Czech Republic
                [23 ]Department of Molecular Medicine, The Scripps Research Institute, Jupiter, FL, USA
                [24 ]Department of Biological Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel
                [25 ]MRC Laboratory of Molecular Biology, Cambridge, UK
                [26 ]Roy J. Carver Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA, USA
                [27 ]Roche Innovation Center Penzberg, Penzberg, Germany
                [28 ]Department of Early Stage Pharmaceutical Development, Genentech, Inc., South San Francisco, CA, USA
                [29 ]Department of Pharmaceutical Chemistry, University of Kansas, Lawrence, KS, USA
                [30 ]Department of Chemistry, York University, Toronto, ON, Canada
                [31 ]Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, Baltimore, MD, USA
                [32 ]Process Development, Amgen, Thousand Oaks, CA, USA
                [33 ]Department of Biochemistry & Molecular Biology, University of Calgary, Calgary, AB, Canada
                [34 ]Department of Pharmacy, University of Copenhagen, Copenhagen, Denmark
                Author notes
                [* ] Correspondence should be addressed to G.R.M., J.E.B., D.C.S. or K.D.R. gmasson@ 123456mrc-lmb.cam.ac.uk ; jeburke@ 123456uvic.ca ; dschriem@ 123456ucalgary.ca ; kasper.rand@ 123456sund.ku.dk
                Article
                EMS83563
                10.1038/s41592-019-0459-y
                6614034
                31249422
                4f379591-267b-4b81-883e-4a98c842c851

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