title: Many overlapping peptides for protein hydrogen exchange experiments by the fragment separation-mass spectrometry method.
abstract: Measurement of the naturally occurring hydrogen exchange (HX) behavior of proteins can in principle provide highly resolved thermodynamic and kinetic information on protein structure, dynamics, and interactions. The HX fragment separation-mass spectrometry method (HX-MS) is able to measure hydrogen exchange in biologically important protein systems that are not accessible to NMR methods. In order to achieve high structural resolution in HX-MS experiments, it will be necessary to obtain many sequentially overlapping peptide fragments and be able to identify and analyze them efficiently and accurately by mass spectrometry. This paper describes operations which, when applied to four different proteins ranging in size from 140 to 908 residues, routinely provides hundreds of useful unique peptides, covering the entire protein length many times over. Coverage in terms of the average number of peptide fragments that span each amino acid exceeds 10. The ability to achieve these results required the integrated application of experimental methods that are described here and a computer analysis program, called ExMS, described in a following paper.
authors.affiliation: Johnson Research Foundation, Department of Biochemistry and Biophysics, Perelman School of Medicine, University of Pennsylvania, 1006 Stellar-Chance Labs, 422 Curie Boulevard, Philadelphia, PA 19104, USA. mayne@mail.med.upenn.edu
authors.forename: Leland
authors.initials: L
authors.lastname: Mayne
authors.forename: Zhong-Yuan
authors.initials: ZY
authors.lastname: Kan
authors.forename: Palaniappan Sevugan
authors.initials: PS
authors.lastname: Chetty
authors.forename: Alec
authors.initials: A
authors.lastname: Ricciuti
authors.forename: Benjamin T
authors.initials: BT
authors.lastname: Walters
authors.forename: S Walter
authors.initials: SW
authors.lastname: Englander
mesh_terms: D041961:Deuterium Exchange Measurement / Q000379:methods*; D004867:Equipment Design; D006801:Humans; D010446:Peptide Fragments / Q000737:chemistry*; D010449:Peptide Mapping / Q000379:methods*; D011506:Proteins / Q000737:chemistry; D011994:Recombinant Proteins; D015203:Reproducibility of Results; D012984:Software; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D010446:Peptide Fragments; D011506:Proteins; D011994:Recombinant Proteins