title: Fast photochemical oxidation of proteins for comparing solvent-accessibility changes accompanying protein folding: data processing and application to barstar.
abstract: Mass spectrometry-based protein footprinting reveals regional and even amino-acid structural changes and fills the gap for many proteins and protein interactions that cannot be studied by X-ray crystallography or NMR spectroscopy. Hydroxyl radical-mediated labeling has proven to be particularly informative in this pursuit because many solvent-accessible residues can be labeled by OH in a protein or protein complex, thus providing more coverage than does specific amino-acid modifications. Finding all the OH-labeling sites requires LC/MS/MS analysis of a proteolyzed sample, but data processing is daunting without the help of automated software. We describe here a systematic means for achieving a comprehensive residue-resolved analysis of footprinting data in an efficient manner, utilizing software common to proteomics core laboratories. To demonstrate the method and the utility of OH-mediated labeling, we show that FPOP easily distinguishes the buried and exposed residues of barstar in its folded and unfolded states. This article is part of a Special Issue entitled: Mass spectrometry in structural biology.
authors.affiliation: Donald Danforth Plant Science Center, Washington University, St. Louis, MO 63132, USA.
authors.forename: Brian C
authors.initials: BC
authors.lastname: Gau
authors.forename: Jiawei
authors.initials: J
authors.lastname: Chen
authors.forename: Michael L
authors.initials: ML
authors.lastname: Gross
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D002853:Chromatography, Liquid / Q000379:methods; D001330:Electronic Data Processing / Q000379:methods*; D017665:Hydroxyl Radical / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D055668:Photochemical Processes; D017510:Protein Folding; D011506:Proteins / Q000737:chemistry*; D020543:Proteome / Q000737:chemistry; D040901:Proteomics / Q000379:methods; D012997:Solvents / Q000737:chemistry
chemical_list: D000596:Amino Acids; D011506:Proteins; D020543:Proteome; D012997:Solvents; D017665:Hydroxyl Radical