title: Pulsed EPR determination of water accessibility to spin-labeled amino acid residues in LHCIIb.
abstract: Membrane proteins reside in a structured environment in which some of their residues are accessible to water, some are in contact with alkyl chains of lipid molecules, and some are buried in the protein. Water accessibility of residues may change during folding or function-related structural dynamics. Several techniques based on the combination of pulsed electron paramagnetic resonance (EPR) with site-directed spin labeling can be used to quantify such water accessibility. Accessibility parameters for different residues in major plant light-harvesting complex IIb are determined by electron spin echo envelope modulation spectroscopy in the presence of deuterated water, deuterium contrast in transversal relaxation rates, analysis of longitudinal relaxation rates, and line shape analysis of electron-spin-echo-detected EPR spectra as well as by the conventional techniques of measuring the maximum hyperfine splitting and progressive saturation in continuous-wave EPR. Systematic comparison of these parameters allows for a more detailed characterization of the environment of the spin-labeled residues. These techniques are applicable independently of protein size and require approximately 10-20 nmol of singly spin-labeled protein per sample. For a residue close to the N-terminus, in a domain unresolved in the existing x-ray structures of light-harvesting complex IIb, all methods indicate high water accessibility.
authors.affiliation: Max-Planck Institute for Polymer Research, Mainz, Germany.
authors.forename: A
authors.initials: A
authors.lastname: Volkov
authors.forename: C
authors.initials: C
authors.lastname: Dockter
authors.forename: T
authors.initials: T
authors.lastname: Bund
authors.forename: H
authors.initials: H
authors.lastname: Paulsen
authors.forename: G
authors.initials: G
authors.lastname: Jeschke
mesh_terms: D000596:Amino Acids / Q000737:chemistry* / Q000378:metabolism; D002021:Buffers; D018360:Crystallography, X-Ray; D003902:Detergents / Q000737:chemistry; D004578:Electron Spin Resonance Spectroscopy; D006801:Humans; D045342:Light-Harvesting Protein Complexes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D018532:Pisum sativum; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D012995:Solubility; D012997:Solvents / Q000737:chemistry; D013113:Spin Labels; D013696:Temperature; D014867:Water / Q000737:chemistry* / Q000378:metabolism
chemical_list: D000596:Amino Acids; D002021:Buffers; D003902:Detergents; D045342:Light-Harvesting Protein Complexes; D010940:Plant Proteins; D012997:Solvents; D013113:Spin Labels; D014867:Water