title: The orientation of a tandem POTRA domain pair, of the beta-barrel assembly protein BamA, determined by PELDOR spectroscopy.
abstract: The outer membrane beta-barrel trans-membrane proteins in gram-negative bacteria are folded into the membrane with the aid of polypeptide transport-associated (POTRA) domains. These domains occur, and probably function, as a tandem array situated on the periplasmic side of the outer membrane. Two crystal structures and one NMR study have attempted to define the structure and articulation of the POTRA domains of the Escherichia coli, prototypic Omp85 protein BamA. We have used pulsed electron paramagnetic resonance (EPR) to determine the distance and distance distribution between (1-Oxyl-2,2,5,5-tetramethylpyrroline-3-methyl) methanethiosulfonate spin labels (MTSSL), placed across the domain interface of the first two POTRA domains of BamA. Our results show tightly defined interdomain distance distributions that indicate a well-defined domain orientation. Examination of the known structures revealed that none of them fitted the EPR data. A combination of EPR and NMR data was used to generate converged structures with defined domain-domain orientation.
authors.affiliation: College of Life Sciences, University of Dundee, Dundee, UK.
authors.forename: R
authors.initials: R
authors.lastname: Ward
authors.forename: M
authors.initials: M
authors.lastname: Zoltner
authors.forename: L
authors.initials: L
authors.lastname: Beer
authors.forename: H
authors.initials: H
authors.lastname: El Mkami
authors.forename: I R
authors.initials: IR
authors.lastname: Henderson
authors.forename: T
authors.initials: T
authors.lastname: Palmer
authors.forename: D G
authors.initials: DG
authors.lastname: Norman
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D029968:Escherichia coli Proteins / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D011487:Protein Conformation; D013113:Spin Labels
chemical_list: D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; D013113:Spin Labels