title: Solid-state NMR study of a 41 kDa membrane protein complex DsbA/DsbB.
abstract: The disulfide bond generation system in E. coli is led by a periplasmic protein, DsbA, and an integral membrane protein, DsbB. Here we present a solid-state NMR (SSNMR) study of a 41 kDa membrane protein complex DsbA/DsbB precipitated in the presence of native lipids to investigate conformational changes and dynamics that occur upon transient complex formation within the electron transfer pathway. Chemical shift changes in the periplasmic enzyme DsbA in three states (wild type, C33S mutant, and in complex with DsbB) reveal structural and/or dynamic information. We report a 4.9 ppm (15)N chemical shift change observed for Pro31 in the active site between the wild type and C33S mutant of DsbA. Additionally, the Pro31 residue remains elusive in the DsbA/DsbB complex, indicating that the dynamics change drastically in the active site between the three states of DsbA. Using three-dimensional SSNMR spectra, partial (13)C and (15)N de novo chemical shift assignments throughout DsbA in the DsbA/DsbB complex were compared with the shifts from DsbA alone to map site-specific chemical shift perturbations. These results demonstrate that there are further structural and dynamic changes of DsbA in the native membrane observed by SSNMR, beyond the differences between the crystal structures of DsbA and the DsbA/DsbB complex.
authors.affiliation: Department of Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, Illinois 61801, United States.
authors.forename: Lindsay J
authors.initials: LJ
authors.lastname: Sperling
authors.forename: Ming
authors.initials: M
authors.lastname: Tang
authors.forename: Deborah A
authors.initials: DA
authors.lastname: Berthold
authors.forename: Anna E
authors.initials: AE
authors.lastname: Nesbitt
authors.forename: Robert B
authors.initials: RB
authors.lastname: Gennis
authors.forename: Chad M
authors.initials: CM
authors.lastname: Rienstra
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D004220:Disulfides / Q000737:chemistry; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism*; D008958:Models, Molecular; D008970:Molecular Weight; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D019704:Protein Disulfide-Isomerases / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004220:Disulfides; C079503:DsbB protein, Bacteria; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D019704:Protein Disulfide-Isomerases; C526648:dsbA protein, E coli