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PNAS Plus: Conformational state of the MscS mechanosensitive channel in solution revealed by pulsed electron-electron double resonance (PELDOR) spectroscopy

DOI: 10.1073/pnas.1202286109 DOI Help
PMID: 23012406 PMID Help

Authors: C. Pliotas (Biomedical Sciences Research Complex, University of St Andrews) , R. Ward (Biomedical Sciences Research Complex, University of St Andrews) , E. Branigan (Biomedical Sciences Research Complex, University of St Andrews) , A. Rasmussen (Institute of Medical Sciences, University of Aberdeen) , G. Hagelueken (Biomedical Sciences Research Complex, University of St Andrews) , H. Huang (Biomedical Sciences Research Complex, University of St Andrews) , S. S. Black (Institute of Medical Sciences, University of Aberdeen) , I. R. Booth (Institute of Medical Sciences, University of Aberdeen) , O. Schiemann (Biomedical Sciences Research Complex, University of St Andrews) , J. H. Naismith (Biomedical Sciences Research Complex, University of St Andrews)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Proceedings Of The National Academy Of Sciences

State: Published (Approved)
Published: September 2012
Diamond Proposal Number(s): 7705

Abstract: The heptameric mechanosensitive channel of small conductance (MscS) provides a critical function in Escherichia coli where it opens in response to increased bilayer tension. Three approaches have defined different closed and open structures of the channel, resulting in mutually incompatible models of gating. We have attached spin labels to cysteine mutants on key secondary structural elements specifically chosen to discriminate between the competing models. The resulting pulsed electron–electron double resonance (PELDOR) spectra matched predicted distance distributions for the open crystal structure of MscS. The fit for the predictions by structural models of MscS derived by other techniques was not convincing. The assignment of MscS as open in detergent by PELDOR was unexpected but is supported by two crystal structures of spin-labeled MscS. PELDOR is therefore shown to be a powerful experimental tool to interrogate the conformation of transmembrane regions of integral membrane proteins.

Journal Keywords: Western; Chromatography; Gel; Crystallography; Electron; Escherichia; Ion; Models; Molecular; Mutagenesis; Patch-Clamp; Protein; Sequence; DNA; Spectrum; Spin Labels

Subject Areas: Biology and Bio-materials


Instruments: I04-1-Macromolecular Crystallography (fixed wavelength)

Other Facilities: ESRF