title: Carr-Purcell Pulsed Electron Double Resonance with Shaped Inversion Pulses.
abstract: Pulsed electron paramagnetic resonance (EPR) spectroscopy allows the determination of distances, in the range of 1.5-8 nm, between two spin-labels attached to macromolecules containing protons. Unfortunately, for hydrophobic lipid-bound or detergent-solubilized membrane proteins, the maximum distance accessible is much lower, because of a strongly reduced coherence time of the electron spins. Here we introduce a pulse sequence, based on a Carr-Purcell decoupling scheme on the observer spin, where each π-pulse is accompanied by a shaped sech/tanh inversion pulse applied to the second spin, to overcome this limitation. This pump/probe excitation scheme efficiently recouples the dipolar interaction, allowing a substantially longer observation time window to be achieved. This increases the upper limit and accuracy of distances that can be determined in membrane protein complexes. We validated the method on a bis-nitroxide model compound and applied this technique to the trimeric betaine transporter BetP. Interprotomer distances as long as 6 nm could be reliably determined, which is impossible with the existing methods.
authors.affiliation: Institute of Physical and Theoretical Chemistry and Center for Biomolecular Magnetic Resonance, Johann Wolfgang Goethe University Frankfurt , 60323 Frankfurt, Germany.
authors.forename: Philipp E
authors.initials: PE
authors.lastname: Spindler
authors.affiliation: Max-Planck Institute for Biophysics , 60438 Frankfurt, Germany.
authors.forename: Izabela
authors.initials: I
authors.lastname: Waclawska
authors.affiliation: Institute of Physical and Theoretical Chemistry and Center for Biomolecular Magnetic Resonance, Johann Wolfgang Goethe University Frankfurt , 60323 Frankfurt, Germany.
authors.forename: Burkhard
authors.initials: B
authors.lastname: Endeward
authors.affiliation: Institute of Physical and Theoretical Chemistry and Center for Biomolecular Magnetic Resonance, Johann Wolfgang Goethe University Frankfurt , 60323 Frankfurt, Germany.
authors.forename: Jörn
authors.initials: J
authors.lastname: Plackmeyer
authors.affiliation: Max-Planck Institute for Biophysics , 60438 Frankfurt, Germany.|Institute of Biophysics and Physical Biochemistry, University of Regensburg , 93053 Regensburg, Germany.
authors.forename: Christine
authors.initials: C
authors.lastname: Ziegler
authors.affiliation: Institute of Physical and Theoretical Chemistry and Center for Biomolecular Magnetic Resonance, Johann Wolfgang Goethe University Frankfurt , 60323 Frankfurt, Germany.
authors.forename: Thomas F
authors.initials: TF
authors.lastname: Prisner
mesh_terms: D002352:Carrier Proteins / Q000737:chemistry*; D004578:Electron Spin Resonance Spectroscopy; D004583:Electrons; D050485:GABA Plasma Membrane Transport Proteins; D008958:Models, Molecular; D009589:Nitrogen Oxides / Q000737:chemistry*
chemical_list: D002352:Carrier Proteins; D050485:GABA Plasma Membrane Transport Proteins; D009589:Nitrogen Oxides; C072207:betaine plasma membrane transport proteins
keywords: DEER; EPR; PELDOR; betaine transporter; membrane protein