Water accessibility in a membrane-inserting peptide comparing Overhauser DNP and pulse EPR methods

Journal Article · Sat May 21 04:00:00 EDT 2016 · Journal of Chemical Physics

DOI:https://doi.org/10.1063/1.4948988· OSTI ID:22658009

Doppelbauer, Maximilian; Garbuio, Luca; Doll, Andrin; Polyhach, Yevhen O.; Jeschke, Gunnar ^[1]

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1. Laboratory of Physical Chemistry, ETH Zurich, Vladimir-Prelog-Weg 2, CH-8093 Zurich (Switzerland)

Water accessibility is a key parameter for the understanding of the structure of biomolecules, especially membrane proteins. Several experimental techniques based on the combination of electron paramagnetic resonance (EPR) spectroscopy with site-directed spin labeling are currently available. Among those, we compare relaxation time measurements and electron spin echo envelope modulation (ESEEM) experiments using pulse EPR with Overhauser dynamic nuclear polarization (DNP) at X-band frequency and a magnetic field of 0.33 T. Overhauser DNP transfers the electron spin polarization to nuclear spins via cross-relaxation. The change in the intensity of the {sup 1}H NMR spectrum of H{sub 2}O at a Larmor frequency of 14 MHz under a continuous-wave microwave irradiation of the nitroxide spin label contains information on the water accessibility of the labeled site. As a model system for a membrane protein, we use the hydrophobic α-helical peptide WALP23 in unilamellar liposomes of DOPC. Water accessibility measurements with all techniques are conducted for eight peptides with different spin label positions and low radical concentrations (10–20 μM). Consistently in all experiments, the water accessibility appears to be very low, even for labels positioned near the end of the helix. The best profile is obtained by Overhauser DNP, which is the only technique that succeeds in discriminating neighboring positions in WALP23. Since the concentration of the spin-labeled peptides varied, we normalized the DNP parameter ϵ, being the relative change of the NMR intensity, by the electron spin concentration, which was determined from a continuous-wave EPR spectrum.

• OSTI ID: 22658009
• Journal Information: Journal of Chemical Physics, Journal Name: Journal of Chemical Physics Journal Issue: 19 Vol. 144; ISSN JCPSA6; ISSN 0021-9606
• Country of Publication: United States
• Language: English

Related Subjects

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
ABUNDANCE
DINITROPHENOL
EXPERIMENTAL DATA
MEMBRANE PROTEINS
MHZ RANGE 01-100
NMR SPECTRA
NUCLEAR MAGNETIC RESONANCE
PEPTIDES
PULSES
SPIN ORIENTATION