Spin-Label EPR for Determining Polarity and Proticity in Biomolecular Assemblies: Transmembrane Profiles

Abstract

Hyperfine couplings and g-values of nitroxyl spin labels are sensitive to polarity and hydrogen bonding in the environment probed. The dependences of these electronic paramagnetic resonance (EPR) properties on environmental dielectric permittivity and proticity are reviewed. Calibrations are given, in terms of the Block-Walker reaction field and local proton donor concentration, for the nitroxides that are commonly used in spin labeling of lipids and proteins. Applications to studies of the transverse polarity profiles in lipid bilayers, which constitute the permeability barrier of biological membranes, are reviewed. Emphasis is given to parallels with the permeation profiles of oxygen and nitric oxide that are determined from spin-label relaxation enhancements by using nonlinear continuouswave EPR and saturation recovery EPR, and with permeation profiles of D 2 O that are determined by using 2 H electron spin echo envelope modulation spectroscopy.

Water Penetration Profile at the Protein-Lipid Interface in Na,K-ATPase Membranes

“…The 14 N-hyperfine splittings in the conventional CW EPR spectra are sensitive to environmental polarity, including the presence of nonbonded and hydrogen-bonded water (2,36). Fig.…”

“…(1,2)). A variety of electron paramagnetic resonance (EPR) parameters are available to characterize the environmental polarity.…”

Determination of the 14N quadrupole coupling constant of nitroxide spin probes by W-band ELDOR-detected NMR

“…The method they used to determine the nqi was electronspin echo envelope modulation (ESEEM) carried out at W-band (95 GHz, $3.5 T). At this magnetic field, the A xx and A yy values of nitroxides are around [13][14][15][16][17][18][19][20] MHz and comparable to the 14 N Larmor frequency, m I $ 10 MHz. This yields intense enough nuclear modulations that can provide the 14 N frequencies along the x and y principal axes of A, which to a good approximation coincide with those of g [18].…”

“…In addition, the sensitivity of the 14 N hyperfine coupling (A zz or a iso ) to the polarity of the probe environment has been successfully used to determine polarity profiles of membranes [5,6], to monitor water penetration depth in a variety of heterogeneous systems such as membranes and micelles [7][8][9][10][11], and to understand the conformation of proteins [12]. The possibility to resolve the g xx component of the g-tensor by high field EPR has also allowed separating the polarity and proticity of the local environment [13,14]. While it is known that in apolar solvents A zz is very sensitive to the dielectric constant e, in polar solvent (e > 25) A zz is mostly dependent on the proticity (H-bonding capability), there is no efficient method to differentiate between polarity and proticity just by the hyperfine interaction [15].…”

Investigation of Liquid‐Phase Inhomogeneity on the Nanometer Scale Using Spin‐Polarized Paramagnetic Probes

“…Examples include an increase in the microwave frequency [8,9] up to 215-240 GHz (very high frequency EPR, VHF/EPR), application of pulsed EPR spectrometers [9,10], as well as advances in targeted spin labeling (site-directed spin labeling, SDSL [11][12][13]). New chemical synthesis methods have resulted in novel probe structures designed, for example, to measure the polarity of the microenvironment surrounding the probe [14], the concentration of oxygen [15], and other properties in both homogeneous and heterogeneous systems.…”