Spin-label CW microwave power saturation and rapid passage with triangular non-adiabatic rapid sweep (NARS) and adiabatic rapid passage (ARP) EPR spectroscopy.
Non-adiabatic rapid passage (NARS) electron paramagnetic resonance (EPR) spectroscopy was introduced by Kittell et al. (2011) as a general purpose technique to collect the pure absorption response. The technique has been used to improve sensitivity relative to sinusoidal magnetic field modulation, increase the range of inter-spin distances that can be measured under near physiological conditions (Kittell et al., 2012), and enhance spectral resolution in copper (II) spectra (Hyde et al., 2013). In the present work, the method is extended to CW microwave power saturation of spin-labeled T4 Lysozyme (T4L). As in the cited papers, rapid triangular sweep of the polarizing magnetic field was superimposed on slow sweep across the spectrum. Adiabatic rapid passage (ARP) effects were encountered in samples undergoing very slow rotational diffusion as the triangular magnetic field sweep rate was increased. The paper reports results of variation of experimental parameters at the interface of adiabatic and non-adiabatic rapid sweep conditions. Comparison of the forward (up) and reverse (down) triangular sweeps is shown to be a good indicator of the presence of rapid passage effects. Spectral turning points can be distinguished from spectral regions between turning points in two ways: differential microwave power saturation and differential passage effects. Oxygen accessibility data are shown under NARS conditions that appear similar to conventional field modulation data. However, the sensitivity is much higher, permitting, in principle, experiments at substantially lower protein concentrations. Spectral displays were obtained that appear sensitive to rotational diffusion in the range of rotational correlation times of 10(-3) to 10(-7) s in a manner that is analogous to saturation transfer spectroscopy.
National Biomedical EPR Center, Department of Biophysics, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI 53226, USA.
Aaron W
AW
Kittell
National Biomedical EPR Center, Department of Biophysics, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI 53226, USA. Electronic address: jshyde@mcw.edu.
James S
JS
Hyde
D000465:Algorithms; D004578:Electron Spin Resonance Spectroscopy / Q000379:methods*; D008872:Microwaves; D011506:Proteins / Q000737:chemistry* / Q000648:ultrastructure*; D012815:Signal Processing, Computer-Assisted
D011506:Proteins
Adiabatic rapid passage; EPR; NARS; Nitroxide; Oxygen accessibility; Paramagnetic ion accessibility; Power saturation; Saturation transfer; Spin-label