EPR Spectra of Spin-labeled Lysozyme in Tetragonal Crystals
Oriented in the Magnetic Field

R. I. Artyukh¹, G. S. Kachalova¹, N. F. Lanina¹, A. P. Mozoleva²,
N. V. Anisimov², and V. P. Timofeev²

¹ Institute of Theoretical and Experimental Biophysics, Russian Academy of Sciences,
Pushchino, Moscow Region, 142292 Russia
² Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, 117984 Russia

Received November 27, 1998

Abstract—Tetragonal crystals of spin-labeled lysozyme were studied. The protein was modified at the histi-
dine residue 15 (the only one in the protein structure) with spin labels containing an imidazolidine-based nitrox-
ide radical. The average number of spin-labeled histidine residues per lysozyme molecule in the crystal was
shown to be close to 0.1. The dependence of the EPR spectra on the crystal orientation with respect to the mag-
netic field was obtained and analyzed by theoretical modeling of the spectra using an original computer pro-
gram, which simulates the quasicrystal EPR spectrum of the nitroxide radical. It was demonstrated that the
directions of the Z axes of the nitroxides on lysozyme molecules in the unit cell of a tetragonal crystal are
reduced to two mutually perpendicular orientations along the crystallographic axes a and b. Without dipole–
dipole interaction between labels in a crystal, the substantial difference of the distance between outer wide
peaks in the triplet EPR spectra from the doubled principal value of the hyperfine structure tensor directly
proves that the magnetic tensors are partially averaged because of the rapid angle-confined reorientations of the
nitroxide at the His-15 end. The noticeable broadening of peaks in the experimental spectra, as compared with
narrow peaks in the spectra of the magnetically diluted radical in single crystal, immediately follows from the
validity of the predicted distribution of nitroxides among clusters that correspond to only certain types of fast
reorientations. The mobility of the label at His-15 of lysozyme in a tetragonal crystal is comparable to the label
mobility in protein solution, since His-15 residues are exposed into wide intermolecular channels of the crystal.

Key words: tetragonal lysozyme crystal, orientation in magnetic field, spin labeling, EPR spectra and their sim-
ulation, dynamics of His-15 side chain

Pleiades Publishing home page | journal home page | top

If you have any problems with this server, contact webmaster.