Jing-Long Du

Electron-electron spin-spin interaction in spin-labeled low-spin methemoglobin

Nitroxyl free radical electron spin relaxation times for spin-labeled low-spin methemoglobins were measured between 6 and 120 K by two-pulse electron spin echo spectroscopy and by saturation recovery electron paramagnetic resonance (EPR). Spin-lattice relaxation times for cyano-methemoglobin and imidazole-methemoglobin were measured between 8 and 25 K by saturation recovery and between 4.2 and 20 K by electron spin echo. At low temperature the iron electron spin relaxation rates are slow relative to the iron-nitroxyl electron-electron spin-spin splitting. As temperature is increased, the relaxation rates for the Fe(III) become comparable to and then greater than the spin-spin splitting, which collapses the splitting in the continuous wave EPR spectra and causes an increase and then a decrease in the nitroxyl electron spin echo decay rate. Throughout the temperature range examined, interaction with the Fe(III) increases the spin lattice relaxation rate (1/T1) for the nitroxyl. The measured relaxation times for the Fe(III) were used to analyze the temperature-dependent changes in the spin echo decays and in the saturation recovery (T1) data for the interacting nitroxyl and to determine the interspin distance, r. The values of r for three spin-labeled methemoglobins were between 15 and 15.5 A, with good agreement between values obtained by electron spin echo and saturation recovery. Analysis of the nitroxyl spin echo and saturation recovery data also provides values of the iron relaxation rates at temperatures where the iron relaxation rates are too fast to measure directly by saturation recovery or electron spin echo spectroscopy. These results demonstrate the power of using time-domain EPR measurements to probe the distance between a slowly relaxing spin and a relatively rapidly relaxing metal in a protein.

Interspin distances determined by time domain EPR of spin-labeled high-spin methemoglobin

Continuous wave (CW) EPR spectra of the nitroxyl signal in high-spin fluoro- and aquo-methemoglobin spin-labeled at the β-93 cysteines exhibited line broadening as the temperature was reduced from 20 to 5 K. In the temperature interval where the CW spectra exhibited broadening, electron spin echo data showed a dramatic increase in the phase memory relaxation rate, 1/Tm, for the nitroxyl. The lineshape changes and changes in 1/Tm are attributed to iron—nitroxyl interaction in the regime where the iron relaxation rate in s−1 is of the same order of magnitude as the iron—nitroxyl spin—spin splitting expressed in s−1. Saturation recovery data for the nitroxyl signal in spin-labeled fluoro- and aquo-methemoglobin between 15 and 150 K demonstrated that interaction with the high-spin iron(III) enhanced the electron spin relaxation rate of the spin label. Electron spin relaxation rates for the ms = ± 1/2 transitions of the high-spin iron(III) were determined by electron spin echo and inversion recovery at 4.2–6 K and by analysis of the temperature-dependent contribution to the iron CW linewidths between about 20 and 150 K. The saturation recovery data for the nitroxyl in the spin-labeled methemoglobins were analyzed to determine the interspin distances, using a modified version of the Bloembergen equation and experimentally determined iron relaxation rates. Interspin distances in the high-spin methemoglobins (15.8–17.5 A) are systematically longer than in the low-spin analogs (15–15.5 A).

Effect of molecular motion on electron spin phase memory times for copper(II) complexes in doped solids

Electron spin phase memory times,Tm, as a function of temperature were measured for Cu(II) bis(diethyldithiocarbamate), Cu(Et2dtc)2; Cu(II) bis(diethyldithiophosphate), Cu(Et2dtp)2; Cu(II) bis(diphenyldithiophosphate), Cu(Ph2dtp)2; Cu(II) tetratolylporphyrin, CuTTP; vanadyl 5-(4-carboxyphenyl)-10,15,20-tritolylporphyrin, VOTTP-COOH; and Ag(II) tetratolylporphyrin, AgTTP, doped into powdered samples of closely-related diamagnetic hosts. For the three metalloporphyrins, the electron spin relaxation rate (1/Tm) increased monotonically with increasing temperature. However the temperature dependence of the relaxation rate was not monotonic for the three other Cu(II) complexes. For Cu(Et2dtc)2 and Cu(Et2dtp)2 the temperature dependence of 1/Tm between about 85 and 130 K is attributed to the effects of methyl rotation, with activation energies of 1.0 kcal/mole. Between 120 and 250 K the 1/Tm data for Cu(Ph2dtp)2 exhibit effects that are attributed to motion of the phenyl rings.

Multifrequency electron paramagnetic resonance of irradiated L-alanine.

The radical generated by gamma-irradiation of crystalline L-alanine was examined by continuous wave (CW) and pulsed electron paramagnetic resonance (EPR) at 1.8, 3.2, 4.9, 9.1 and 19.4 GHz. The spin-flip satellite lines that make a prominent contribution to the saturated spectra at 9.1 GHz are less conspicuous at lower frequencies because of overlap with the allowed transitions. The spin-lattice relaxation times measured by long-pulse saturation recovery and phase memory times measured by electron spin echo increase with increasing microwave frequency.

Electron spin relaxation rates for nitridochromium(V) tetratolylporphyrin and nitridochromium(V) octaethylporphyrin in Frozen solution

T1 andTm for nitridochromium(V) tetratolylporphyrin and nitridochromium(V) octaethylporphyrin were measured by saturation recovery and electron spin echo EPR, respectively, between 10 and 130 K. The temperature dependence of 1/T1 was similar to that observed previously for chromium(V) complexes of hydroxycarboxylic acids. The spin lattice relaxation rate was faster in the perpendicular plane (the porphyrin plane) than normal to this plane. 1/Tm was orientation dependent with the fastest rates observed for orientations intermediate between the principal axes. The orientation dependence of 1/Tm increased with increasing temperature and decreasing rigidity of the matrix, and is attributed to molecular motion.

Enhancement of electron spin relaxation rates of metalloporphyrins due to interaction with a faster relaxing metal bound to an appended bipyridyl

Abstract Complexes of 5,10,15-tri-p-tolyl 20-4-(4-methyl) bipyridyl) porphyrin were prepared with Zn(II), Cu(II) or vanadyl coordinated to the porphyrin and bis(hexafluoroacetylacetonato)copper(II), Cu(hfac)2, or bis(ethylxanthato)nickel(II), Ni(ex)2, bound to the bipyridyl. Continuous wave EPR, electron spin echo and saturation recovery measurements were performed to determine the effect of metal-metal interaction on the EPR lineshapes between 6 K and room temperature and on the electron spin relaxation rates between about 10 and 120 K. Metal-metal exchange was larger for the Cu(II) porphyrins than for the vanadyl porphyrins. For the Cu(hfac)2 adducts the spin-spin interaction caused the relaxation rate for the slower relaxing Cu(II) or vanadyl in the porphyrin to become approximately equal to that for the faster relaxing Cu(hfac)2 moiety. The zero-field splitting for the Ni(II) complex was too large to permit detection of Ni(II) EPR signals at X-band, but the effect of the Ni(II) on the slower relaxing Cu(II) or vanadyl provided information on the relaxation rates for the Ni(II). Although the electron spin relaxation rates for the Ni(II) were orders of magnitude faster than for the Cu(II) in the Cu(hfac)2 moiety, this did not result in proportionately larger effects of the Ni(II) on the relaxation rates for the slower relaxing centers because of the large separation between the Ni(II) and Cu(II) or vanadyl transition energies.