Determination of the rotational correlation time and the rigid limit hyperfine coupling from the dependence of spin-label EPR spectra on microwave frequency

Abstract

A method using nitroxide radical spin labels for determining both the isotropic rotational correlation time and the “rigid limit” value of the nitrogen hyperfine coupling along the p z orbital ( A z ) is described. The method requires measurement of the same sample under unchanged conditions at both X(9.5 GHz)- and Q(35 GHz)-bands, and is valid for rotational correlation times between 10 −6 and 10 −8 sec. In this context the “rigid limit” value of A z is in the absence of rotational diffusion but with fast restricted librational motions unchanged, and its measurement becomes a useful spectroscopic observable. Extension of the method to anisotropic motion, particularly as may occur in phospholipid bilayers, is discussed.