| Abstract: | A method of monitoring slow rotational motions of proteins from the decay of the intrinsic phosphorescence is described. The phosphorescence is excited with a 10-μsec pulse of vertically polarized light from an air gap lamp, and the anisotropy was computed as a function of time from the simultaneously detected vertically and horizontally polarized components of the emission. The approach is illustrated with time-dependent measurements of the anisotropy of the tryptophan phosphorescence of Staphylococcus aureus nuclease, bovine carbonic anhydrase, and liver alcohol dehydrogenase in glycerol-phosphate buffer between ?90 and ?70°C. The temperature- and molecular-weight dependence of the exponential decays in the anisotropy indicate that overall rotation of the proteins is at the origin of the depolarization. The potential of the approach as a probe of the slow rotational motions of proteins in membranes and other macromolecular complexes is stressed. |
