Conventional and saturation transfer (electron spin resonance) techniques are used to study the motional properties of several spin labels introduced in three proteins: lysozyme, sperm whale myoglobin and human hemoglobin. The mobilities of a maleimide spin label which binds covalently to the proteins, as well as of two small probes TEMPO and PD-TEMPOL were monitored in the temperature range from –−10 to −150 °C for samples in the dry and solution states. The three proteins show a similar temperature dependence as indicated by the parameters 2Azz and ΔH. A small linear increase in 2Azz with decrease in temperature is observed for the dry samples. For the proteins in solution, on the other hand, the 2Azztemperature dependence shows a change of behaviour around −60 °C that is related to the freezing of the water molecules in the hydration shell. The changes observed for the parameter ΔH are such that at temperatures below −60 °C ΔH is greater for the solution sample, while at temperatures above −60 °C ΔH is greater for the dry sample. Saturation transfer measurements show that the motion of the spin label is very restricted in all systems (τc > 10−5s) in the temperature range studied, so that the residual librational motion of the label is sensitive to the hydration, being responsible for the observed changes of the esr parameters with temperature.
Spin label saturation transfer ESR studies of protein-lipid interactions in Photosystem II-enriched membranes