Interpretation of the electron spin resonance spectra of nitroxide-maleimide-labelled proteins and the use of this technique in the study of albumin and biomembranes.
The composite ESR spectra of nitroxide-maleimide spin-labelled albumin and mitochondrial membranes were investigated. It was found that the mobile spectrum (so called "weakly immobilized") is due to the probe free in solution; it could be removed by dialysis, by lowering the ratio of spin label to albumin or by using acid pH values. Similar degrees of mobility were noted with spectra of spin label in solutions with viscosities similar to that of albumin. The immobile spectrum (so called "strongly immobilized") is due to the binding of N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidinyl)-maleimide to the proteins. Studies with modified albumins have shown that in this binding the free SH and amino groups are involved, but the three-dimensional structure offering conditions for hydrophobic binding is also required. The spectral parameters useful for evaluating conformational changes of proteins and an application to mitochondrial membranes are described.
D000818:Animals; D001665:Binding Sites; D003497:Cyclic N-Oxides; D004578:Electron Spin Resonance Spectroscopy; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008301:Maleimides; D008566:Membranes / Q000648:ultrastructure; D008930:Mitochondria, Liver / Q000648:ultrastructure*; D011485:Protein Binding; D011487:Protein Conformation; D051381:Rats; D012710:Serum Albumin, Bovine; D013113:Spin Labels; D013696:Temperature; D014783:Viscosity
D003497:Cyclic N-Oxides; D008301:Maleimides; D013113:Spin Labels; D012710:Serum Albumin, Bovine