Measuring distances in proteins by saturation-recovery EPR.
We describe a protocol for detecting electron spin-spin interactions between a radical and a metal ion in a protein or protein complex by saturation-recovery electron paramagnetic resonance (EPR). This protocol can be used with a protein containing an endogenous metal center and either an endogenous or synthetic radical species. We suggest a two-step approach whereby dipole-dipole or exchange interactions are first detected by continuous-wave EPR experiments and then quantified by saturation-recovery EPR. The latter measurements make it possible to measure long distances to within a few Angstroms. The protocol for making distance measurements by saturation-recovery EPR will take approximately 6 days to complete.
Hirsh
Donald J
DJ
Department of Chemistry, The College of New Jersey, PO Box 7718, Ewing, New Jersey 08628, USA. hirsh@tcnj.edu
Brudvig
Gary W
GW
D004574:Electromagnetic Fields; D004578:Electron Spin Resonance Spectroscopy / Q000295:instrumentation / Q000379:methods*; D005609:Free Radicals; D008872:Microwaves; D045332:Photosystem II Protein Complex / Q000737:chemistry; D011506:Proteins / Q000737:chemistry* / Q000302:isolation & purification; D011789:Quantum Theory; D015203:Reproducibility of Results; D014443:Tyrosine
D005609:Free Radicals; D045332:Photosystem II Protein Complex; D011506:Proteins; D014443:Tyrosine