Determining α-helical and β-sheet secondary structures via pulsed electron spin resonance spectroscopy.

A new method has been developed to determine α-helical and β-sheet secondary structural components of aqueous and membrane-bound proteins using pulsed electron paramagnetic resonance (EPR) spectroscopy. The three-pulse electron spin echo envelope modulation (ESEEM) technique was used to detect weakly coupled (2)H-labeled nuclei on side chains in the proximity of a strategically placed nitroxide spin-label up to 8 Å away. Changes in the ESEEM spectra for different samples correlate directly to periodic structural differences between α-helical and β-sheet motifs. These distinct trends were demonstrated with α-helical (M2δ subunit of the acetylcholine receptor) and β-sheet (ubiquitin) peptides in biologically relevant sample environments.