Nitroxide side-chain dynamics in a spin-labeled helix-forming peptide revealed by high-frequency (139.5-GHz) EPR spectroscopy.
High-frequency electron paramagnetic resonance (EPR) spectroscopy has been performed on a nitroxide spin-labeled peptide in fluid aqueous solution. The peptide, which follows the single letter sequence, was reacted with the methanethiosulfonate spin label at the cysteine sulfur. The spin sensitivity of high-frequency EPR is excellent with less than 20 pmol of sample required to obtain spectra with good signal-to-noise ratios. Simulation of the temperature-dependent spectral lineshapes reveals the existence of local anisotropic motion about the nitroxide N-O bond with a motional anisotropy tau( perpendicular)/tau( parallel) ( identical with N) approaching 2.6 at 306 K. Comparison with previous work on rigidly labeled peptides suggests that the spin label is reorienting about its side-chain tether. This study demonstrates the feasibility of performing 140-GHz EPR on biological samples in fluid aqueous solution.
Bennati
M
M
Department of Chemistry and Biochemistry, University of California, Santa Cruz, California 95064, USA.
Gerfen
G J
GJ
Martinez
G V
GV
Griffin
R G
RG
Singel
D J
DJ
Millhauser
G L
GL
D000595:Amino Acid Sequence; D016880:Anisotropy; D004578:Electron Spin Resonance Spectroscopy; D010455:Peptides / Q000737:chemistry*; D011487:Protein Conformation; D013113:Spin Labels; D013696:Temperature
D010455:Peptides; D013113:Spin Labels