Exploring Sulfur Sites in Proteins via Triple-Resonance 1H-Detected 77Se NMR

NMR spectroscopy has been applied to virtually all sites within proteins and biomolecules; however, the observation of sulfur sites remains very challenging. Recent studies have examined Se-77 as a replacement for sulfur and applied Se-77 NMR in both the solution and solid states. As a spin-1/2 nuclide, Se-77 is attractive as a probe of sulfur sites, and it has a very large chemical shift range (due to a large chemical shift anisotropy), which makes it potentially very sensitive to structural and/or binding interactions as well as dynamics. Despite being a spin-1/2 nuclide, there have been rather limited studies of Se-77, and the ability to use H-1-indirect detection has been sparse. Some examples exist, but in the absence of a directly bonded, nonexchangeable H-1, these have been largely limited to smaller molecules. We develop and illustrate approaches using double-labeling of C-13 and Se-77 in proteins that enable more sensitive triple-resonance schemes via multistep coherence transfers and H-1-detection. These methods require specialized hardware and decoupling schemes, which we developed and will be discussed.

Automated summary

NMR spectroscopy has been widely used in proteins and biomolecules, but observing sulfur sites has been challenging. Recent studies have explored the use of Se-77 as a replacement for sulfur and applied Se-77 NMR in both solution and solid states. A double-labeling method using C-13 and Se-77 in proteins has been developed, enabling more sensitive triple-resonance schemes.