Solid-State NMR/Dynamic Nuclear Polarization of Polypeptides in Planar Supported Lipid Bilayers

Dynamic nuclear polarization has been developed to overcome the limitations of the inherently low signal intensity of NMR spectroscopy. This technique promises to be particularly useful for solid-state NMR spectroscopy where the signals are broadened over a larger frequency range and most investigations rely on recording low gamma nuclei. To extend the range of possible investigations, a triple-resonance flat-coil solid-state NMR probe is presented with microwave irradiation capacities allowing the investigation of static samples at temperatures of 100 K, including supported lipid bilayers. The probe performance allows for two-dimensional separated local field experiments with high-power Lee-Goldberg decoupling and cross-polarization under simultaneous irradiation from a gyrotron microwave generator. Efficient cooling of the sample turned out to be essential for best enhancements and line shape and necessitated the development of a dedicated cooling chamber. Furthermore, a new membrane-anchored biradical is presented, and the geometry of supported membranes was optimized not only for good membrane alignment, handling, stability, and filling factor of the coil but also for heat and microwave dissipation. Enhancement factors of 17-fold were obtained, and a two-dimensional PISEMA spectrum of a transmembrane helical peptide was obtained in less than 2 h.