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Fast magic angle spinning (MAS) NMR spectroscopy is emerging as an essential analytical and structural biology technique. Large resolution and sensitivity enhancements observed under fast MAS conditions enable structural and dynamics analysis of challenging systems, such as large macromolecular assemblies and isotopically dilute samples, using only a fraction of material required for conventional experiments. Homonuclear dipolar-based correlation spectroscopy constitutes a centerpiece in the MAS NMR methodological toolbox, and is used essentially in every biological and organic system for deriving resonance assignments and distance restraints information necessary for structural analysis. Under fast MAS conditions (rotation frequencies above 35-40 kHz), dipolar-based techniques that yield multi-bond correlations and non-trivial distance information are ineffective and suffer from low polarization transfer efficiency. To overcome this limitation, we have developed a family of experiments, CORD-RFDR. These experiments exploit the advantages of both zero-quantum RFDR and spin-diffusion based CORD methods, and exhibit highly efficient and broadband dipolar recoupling across the entire spectrum, for both short-range and long-range correlations. We have verified the performance of the CORD-RFDR sequences experimentally on a U-(13)C,(15)N-MLF tripeptide and by numerical simulations. We demonstrate applications of 2D CORD-RFDR correlation spectroscopy in dynein light chain LC8 and HIV-1 CA tubular assemblies. In the CORD-RFDR spectra of LC8 acquired at the MAS frequency of 40 kHz, many new intra- and inter-residue correlations are detected, which were not observed with conventional dipolar recoupling sequences. At a moderate MAS frequency of 14 kHz, the CORD-RFDR experiment exhibits excellent performance as well, as demonstrated in the HIV-1 CA tubular assemblies. Taken together, the results indicate that CORD-RFDR experiment is beneficial in a broad range of conditions, including both high and moderate MAS frequencies and magnetic fields.

journal of biomolecular nmr

Combined zero-quantum and spin-diffusion mixing for efficient homonuclear correlation spectroscopy under fast MAS: broadband recoupling and detection of long-range correlations