C 13 – C 13 dipolar recoupling under very fast magic angle spinning in solid-state nuclear magnetic resonance: Applications to distance measurements, spectral assignments, and high-throughput secondary-structure determination

A technique is presented to recouple homonuclear dipolar couplings between dilute spin pairs such as C13–C13 systems under very fast magic angle spinning (MAS) in solid-state nuclear magnetic resonance (NMR) spectroscopy. The presented technique, finite pulse rf driven recoupling (fpRFDR), restores homonuclear dipolar interactions based on constructive usage of finite pulse-width effects in a phase- and symmetry-cycled π-pulse train in which a rotor-synchronous π pulse is applied every rotation period. The restored effective dipolar interaction has the form of a zero-quantum dipolar Hamiltonian for static solids, whose symmetry in spin space is different from that obtained by conventional rf driven recoupling (RFDR) techniques. It is demonstrated that the efficiency of recoupling by fpRFDR is not strongly dependent on chemical shift differences or resonance offsets in contrast to previous recoupling methods under very fast MAS. To realize distance measurements without effects of spin relaxation, a constant-time version of fpRFDR (CT-fpRFDR) is introduced, in which the effective evolution period is varied by refocusing dipolar evolution with a rotor-synchronized solid echo while the total recoupling period is kept constant. From CT-fpRFDR experiments at a spinning speed of 30.3 kHz in a field of 17.6 T, the C13–C13 distance of 1-13CAla–1-13CGly–Gly was determined to be 3.27 Å, which agrees well with the value of 3.20 Å obtained by x-ray diffraction. Also, two-dimensional (2D) C13/C13 chemical-shift correlation NMR spectrum in a field of 9.4 T was obtained with fpRFDR for fibrils of the segmentally C13- and N15-labeled Alzheimer’s β-Amyloid fragments, Aβ16–22 (residues 16–22 taken from the 40-residue Aβ peptide) in which Leu-17 through Ala-21 are uniformly C13- and N15-labeled. Most C13 resonances for the main chain as well as for the side chains are assigned based on 2D C13/C13 chemical-shift correlation patterns specific to amino-acid types. Examination of the obtained C13 chemical shifts revealed the formation of β-strand across the entire molecule of Aβ16–22. Possibility of high throughput determination of global main-chain structures based on C13 shifts obtained from 2D C13/C13 chemical-shift correlation under very fast MAS is also discussed for uniformly/segmentally C13-labeled protein/peptide samples.