Resolution Enhancement in Multidimensional Solid-State NMR Spectroscopy of Proteins Using Spin-State Selection

Abstract

We show that the resolution of homonuclear multidimensional solid-state NMR correlation experiments can be significantly improved using transition selection and spin-state-selective polarization transfer techniques. The selection and transfer of single states allow the removal of the J-coupling contribution from the line width in both the direct and indirect spectral dimensions. This is demonstrated with a new spin-state-selective CO−Cα correlation experiment, applied to a microcrystalline 85-residue protein. With this new sequence, all four components of the CO−Cα cross-peaks are separated into different spectra, obtained by linear combination of four recorded data sets. Line narrowing of up to 44% was obtained on the protein sample for the spin-state-selective CO−Cα spectrum compared to a standard spin-diffusion experiment. The new technique also allows an easy distinction between “direct” and “relayed” transfer cross-peaks.

Narrow carbonyl resonances in proton-diluted proteins facilitate NMR assignments in the solid-state

“…In contrast, 13 C detection in the direct dimension yields broad lines due to the evolution of homonuclear scalar couplings. Decoupling can here in principle be achieved using spin state selective experiments (Duma et al 2003;Laage et al 2009), by application of homonuclear decoupling in the direct dimension (Chevelkov et al 2005) or by processing the data using J-deconvolution (Agarwal et al 2009). These techniques, however, usually compromise the sensitivity of the experiment.…”

Solid‐State NMR Spectroscopy of a Paramagnetic Protein: Assignment and Study of Human Dimeric Oxidized CuII–ZnII Superoxide Dismutase (SOD)

“…As a result, we could detect almost all the backbone and side-chain 13 C spins, including histidine residues coordinating to the copper center, therefore extracting information as close as 5 from the metal center ( 13 C a = 98 %, 13 C b = 98 %, 13 C sidechain = 95 %; owing to spectral crowding, the assignment of the carbonyl region requires dedicated experiments, for instance a 3D NCOCA experiment, [18] or alternatively spinstate-selective [19] or homodecoupled experiments, [20] and was not attempted here).…”

NMR Spectroscopy of Paramagnetic Metalloproteins

“…A more valuable approach is the inclusion in the experiments of in-phase/antiphase selection filters (IPAP). [125][126][127] The removal of the coupling is accomplished by recording two FIDs for each increment, one for the antiphase and one for the in-phase components; each pair of FIDs is then combined to separate the two multiplet components. These are then shifted to the center of the original multiplet (by J COC a/2 Hz) and summed to obtain a singlet.…”