Simultaneous acquisition of 2D and 3D solid-state NMR experiments for sequential assignment of oriented membrane protein samples

Abstract

We present a new method called DAISY (Dual Acquisition orIented ssNMR spectroScopY) for the simultaneous acquisition of 2D and 3D oriented solid-state NMR experiments for membrane proteins aligned in mechanically or magnetically lipid bilayers. DAISY utilizes dual acquisition of sine and cosine dipolar or chemical shift coherences and long living 15N longitudinal polarization to obtain two multi-dimensional spectra, simultaneously. In these new experiments, the first acquisition gives the polarization inversion spin exchange at the magic angle (PISEMA) or heteronuclear correlation (HETCOR) spectra, the second acquisition gives PISEMA-mixing or HETCOR-mixing spectra, where the mixing element enables inter-residue correlations through 15N-15N homonuclear polarization transfer. The analysis of the two 2D spectra (first and second acquisitions) enables one to distinguish 15N-15N inter-residue correlations for sequential assignment of membrane proteins. DAISY can be implemented in 3D experiments that include the polarization inversion spin exchange at magic angle via I spin coherence (PISEMAI) sequence, as we show for the simultaneous acquisition of 3D PISEMAI-HETCOR and 3D PISEMAI-HETCOR-mixing experiments.

Supramolecular Organization of Apolipoprotein-A-I-Derived Peptides within Disc-like Arrangements

“…One approach is based on fast magic angle sample spinning, which averages dipolar interactions and chemical shift anisotropies (2,(7)(8)(9)(10)(11)(12). Alternatively, samples oriented relative to the magnetic field direction provide well-resolved spectra where the resonance positions are indicators of the alignment of bonds and molecules relative to the magnetic field direction (6,8,13). Uniaxial sample alignment has been achieved by preparing stacks of lipid bilayers supported on glass surfaces (14) or by incorporating proteins in bicelles or large nanodiscs, i.e., lipid bilayer systems that can spontaneously orient relative to the magnetic field direction (8,13,15,16).…”

Enhancing NMR Sensitivity of Natural‐Abundance Low‐γ Nuclei by Ultrafast Magic‐Angle‐Spinning Solid‐State NMR Spectroscopy

“…In particular, for a heterogeneous sample with a broad range of 13 C T 1 values, a short T 1z filter delay between two neighboring CP periods will not be able to recover the lost 1 H magnetization during the previous CP, while a long T 1z filter delay will result in a significant signal loss (due to 13 C T 1 relaxation). Though other recent approaches that employed multiple acquisitions to utilize the residual magnetization of low‐γ nuclei are valuable to obtain multiple 2D spectra of labeled biological solids, the number of acquisitions is quite limited by the initial signal‐contact proton‐enhanced CP . On the other hand, using the MCP approach proposed in this study could significantly broaden the benefits of such approaches under ultrafast MAS.…”

Practical aspects of the simultaneous collection of COSY and TOCSY spectra

“…One strategy refers to the residual magnetization of low‐gamma nuclei that is too weak for direct observation being transferred to high‐gamma nuclei for detection. Examples have been reported in different multidimensional magic angle spinning solid‐state NMR experiments using commercial probes and standard one receiver configuration, in liquid‐state protein NMR applications and also in multiple receiver systems . On the other hand, a set of gradient‐enhanced homonuclear and heteronuclear experiments have demonstrated the feasibility of the afterglow strategy in small‐molecule NMR spectroscopy, where the residual signal originated on the same high‐gamma nuclei can represent 15–20% of the total signal .…”