Magnetic-Alignment of Polymer Nanodiscs Probed by Solid-State NMR Spectroscopy

Abstract

The ability of amphipathic polymers to self-assemble with lipids and form nanodiscs has been a boon for the field of functional reconstitution of membrane proteins. In a field dominated by detergent micelles, a unique feature of polymer nanodiscs is their much-desired ability to align in the presence of an external magnetic field. Magnetic alignment facilitates the application of solidstate NMR spectroscopy and aids in the measurement of residual dipolar couplings (RDCs) via well-established solution NMR spectroscopy. In this study, we comprehensively investigate the magnetic-alignment properties of SMA-QA polymer based nanodiscs by using 31 P and 14 N solidstate NMR experiments under static conditions. The results reported herein demonstrate the spontaneous magnetic-alignment of large-size (≥ 20 nm diameter) SMA-QA nanodiscs (also called as macro-nanodiscs) with the lipid-bilayer-normal perpendicular to the magnetic field direction. Consequently, the orientation of macro-nanodiscs are further shown to flip their alignment axis parallel to the magnetic field direction upon the addition of a paramagnetic lanthanide salt. These results demonstrate the use of SMA-QA polymer nanodiscs for solid-state NMR applications including structural studies on membrane proteins. 14 N NMR Experiments: Nitrogen-14 NMR spectra were acquired using a Bruker 400 MHz solidstate NMR spectrometer and a 5 mm double-resonance probe operating at the 14 N resonance frequency of 28.910 MHz. 14 N NMR spectra were recorded using the quadrupole-echo pulse sequence 46 with a 90° pulse length of 8 µs and an echo-delay of 80 µs. 14 N magnetization was acquired using 25 ms acquisition time, 20,000 scans and a recycle delay of 0.9 s with no 1 H decoupling.

Synthesis, Characterization, and Nanodisc formation of Non-ionic Polymers

“…An unique property of large-size (>20 nm in diameter) nanodiscs (called as macro-nanodiscs) is their ability to align in the presence of an external magnetic field. This is a much needed property to study membrane proteins using solid-state NMR techniques; [14][15][16] and also useful for solution NMR applications via the measurement of residual dipolar couplings (RDCs) [17][18][19][20] from watersoluble molecules by utilizing nanodiscs as the alignment medium. 21 To examine the magnetic-alignment properties of the non-ionic inulin based nanodiscs, static solid-state NMR experiments were performed.…”