Abstract
Lipid-nanodiscs have been shown to be an exciting innovation as a membrane-mimicking system for studies on membrane proteins by a variety of biophysical techniques, including NMR spectroscopy. Although NMR spectroscopy is unique in enabling the atomic-resolution investigation of dynamic structures of membrane-associated molecules, it, unfortunately, suffers from intrinsically low sensitivity. The long data acquisition often used to enhance the sensitivity is not desirable for sensitive membrane proteins. Instead, paramagnetic relaxation enhancement (PRE) has been used to reduce NMR data acquisition time or to reduce the amount of sample required to acquire an NMR spectra. However, the PRE approach involves the introduction of external paramagnetic probes in the system, which can induce undesired changes in the sample and on the observed NMR spectra. For example, the addition of paramagnetic ions, as frequently used, can denature the protein via direct interaction and also through sample heating. In this study, we show how the introduction of paramagnetic tags on the outer belt of polymer-nanodiscs can be used to speed-up data acquisition by significantly reducing the spin-lattice relaxation (T1) times with minimum-to-no alteration of the spectral quality. Our results also demonstrate the feasibility of using different types of paramagnetic ions (Eu3+, Gd3+, Dy3+, Er3+, Yb3+) for NMR studies on lipid-nanodiscs. Experimental results characterizing the formation of lipid-nanodiscs by the metal-chelated polymer, and their increased tolerance toward metal ions are also reported.