Paramagnetic doping of a 7TM membrane protein in lipid bilayers by Gd 3+-complexes for solid-state NMR spectroscopy

Ullrich, S. J., Hölper, S., & Glaubitz, C.

Journal of biomolecular NMR 58.1 (2014): 27-35.

A considerable limitation of NMR spectroscopy is its inherent low sensitivity. Approximately 90 % of the measuring time is used by the spin system to return to its Boltzmann equilibrium after excitation, which is determined by (1)H-T1 in cross-polarized solid-state NMR experiments. It has been shown that sample doping by paramagnetic relaxation agents such as Cu(2+)-EDTA accelerates this process considerably resulting in enhanced sensitivity. Here, we extend this concept to Gd(3+)-complexes. Their effect on (1)H-T1 has been assessed on the membrane protein proteorhodopsin, a 7TM light-driven proton pump. A comparison between Gd(3+)-DOTA, Gd(3+)-TTAHA, covalently attached Cu(2+)-EDTA-tags and Cu(2+)-EDTA reveals a 3.2-, 2.6-, 2.4- and 2-fold improved signal-to-noise ratio per unit time due to longitudinal paramagnetic relaxation enhancement. Furthermore, Gd(3+)-DOTA shows a remarkably high relaxivity, which is 77-times higher than that of Cu(2+)-EDTA. Therefore, an order of magnitude lower dopant concentration can be used. In addition, no line-broadening effects or peak shifts have been observed on proteorhodopsin in the presence of Gd(3+)-DOTA. These favourable properties make it very useful for solid-state NMR experiments on membrane proteins.

Paramagnetic-doping-of-a-7TM-membrane-protein-in-lipid-bilayers-by-Gd-complexes-for-solid-state-NMR-spectroscopy

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