Magic Angle Spinning NMR of Proteins: High-Frequency Dynamic Nuclear Polarization and ¹H Detection

Abstract

Magic angle spinning (MAS) NMR studies of amyloid and membrane proteins and large macromolecular complexes are an important new approach to structural biology. However, the applicability of these experiments, which are based on (13)C- and (15)N-detected spectra, would be enhanced if the sensitivity were improved. Here we discuss two advances that address this problem: high-frequency dynamic nuclear polarization (DNP) and (1)H-detected MAS techniques. DNP is a sensitivity enhancement technique that transfers the high polarization of exogenous unpaired electrons to nuclear spins via microwave irradiation of electron-nuclear transitions. DNP boosts NMR signal intensities by factors of 10(2) to 10(3), thereby overcoming NMR's inherent low sensitivity. Alternatively, it permits structural investigations at the nanomolar scale. In addition, (1)H detection is feasible primarily because of the development of MAS rotors that spin at frequencies of 40 to 60 kHz or higher and the preparation of extensively (2)H-labeled proteins.

High resolution observed in 800 MHz DNP spectra of extremely rigid type III secretion needles

“…In a previous study conducted at 600 MHz (Fricke et al 2014), this system exhibited one of the best resolutions ever achieved for a protein sample under DNP conditions (Can et al 2015;Su et al 2015). Increasing the field strength to 800 MHz and using the biradical AMUPol (Sauvee et al 2013), the observed line-widths at half height are about 22 % smaller, improving the overall resolution of the spectrum sizably.…”

Atomic Resolution Structure of Monomorphic Aβ42 Amyloid Fibrils

“…7–9 In particular, MAS NMR has provided information on backbone conformations, supramolecular organization, and registry of interstrand arrangements of amyloid fibrils. This has led to an atomic resolution structure of amyloid fibrils formed by a small peptide derived from transthyretin (TTR 105–115 ), which was determined utilizing a combination of MAS NMR spectra and cryo-electron microscopy.…”

Fast magic-angle sample spinning solid-state NMR at 60–100 kHz for natural abundance samples

“…For example, the 99% of carbon is 12 C with spin quantum number, I = 0 (NMR inactive) and 99.6% of nitrogen is quadrupolar ( 14 N) with I = 1, while only 1.1% ( 13 C) and 0.37% ( 15 N) are NMR active nuclei with I = 1/2. Therefore, there is a significant reduction of the sensitivity in ssNMR spectra of 13 There are many excellent research articles and reviews that highlight the role of fast 4 MAS for biomolecular NMR applications 21,23,25,26,27,28,29,30,31,32 .…”