Probing a cell-embedded megadalton protein complex by DNP-supported solid-state NMR

Kaplan, Mohammed; Cukkemane, Abhishek; Zundert, Gydo C. P. van; Narasimhan, Siddarth; Daniëls, Mark A.; Mance, Deni; Waksman, Gabriel; Bonvin, Alexandre M. J. J.; Fronzes, Rémi; Folkers, Gert E.; Baldus, Marc

doi:10.1038/nmeth.3406

Cited by 131 publications

(155 citation statements)

References 32 publications

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“…The obtained results are also of interest for their implications for variable temperature studies of membrane-bound proteins and peptides, as well as whole cells, by MAS ssNMR. This notably includes the ongoing efforts to apply low-temperature dynamic nuclear polarization (DNP) to membrane-embedded proteins and whole cells (69,70).…”

Section: Discussionmentioning

confidence: 99%

MAS 1 H NMR Probes Freezing Point Depression of Water and Liquid-Gel Phase Transitions in Liposomes

Mandal

Wel

2016

Biophysical Journal

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The lipid bilayer typical of hydrated biological membranes is characterized by a liquid-crystalline, highly dynamic state. Upon cooling or dehydration, these membranes undergo a cooperative transition to a rigidified, more-ordered, gel phase. This characteristic phase transition is of significant biological and biophysical interest, for instance in studies of freezing-tolerant organisms. Magic-angle-spinning (MAS) solid-state NMR (ssNMR) spectroscopy allows for the detection and characterization of the phase transitions over a wide temperature range. In this study we employ MAS 1 H NMR to probe the phase transitions of both solvent molecules and different hydrated phospholipids, including tetraoleoyl cardiolipin (TOCL) and several phosphatidylcholine lipid species. The employed MAS NMR sample conditions cause a previously noted substantial reduction in the freezing point of the solvent phase. The effect on the solvent is caused by confinement of the aqueous solvent in the small and densely packed MAS NMR samples. In this study we report and examine how the freezing point depression also impacts the lipid phase transition, causing a ssNMR-observed reduction in the lipids' melting temperature (T m ). The molecular underpinnings of this phenomenon are discussed and compared with previous studies of membrane-associated water phases and the impact of membrane-protective cryoprotectants.

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Section: Discussionmentioning

confidence: 99%

MAS 1 H NMR Probes Freezing Point Depression of Water and Liquid-Gel Phase Transitions in Liposomes

Mandal

Wel

2016

Biophysical Journal

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“…It has been shown that DNP can drastically increase the sensitivity in solid-state NMR (ssNMR) experiments (Ni et al 2013;Sauvee et al 2013;Song et al 2006) and it has been successfully applied to several biological systems (Becker-Baldus et al 2015;Debelouchina et al 2013;Fricke et al 2015;Gupta et al 2016;Kaplan et al 2015;Koers et al 2014;Potapov et al 2015). Unfortunately, those experiments need to be carried out at cryogenic temperatures as the enhancement factor decreases with increasing temperature (Rosay et al 2010;Sauvee et al 2013;Zagdoun et al 2013).…”

Section: Contentmentioning

confidence: 99%

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

et al. 2016

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The cryogenic temperatures at which dynamic nuclear polarization (DNP) solid-state NMR experiments need to be carried out cause line-broadening, an effect that is especially detrimental for crowded protein spectra. By increasing the magnetic field strength from 600 to 800 MHz, the resolution of DNP spectra of type III secretion needles (T3SS) could be improved by 22 %, indicating that inhomogeneous broadening is not the dominant effect that limits the resolution of T3SS needles under DNP conditions. The outstanding spectral resolution of this system under DNP conditions can be attributed to its low overall flexibility.

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“…The development of a general nuclear hyperpolarization technique at arbitrary fields would enable measurements of biomolecules and reaction dynamics that were not accessible by the present techniques while decreasing routine NMR measurement times by orders of magnitude [1]. Several approaches to dynamic nuclear polarization (DNP) processes have been demonstrated that enhance nuclear spin polarization; however, the majority are limited to specific fields [2][3][4][5], low temperatures [6,7], specific molecules [8], or require microwave irradiation of the sample [8,9]. Low temperature is particularly problematic for liquid-state biological samples, where freezing leads to a loss of spectral resolution [10].…”

Section: Introductionmentioning

confidence: 99%

All-optical hyperpolarization of electron and nuclear spins in diamond

et al. 2017

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Low thermal polarization of nuclear spins is a primary sensitivity limitation for nuclear magnetic resonance. Here we demonstrate optically pumped (microwave-free) nuclear spin polarization of 13 C and 15 N in 15 N-doped diamond.15 N polarization enhancements up to −2000 above thermal equilibrium are observed in the paramagnetic system N s 0 . Nuclear spin polarization is shown to diffuse to bulk 13 C with NMR enhancements of −200 at room temperature and −500 at 240 K, enabling a route to microwave-free high-sensitivity NMR study of biological samples in ambient conditions.

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Probing a cell-embedded megadalton protein complex by DNP-supported solid-state NMR

Cited by 131 publications

References 32 publications

MAS 1 H NMR Probes Freezing Point Depression of Water and Liquid-Gel Phase Transitions in Liposomes

MAS 1 H NMR Probes Freezing Point Depression of Water and Liquid-Gel Phase Transitions in Liposomes

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

All-optical hyperpolarization of electron and nuclear spins in diamond

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