Proton hyperpolarisation preserved in long-lived states.

International audienceThe polarisation of abundant protons, rather than dilute nuclei with low gyromagnetic ratios, can be enhanced in less than 10 min using dissolution DNP and converted into a long-lived state delocalised over an ensemble of three coupled protons. The process is more straightforward than the hyperpolarisation of heteronuclei followed by magnetisation transfer to protons

SAA fibrils involved in AA amyloidosis are similar in bulk and by single particle reconstitution: a MAS solid-state NMR study

AA amyloidosis is one of the most prevalent forms of systemic amyloidosis and affects both humans and other vertebrates. In this study, we compare MAS solid-state NMR data with a recent cryo-EM study of fibrils involving full-length murine SAA1.1. We address the question whether the specific requirements for the reconstitution of an amyloid fibril structure by cryo-EM can potentially yield a bias towards a particular fibril polymorph. We employ fibril seeds extracted from in to vivo material to imprint the fibril structure onto the biochemically produced protein. Sequential assignments yield the secondary structure elements in the fibril state. Long-range DARR and PAR experiments confirm largely the topology observed in the ex-vivo cryo-EM study. We find that the β-sheets identified in the NMR experiments are similar to the β-sheets found in the cryo-EM study, with the exception of amino acids 33–42. These residues cannot be assigned by solid-state NMR, while they adopt a stable β-sheet in the cryo-EM structure. We suggest that the differences between MAS solid-state NMR and cryo-EM data are a consequence of a second conformer involving residues 33–42. Moreover, we were able to characterize the dynamic C-terminal tail of SAA in the fibril state. The C-terminus is flexible, remains detached from the fibrils, and does not affect the SAA fibril structure as confirmed further by molecular dynamics simulations. As the C-terminus can potentially interact with other cellular components, binding to cellular targets can affect its accessibility for protease digestion

Long lived states in high field NMR spectroscopy for the study of very slow dynamic processes in solution state

In solution-state NMR spectroscopy, dynamic processes can be probed provided the nuclear relaxation rate constant does not exceed the rate constant of the process. Phenomena occurring on a time scale longer than the spin-lattice relaxation time constant, T1, were not amenable to study by conventional NMR. The discovery of long-lived nuclear spin states (LLS) in 2004, by M. Levitt, M. Carravetta and O. W. Johannessen, showed that lifetimes of magnetization can be much higher than T1. LLS experiments can be performed either in zero or in high magnetic fields for scalar-coupled proton spin pairs. This thesis contributes methods for exciting and preserving LLS in high field for a wide class of molecules, including sugars, amino acids and nucleotides. Slow diffusion has been measured for a mixture of molecules having different NMR parameters using LLS. Conformational exchange could also be followed by exchange spectroscopy in BPTI. Recently, LLS have been observed in highly mobile parts (Gly-75 and 76) of Ubiquitin. Dynamic Nuclear Polarization (DNP) is one of the methods to overcome the inherent low sensitivity of NMR spectroscopy. We have conceived an experiment to preserve DNP enhanced magnetization by conversion into LLS. A way of improving resolution and sensitivity of NMR has been designed by creation of long-lived coherences (LLC) in biomolecules

Method for nuclear magnetic resonance (NMR) spectroscopy measurements using long-lived coherences (LLC)

A method for nuclear magnetic resonance (NMR) spectroscopy of a sample comprises preparation of the sample and carrying out an NMR spectroscopy measurement. Preparation includes excitation of long lived coherences (LLC) between the singlet state S0 and the central triplet state T0 of nuclei of the sample. The thermal equilibrium Boltzmann distribution (Iz+Sz) is thereby transformed into a difference (Iz-Sz), which is flipped to the transverse plane, and irradiation of the sample with an rf-field is initiated. The LLC is sustained by maintaining the rf-irradiation during an interval t1 and the LLC is converted into observable magnetisation by interrupting the rf-irradiation. The method allows nuclear magnetic resonance spectroscopy measurements with improved spectral resolution

Long-lived coherences for homogeneous line narrowing in spectroscopy.

International audienceLine broadening, which can arise from inhomogeneities or homogeneous relaxation effects that lead to finite lifetimes of quantum states, is the Achilles' heel of many forms of spectroscopy. We show that line broadening may be considerably reduced by exploiting long lifetimes associated with superpositions of quantum states with different symmetry, termed long-lived coherences. In proton NMR of arbitrary molecules (including proteins) in isotropic solution, the slow oscillatory decays of long-lived coherences can yield spectra with very high resolution. This improvement opens the way to high-field magnetic resonance of molecular assemblies that are almost an order of magnitude larger than could be hitherto studied. Coherences between states of different symmetry may be useful in other forms of spectroscopy to cancel unwanted line broadening effects

Methods for nuclear magnetic resonance (NMR) or magnetic resonance imaging (MRI) measurements using long-lived states

A method for nuclear magnetic resonance (NMR) or magnetic resonance imaging (MRI) measurements, includes creation of enhanced polarization of nuclei of a first kind within a sample in a magnetic field at cryogenic temperatures and transfer of the polarized sample to room temperature. The enhanced polarization of nuclei of the first kind is thereby transformed into long-lived states (LLS) of nuclei of a second kind and these LLS are sustained. The LLS is at least partially converted into observable magnetization and an NMR or MRI measurement is carried out. The method allows one to extend the time needed between hyperpolarized magnetization and NMR detection

Long-lived States in Multiple-Spin Systems

Long-lived spin states are excited in molecules featuring more than two isolated coupled spins, including amino acids. The figure shows the exponential recovery with the longest time-constant in aspartic acid, T1max=5.842±0.004 s, and of the decay of the long-lived state, TLLS=10.9±0.2 s). An improvement in spin memory by a factor 2 compared to longitudinal spin-lattice relaxation time constants is obtained for most systems

Measurement of slow diffusion coefficients of molecules with arbitrary scalar couplings via long-lived spin states.

International audienceNew experiments are described for the determination of very slow diffusion constants by nuclear magnetic resonance (NMR) using long-lived (singlet) states. These experiments are suitable for molecules or conformations featuring a wide range of J-couplings