Wednesday, July 30, 2014

Hydration dynamics as an intrinsic ruler for refining protein structure at lipid membrane interfaces

Cheng, C.-Y., et al., Hydration dynamics as an intrinsic ruler for refining protein structure at lipid membrane interfaces. Proc. Nat. Aca. Sci. USA, 2013. 110(42): p. 16838-16843.

Knowing the topology and location of protein segments at water–membrane interfaces is critical for rationalizing their functions, but their characterization is challenging under physiological conditions. Here, we debut a unique spectroscopic approach by using the hydration dynamics gradient found across the phospholipid bilayer as an intrinsic ruler for determining the topology, immersion depth, and orientation of protein segments in lipid membranes, particularly at water–membrane interfaces. This is achieved through the site-specific quantification of translational diffusion of hydration water using an emerging tool, 1H Overhauser dynamic nuclear polarization (ODNP)-enhanced NMR relaxometry. ODNP confirms that the membrane-bound region of α-synuclein (αS), an amyloid protein known to insert an amphipathic α-helix into negatively charged phospholipid membranes, forms an extended α-helix parallel to the membrane surface. We extend the current knowledge by showing that residues 90–96 of bound αS, which is a transition segment that links the α-helix and the C terminus, adopt a larger loop than an idealized α-helix. The unstructured C terminus gradually threads through the surface hydration layers of lipid membranes, with the beginning portion residing within 5–15 Å above the phosphate level, and only the very end of C terminus surveying bulk water. Remarkably, the intrinsic hydration dynamics gradient along the bilayer normal extends to 20–30 Å above the phosphate level, as demonstrated with a peripheral membrane protein, annexin B12. ODNP offers the opportunity to reveal previously unresolvable structure and location of protein segments well above the lipid phosphate, whose structure and dynamics critically contribute to the understanding of functional versatility of membrane proteins.

Monday, July 28, 2014

Hyperpolarization without persistent radicals for in vivo real-time metabolic imaging

Eichhorn, T.R., et al., Hyperpolarization without persistent radicals for in vivo real-time metabolic imaging. Proc. Nat. Aca. Sci. USA, 2013. 110(45): p. 18064-18069.

Hyperpolarized substrates prepared via dissolution dynamic nuclear polarization have been proposed as magnetic resonance imaging (MRI) agents for cancer or cardiac failure diagnosis and therapy monitoring through the detection of metabolic impairments in vivo. The use of potentially toxic persistent radicals to hyperpolarize substrates was hitherto required. We demonstrate that by shining UV light for an hour on a frozen pure endogenous substance, namely the glucose metabolic product pyruvic acid, it is possible to generate a concentration of photo-induced radicals that is large enough to highly enhance the 13C polarization of the substance via dynamic nuclear polarization. These radicals recombine upon dissolution and a solution composed of purely endogenous products is obtained for performing in vivo metabolic hyperpolarized 13C MRI with high spatial resolution. Our method opens the way to safe and straightforward preclinical and clinical applications of hyperpolarized MRI because the filtering procedure mandatory for clinical applications and the associated pharmacological tests necessary to prevent contamination are eliminated, concurrently allowing a decrease in the delay between preparation and injection of the imaging agents for improved in vivo sensitivity.

Friday, July 25, 2014

Flux through hepatic pyruvate carboxylase and phosphoenolpyruvate carboxykinase detected by hyperpolarized 13C magnetic resonance

Merritt, M.E., et al., Flux through hepatic pyruvate carboxylase and phosphoenolpyruvate carboxykinase detected by hyperpolarized 13C magnetic resonance. Proc. Nat. Aca. Sci. USA, 2011. 108(47): p. 19084-19089.

In the heart, detection of hyperpolarized [13C]bicarbonate and 13CO2 by magnetic resonance (MR) after administration of hyperpolarized [1-13C]pyruvate is caused exclusively by oxidative decarboxylation of pyruvate via the pyruvate dehydrogenase complex (PDH). However, liver mitochondria possess alternative anabolic pathways accessible by [1-13C]pyruvate, which may allow a wider diagnostic range for hyperpolarized MR compared with other tissue. Metabolism of hyperpolarized [1-13C]pyruvate in the tricarboxylic acid (TCA) cycle was monitored in the isolated perfused liver from fed and fasted mice. Hyperpolarized [1-13C]pyruvate was rapidly converted to [1-13C]lactate, [1-13C]alanine, [1-13C]malate, [4-13C]malate, [1-13C]aspartate, [4-13C]aspartate, and [13C]bicarbonate. Livers from fasted animals had increased lactate:alanine, consistent with elevated NADH:NAD+. The appearance of asymmetrically enriched malate and aspartate indicated high rates of anaplerotic pyruvate carboxylase activity and incomplete equilibration with fumarate. Hyperpolarized [13C]bicarbonate was also detected, consistent with multiple mechanisms, including cataplerotic decarboxylation of [4-13C]oxaloacetate via phosphoenolpyruvate carboxykinase (PEPCK), forward TCA cycle flux of [4-13C]oxaloacetate to generate 13CO2 at isocitrate dehydrogenase, or decarboxylation of [1-13C]pyruvate by PDH. Isotopomer analysis of liver glutamate confirmed that anaplerosis was sevenfold greater than flux through PDH. In addition, signal from [4-13C]malate and [4-13C]aspartate was markedly blunted and signal from [13C]bicarbonate was completely abolished in livers from PEPCK KO mice, indicating that the major pathway for entry of hyperpolarized [1-13C]pyruvate into the hepatic TCA cycle is via pyruvate carboxylase, and that cataplerotic flux through PEPCK is the primary source of [13C]bicarbonate. We conclude that MR detection of hyperpolarized TCA intermediates and bicarbonate is diagnostic of pyruvate carboxylase and PEPCK flux in the liver.

Wednesday, July 23, 2014

Hyperpolarized 13C dehydroascorbate as an endogenous redox sensor for in vivo metabolic imaging

Keshari, K.R., et al., Hyperpolarized 13C dehydroascorbate as an endogenous redox sensor for in vivo metabolic imaging. Proc. Nat. Aca. Sci. USA, 2011. 108(46): p. 18606-18611.

Reduction and oxidation (redox) chemistry is involved in both normal and abnormal cellular function, in processes as diverse as circadian rhythms and neurotransmission. Intracellular redox is maintained by coupled reactions involving NADPH, glutathione (GSH), and vitamin C, as well as their corresponding oxidized counterparts. In addition to functioning as enzyme cofactors, these reducing agents have a critical role in dealing with reactive oxygen species (ROS), the toxic products of oxidative metabolism seen as culprits in aging, neurodegenerative disease, and ischemia/ reperfusion injury. Despite this strong relationship between redox and human disease, methods to interrogate a redox pair in vivo are limited. Here we report the development of [1-13C] dehydroascorbate [DHA], the oxidized form of Vitamin C, as an endogenous redox sensor for in vivo imaging using hyperpolarized 13C spectroscopy. In murine models, hyperpolarized [1-13C] DHA was rapidly converted to [1-13C] vitamin C within the liver, kidneys, and brain, as well as within tumor in a transgenic prostate cancer mouse. This result is consistent with what has been previously described for the DHA/Vitamin C redox pair, and points to a role for hyperpolarized [1-13C] DHA in characterizing the concentrations of key intracellular reducing agents, including GSH. More broadly, these findings suggest a prognostic role for this new redox sensor in determining vulnerability of both normal and abnormal tissues to ROS.

Monday, July 21, 2014

Storage of nuclear magnetization as long-lived singlet order in low magnetic field

This week I will catch up with some articles that were published in the Proceedings of the National Academy of Sciences that slipped through the crack.

Pileio, G., M. Carravetta, and M.H. Levitt, Storage of nuclear magnetization as long-lived singlet order in low magnetic field. Proc. Nat. Aca. Sci. USA, 2010. 107(40): p. 17135-17139.

Hyperpolarized nuclear states provide NMR signals enhanced by many orders of magnitude, with numerous potential applications to analytical NMR, in vivo NMR, and NMR imaging. However, the lifetime of hyperpolarized magnetization is normally limited by the relaxation time constant T1, which lies in the range of milliseconds to minutes, apart from in exceptional cases. In many cases, the lifetime of the hyperpolarized state may be enhanced by converting the magnetization into nuclear singlet order, where it is protected against many common relaxation mechanisms. However, all current methods for converting magnetization into singlet order require the use of a high-field, high-homogeneity NMR magnet, which is incompatible with most hyperpolarization procedures. We demonstrate a new method for converting magnetization into singlet order and back again. The new technique is suitable for magnetically inequivalent spin-pair systems in weak and inhomogeneous magnetic fields, and is compatible with known hyperpolarization technology. The method involves audio-frequency pulsed irradiation at the low-field nuclear Larmor frequency, employing coupling-synchronized trains of 180° pulses to induce singlet–triplet transitions. The echo trains are used as building blocks for a pulse sequence called M2S that transforms longitudinal magnetization into long-lived singlet order. The time-reverse of the pulse sequence, called S2M, converts singlet order back into longitudinal magnetization. The method is demonstrated on a solution of 15N-labeled nitrous oxide. The magnetization is stored in low magnetic field for over 30 min, even though the T1 is less than 3 min under the same conditions.

Monday, July 14, 2014

Rocky Mountain Conference on Analytical Chemistry

This week I will be at the Rocky Mountain Conference on Analytical Chemistry. The conference program looks very promising with a lot of different talks on DNP-NMR spectroscopy. So no posts for this week. If you are there too say hi to me.

Friday, July 11, 2014

Room temperature hyperpolarization of nuclear spins in bulk

Tateishi, K., et al., Room temperature hyperpolarization of nuclear spins in bulk. Proc. Nat. Aca. Sci. USA, 2014. 111(21): p. 7527-7530.

Dynamic nuclear polarization (DNP), a means of transferring spin polarization from electrons to nuclei, can enhance the nuclear spin polarization (hence the NMR sensitivity) in bulk materials at most 660 times for 1H spins, using electron spins in thermal equilibrium as polarizing agents. By using electron spins in photo-excited triplet states instead, DNP can overcome the above limit. We demonstrate a 1H spin polarization of 34%, which gives an enhancement factor of 250,000 in 0.40 T, while maintaining a bulk sample (∼0.6 mg, ∼0.7 × 0.7 × 1 mm3) containing >1019 1H spins at room temperature. Room temperature hyperpolarization achieved with DNP using photo-excited triplet electrons has potentials to be applied to a wide range of fields, including NMR spectroscopy and MRI as well as fundamental physics.

Wednesday, July 9, 2014

Spin dynamic simulations of solid effect DNP: the role of the relaxation superoperator

Karabanov, A., G. Kwiatkowski, and W. Köckenberger, Spin dynamic simulations of solid effect DNP: the role of the relaxation superoperator. Mol. Phys., 2014: p. 1-17.

Relaxation plays a crucial role in the spin dynamics of dynamic nuclear polarisation. We review here two different strategies that have recently been used to incorporate relaxation in models to predict the spin dynamics of solid effect dynamic nuclear polarisation. A detailed explanation is provided on how the Lindblad?Kossakowski form of the master equation can be used to describe relaxation in a spin system. Fluctuations of the spin interactions with the environment as a cause of relaxation are discussed and it is demonstrated how the relaxation superoperator acting in Liouville space on the density operator can be derived in the Lindblad?Kossakowski form by averaging out non-secular terms in an appropriate interaction frame. Furthermore we provide a formalism for the derivation of the relaxation superoperator starting with a choice of a basis set in Hilbert space. We show that the differences in the prediction of the nuclear polarisation dynamics that are found for certain parameter choices arise from the use of different interaction frames in the two different strategies. In addition, we provide a summary of different relaxation mechanisms that need to be considered to obtain more realistic spin dynamic simulations of solid effect dynamic nuclear polarisation.

Monday, July 7, 2014

36th Magnetic Resonance Discussion Group Meeting of the Gesellschaft Deutscher Chemiker (GDCh)

From the Ampere Magnetic Resonance List

Dear Colleagues,

The Leibniz-Institut für Molekulare Pharmakologie (FMP), the Physikalisch-Technische Bundesanstalt (PTB), the Max-Delbrück-Centrum (MDC) and the Freie Universität Berlin will be hosting the 36th Magnetic Resonance Discussion Group Meeting of the Gesellschaft Deutscher Chemiker (GDCh) from Sep 29-Oct 2, 2014. The meeting will be hold in conjunction with a DIP satellite meeting "Progress in DNP".

The program will cover a wide range of magnetic resonance topics addressing participants from both academia and industry. These topics will include but are not limited to solution and solid-state NMR spectroscopy, magnetic resonance imaging, and EPR spectroscopy. Applications cover the fields of small molecules and materials, of proteins and nucleic acids, medical imaging, polymers and pharmaceutical compounds, EPR and hyperpolarization, and computational methods.

For further details please follow this link:

Please forward this announcement to anyone who might be interested.

Hartmut Oschkinat Thomas Risse Leif Schröder

This is the AMPERE MAGNETIC RESONANCE mailing list:

NMR web database:

On the Potential of Hyperpolarized Water in Biomolecular NMR Studies

Harris, T., O. Szekely, and L. Frydman, On the Potential of Hyperpolarized Water in Biomolecular NMR Studies. The Journal of Physical Chemistry B, 2014. 118(12): p. 3281-3290.

A main obstacle arising when using ex situ hyperpolarization to increase the sensitivity of biomolecular NMR is the fast relaxation that macromolecular spins undergo upon being transferred from the polarizer to the spectrometer, where their observation takes place. To cope with this limitation, the present study explores the use of hyperpolarized water as a means to enhance the sensitivity of nuclei in biomolecules. Methods to achieve proton polarizations in excess of 5% in water transferred into the NMR spectrometer were devised, as were methods enabling this polarization to last for up to 30 s. Upon dissolving amino acids and polypeptides sited at the spectrometer into such hyperpolarized water, a substantial enhancement of certain biomolecular amide and amine proton resonances was observed. This exchange-driven 1H enhancement was further passed on to side-chain and to backbone nitrogens, owing to spontaneous one-bond Overhauser processes. 15N signal enhancements >500 over 11.7 T thermal counterparts could thus be imparted in a kinetic process that enabled multiscan signal averaging. Besides potential bioanalytical uses, this approach opens interesting possibilities in the monitoring of dynamic biomolecular processes, including solvent accessibility and exchange process.

Friday, July 4, 2014

Dynamic Nuclear Polarization NMR of Low-γ Nuclei: Structural Insights into Hydrated Yttrium-Doped BaZrO3

Blanc, F., et al., Dynamic Nuclear Polarization NMR of Low-γ Nuclei: Structural Insights into Hydrated Yttrium-Doped BaZrO3. The Journal of Physical Chemistry Letters, 2014: p. 2431-2436.

We demonstrate that solid-state NMR spectra of challenging nuclei with a low gyromagnetic ratio such as yttrium-89 can be acquired quickly with indirect dynamic nuclear polarization (DNP) methods. Proton to 89Y cross polarization (CP) magic angle spinning (MAS) spectra of Y3+ in a frozen aqueous solution were acquired in minutes using the AMUPol biradical as a polarizing agent. Subsequently, the detection of the 89Y and 1H NMR signals from technologically important hydrated yttrium-doped zirconate ceramics, in combination with DFT calculations, allows the local yttrium and proton environments present in these protonic conductors to be detected and assigned to different hydrogen-bonded environments.

Wednesday, July 2, 2014

Spectral editing through laser-flash excitation in two-dimensional photo-CIDNP MAS NMR experiments

Gupta, K.B.S.S., et al., Spectral editing through laser-flash excitation in two-dimensional photo-CIDNP MAS NMR experiments. J. Magn. Reson., 2014(0).

In solid-state photochemically induced dynamic nuclear polarization (photo-CIDNP) MAS NMR experiments, strong signal enhancement is observed from molecules forming a spin-correlated radical pair in a rigid matrix. Two-dimensional 13C-13C dipolar-assisted rotational resonance (DARR) photo-CIDNP MAS NMR experiments have been applied to obtain exact chemical shift assignments from those cofactors. Under continuous illumination, the signals are enhanced via three-spin mixing (TSM) and differential decay (DD) and their intensity corresponds to the electron spin density in pz orbitals. In multiple-13C labeled samples, spin diffusion leads to propagation of signal enhancement to all 13C spins. Under steady-state conditions, direct signal assignment is possible due to the uniform signal intensity. The original intensities, however, are inaccessible and the information of the local electron spin density is lost. Upon laser-flash illumination, the signal is enhanced via the classical radical pair mechanism (RPM). The obtained intensities are related to isotropic hyperfine interactions aiso and both enhanced absorptive and emissive lines can be observed due to differences in the sign of the local isotropic hyperfine interaction. Exploiting the mechanism of the polarization, selectivity can be increased by the novel time-resolved two-dimensional dipolar-assisted rotational resonance (DARR) MAS NMR experiment which simplifies the signal assignment compared to complex spectra of the same RCs obtained by continuous illumination. Here we present two-dimensional time-resolved photo-CIDNP MAS NMR experiments providing both directly: signal assignment and spectral editing by sign and strength of aiso. Hence, this experiment provides a direct key to the electronic structure of the correlated radical pair.