Dynamic Nuclear Polarization of ¹⁷O: Direct Polarization

Dynamic nuclear polarization of ¹⁷O was studied using four different polarizing agents: the biradical TOTAPOL and the monoradicals trityl and SA-BDPA, as well as a mixture of the latter two. Field profiles, DNP mechanisms, and enhancements were measured to better understand and optimize directly polarizing this low-gamma quadrupolar nucleus using both mono- and biradical polarizing agents. Enhancements were recorded at <88 K and were >100 using the trityl (OX063) radical and <10 with the other polarizing agents. The >10 000-fold savings in acquisition time enabled a series of biologically relevant small molecules to be studied with small sample sizes and the measurement of various quadrupolar parameters. The results are discussed with comparison to room temperature studies and GIPAW quantum chemical calculations. These experimental results illustrate the strength of high field DNP and the importance of radical selection for studying low-gamma nuclei

Water-Soluble Narrow-Line Radicals for Dynamic Nuclear Polarization

The synthesis of air-stable, highly water-soluble organic radicals containing a 1,3-bis­(diphenylene)-2-phenylallyl (BDPA) core is reported. A sulfonated derivative, SA-BDPA, retains the narrow electron paramagnetic resonance linewidth (<30 MHz at 5 T) of the parent BDPA in highly concentrated glycerol/water solutions (40 mM), which enables its use as polarizing agent for solid effect dynamic nuclear polarization (SE DNP). A sensitivity enhancement of 110 was obtained in high-field magic-angle-spinning (MAS) NMR experiments. The ease of synthesis and high maximum enhancements obtained with the BDPA-based radicals constitute a major advance over the trityl-type narrow-line polarization agents

Equilibration of Tyrosyl Radicals (Y₃₅₆^•, Y₇₃₁^•, Y₇₃₀^•) in the Radical Propagation Pathway of the Escherichia coli Class Ia Ribonucleotide Reductase

Escherichia coli ribonucleotide reductase is an α2β2 complex that catalyzes the conversion of nucleotides to deoxynucleotides using a diferric tyrosyl radical (Y₁₂₂^•) cofactor in β2 to initiate catalysis in α2. Each turnover requires reversible long-range proton-coupled electron transfer (PCET) over 35 Å between the two subunits by a specific pathway (Y₁₂₂^• ⇆ [W₄₈?] ⇆ Y₃₅₆ within β to Y₇₃₁ ⇆ Y₇₃₀ ⇆ C₄₃₉ within α). Previously, we reported that a β2 mutant with 3-nitrotyrosyl radical (NO₂Y^•; 1.2 radicals/β2) in place of Y₁₂₂^• in the presence of α2, CDP, and ATP catalyzes formation of 0.6 equiv of dCDP and accumulates 0.6 equiv of a new Y^• proposed to be located on Y₃₅₆ in β2. We now report three independent methods that establish that Y₃₅₆ is the predominant location (85–90%) of the radical, with the remaining 10–15% delocalized onto Y₇₃₁ and Y₇₃₀ in α2. Pulsed electron–electron double-resonance spectroscopy on samples prepared by rapid freeze quench (RFQ) methods identified three distances: 30 ± 0.4 Å (88% ± 3%) and 33 ± 0.4 and 38 ± 0.5 Å (12% ± 3%) indicative of NO₂Y₁₂₂^•–Y₃₅₆^•, NO₂Y₁₂₂^•–NO₂Y₁₂₂^•, and NO₂Y₁₂₂^•–Y_731(730)^•, respectively. Radical distribution in α2 was supported by RFQ electron paramagnetic resonance (EPR) studies using Y₇₃₁(3,5-F₂Y) or Y₇₃₀(3,5-F₂Y)-α2, which revealed F₂Y^•, studies using globally incorporated [β-²H₂]Y-α2, and analysis using parameters obtained from 140 GHz EPR spectroscopy. The amount of Y^• delocalized in α2 from these two studies varied from 6% to 15%. The studies together give the first insight into the relative redox potentials of the three transient Y^• radicals in the PCET pathway and their conformations

Template-Mediated Formation of Colloidal Two-Dimensional Tin Telluride Nanosheets and the Role of the Ligands

We report the colloidal synthesis of 2D SnTe nanosheets through precursor hot injection in a nonpolar solvent. During the reaction, an important intermediateSn-templateis formed which defines the confined growth of SnTe. This “flake-like” structure gives the first evidence for the possible 2D morphology formation prior to the anion precursor injection (TOP-Te). Additionally, we explore the role of each ligand in the reaction process. Thus, we explain the formation and morphology evolution of 2D SnTe nanostructures from a mechanism perspective as well as the role of each ligand on the molecular scale. The interplay of ligands provides the necessary conditions for the realization of stable low-dimensional SnTe nanomaterials with tunable size and shape

Dynamic Nuclear Polarization with a Water-Soluble Rigid Biradical

A new biradical polarizing agent, bTbtk-py, for dynamic nuclear polarization (DNP) experiments in aqueous media is reported. The synthesis is discussed in light of the requirements of the optimum, theoretical, biradical system. To date, the DNP NMR signal enhancement resulting from bTbtk-py is the largest of any biradical in the ideal glycerol/water solvent matrix, ε = 230. EPR and X-ray crystallography are used to characterize the molecule and suggest approaches for further optimizing the biradical distance and relative orientation

Dynamic Nuclear Polarization of Sedimented Solutes

Using the 480 kDa iron-storage protein complex, apoferritin (ApoF), as an example, we demonstrate that sizable dynamic nuclear polarization (DNP) enhancements can be obtained on sedimented protein samples. In sedimented solute DNP (SedDNP), the biradical polarizing agent is co-sedimented with the protein, but in the absence of a glass-forming agent. We observe DNP enhancement factors ε > 40 at a magnetic field of 5 T and temperatures below 90 K, indicating that the protein sediment state is “glassy” and suitable to disperse the biradical polarizing agent upon freezing. In contrast, frozen aqueous solutions of ApoF yield ε ≈ 2. Results of SedDNP are compared to those obtained from samples prepared using the traditional glass-forming agent glycerol. Collectively, these and results from previous investigations suggest that the sedimented state can be functionally described as a “microcrystalline glass” and in addition provide a new approach for preparation of samples for DNP experiments

High-Field ¹³C Dynamic Nuclear Polarization with a Radical Mixture

We report direct ¹³C dynamic nuclear polarization at 5 T under magic-angle spinning (MAS) at 82 K using a mixture of monoradicals with narrow EPR linewidths. We show the importance of optimizing both EPR linewidth and electron relaxation times by studying direct DNP of ¹³C using SA-BDPA and trityl radical, and achieve ¹³C enhancements above 600. This new approach may be best suited for dissolution DNP and for studies of ¹H depleted biological and other nonprotonated solids

Dynamic Nuclear Polarization of ¹H, ¹³C, and ⁵⁹Co in a Tris(ethylenediamine)cobalt(III) Crystalline Lattice Doped with Cr(III)

The study of inorganic crystalline materials by solid-state NMR spectroscopy is often complicated by the low sensitivity of heavy nuclei. However, these materials often contain or can be prepared with paramagnetic dopants without significantly affecting the structure of the crystalline host. Dynamic nuclear polarization (DNP) is generally capable of enhancing NMR signals by transferring the magnetization of unpaired electrons to the nuclei. Therefore, the NMR sensitivity in these paramagnetically doped crystals might be increased by DNP. In this paper we demonstrate the possibility of efficient DNP transfer in polycrystalline samples of [Co­(en)₃Cl₃]₂·NaCl·6H₂O (en = ethylenediamine, C₂H₈N₂) doped with Cr­(III) in varying concentrations between 0.1 and 3 mol %. We demonstrate that ¹H, ¹³C, and ⁵⁹Co can be polarized by irradiation of Cr­(III) with 140 GHz microwaves at a magnetic field of 5 T. We further explain our findings on the basis of electron paramagnetic resonance spectroscopy of the Cr­(III) site and analysis of its temperature-dependent zero-field splitting, as well as the dependence of the DNP enhancement factor on the external magnetic field and microwave power. This first demonstration of DNP transfer from one paramagnetic metal ion to its diamagnetic host metal ion will pave the way for future applications of DNP in paramagnetically doped materials or metalloproteins

Host–Guest Complexes as Water-Soluble High-Performance DNP Polarizing Agents

Dynamic nuclear polarization (DNP) enhances the sensitivity of solid-state NMR (SSNMR) spectroscopy by orders of magnitude and, therefore, opens possibilities for novel applications from biology to materials science. This multitude of opportunities implicates a need for high-performance polarizing agents, which integrate specific physical and chemical features tailored for various applications. Here, we demonstrate that for the biradical bTbK in complex with captisol (CAP), a β-cyclodextrin derivative, host–guest assembling offers a new and easily accessible approach for the development of new polarizing agents. In contrast to bTbK, the CAP-bTbK complex is water-soluble and shows significantly improved DNP performance compared to the commonly used DNP agent TOTAPOL. Furthermore, NMR and EPR data reveal improved electron and nuclear spin relaxation properties for bTbK within the host molecule. The numerous possibilities to functionalize host molecules will permit designing novel radical complexes targeting diverse applications

Dynamic Nuclear Polarization Study of Inhibitor Binding to the M2_18–60 Proton Transporter from Influenza A

We demonstrate the use of dynamic nuclear polarization (DNP) to elucidate ligand binding to a membrane protein using dipolar recoupling magic angle spinning (MAS) NMR. In particular, we detect drug binding in the proton transporter M2_18–60 from influenza A using recoupling experiments at room temperature and with cryogenic DNP. The results indicate that the pore binding site of rimantadine is correlated with previously reported widespread chemical shift changes, suggesting functional binding in the pore. Futhermore, the ¹⁵N-labeled ammonium of rimantadine was observed near A30 ¹³Cβ and G34 ¹³Cα, suggesting a possible hydrogen bond to A30 carbonyl. Cryogenic DNP was required to observe the weaker external binding site(s) in a ZF-TEDOR spectrum. This approach is generally applicable, particularly for weakly bound ligands, in which case the application of MAS NMR dipolar recoupling requires the low temperatures to quench dynamic exchange processes. For the fully protonated samples investigated, we observed DNP signal enhancements of ∼10 at 400 MHz using only 4–6 mM of the polarizing agent TOTAPOL. At 600 MHz and with DNP, we measured a distance between the drug and the protein to a precision of 0.2 Å