Browsing by Subject "DNP"
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Item Application of Dissolution Dynamic Nuclear Polarization to the Characterization of Reactions Involving Large Molecules(2013-03-13) Lee, YoungbokNuclear magnetic resonance (NMR) spectroscopy is one of the most important analytical tools for organic and biological chemistry. It provides not only detailed information on the structure of small molecules and macromolecules, but also on molecular interactions. Because of the inherent low sensitivity of NMR, a long signal averaging time or a high spin concentration is often required. A variety of methods have been explored to improve the sensitivity of NMR. Especially, large signal gains can be obtained by hyperpolarization of the nuclear spins. NMR signals of hyperpolarized samples are enhanced by several orders of magnitude. Dissolution Dynamic Nuclear Polarization (D-DNP) is a versatile technique capable of polarizing many different nuclei in the solid state, and subsequently providing a hyperpolarized liquid sample following a dissolution step. The resulting signal enhancement has made it possible to obtain detailed information in research fields as varied as metabolic imaging or enzyme catalysis. This dissertation aims to extend the applicability of D-DNP into new areas of chemistry, which involve the characterization of interactions and reactions involving large molecules. In a first project, fluorine hyperpolarization is exploited to investigate protein-ligand interactions. The enhancement of 19F signal allows for the detection of submicromolar concentrations of fluorinated ligands in the strong-, intermediate-, and weak-binding regimes. Several NMR parameters are utilized to observe ligand binding to the macromolecule, and to determine dissociation constants. In a second project, competitive binding of ligands to the same binding pocket on a protein is investigated. Here, polarization flows from a first ligand hyperpolarized on protons to the protein, and then to the second ligand. The buildup in function of time of the signals due to this relayed nuclear Overhauser effect contains structural information on the binding epitope. In a third project, the aim is to directly detect a larger molecule, a polymer, which has been synthesized starting from hyperpolarized monomers. Using DNP, single scan observation of 13C, a common nucleus with large chemical shift dispersion, is possible. Time resolved 13C NMR spectroscopy in combination with kinetic models permits the description of polymerization reaction of the living anionic polymerization of styrene. In summary, several approaches have been investigated for utilizing a large hyperpolarization initially produced on small molecules, for the benefit of characterizing properties of macromolecules. These developments extend the capabilities of D-DNP and demonstrate the potential for leading to new applications in fields as diverse as drug discovery and polymer science.Item Delayed neutrons from the neutron irradiation of ???U(Texas A&M University, 2008-10-10) Heinrich, Aaron DavidA series of experiments was performed with the Texas A&M University Nuclear Science Center Reactor (NSCR) to verify ???U delayed neutron emission rates. A custom device was created to accurately measure a sample's pneumatic flight time and the Nuclear Science Center's (NSC's) pneumatic transfer system (PTS) was redesigned to reduce a sample's pneumatic flight time from over 1,600 milliseconds to less than 450 milliseconds. Four saturation irradiations were performed at reactor powers of 100 and 200 kW for 300 seconds and one burst irradiation was performed using a $1.61 pulse producing 19.11 MW-s of energy. Experimental results agreed extremely well with those of Keepin. By comparing the first ten seconds of collected data, the first saturation irradiation deviated ~1.869% with a dead time of 2 microseconds, while the burst irradiation deviated ~0.303% with a dead time of 5 microseconds. Saturation irradiations one, three and four were normalized to the initial count rate of saturation irradiation two to determine the system reproducibility, and deviated ~0.449%, ~0.343% and ~0.389%, respectively.Item Nuclear Magnetic Resonance based Characterization of the Protein Binding Pocket using Hyperpolarized Ligand(2014-08-04) Min, HlaingIn the drug design process, the structural determination of the protein-ligand binding interface and understanding how the drug binds to the target protein at the protein binding pocket is essential. In the past few years, Dynamic Nuclear Polarization (DNP) combined with Nuclear Magnetic Resonance (NMR) has emerged as a new tool for studying interactions between different molecules. In this study, the DNP-NMR technique was employed for characterization of the protein binding pocket through binding of the hyperpolarized ligand to the protein. Trypsin and benzamidine were chosen as models for the protein and the ligand because the binding of benzamidine to trypsin is well-known. Several enhanced NMR signals of trypsin appeared from the binding of hyperpolarized benzamidine to trypsin. A significant finding was that those trypsin signals were non-uniformly enhanced when compared with the trypsin signals in the conventional (non-hyperpolarized) NMR spectrum, suggesting that a specific region of the protein, most likely the protein-binding pocket proximal to the bound hyperpolarized ligand, is selectively polarized. The polarization transfer process was described mathematically by fitting model equations to the enhanced signal intensities of both the protein and the ligand. A fit parameter was evaluated, which assuming the presence of a single spin on protein and ligand can be interpreted as a cross-relaxation rate (?_(DNP)), that can provide spatial information between the two spins. Saturation Transfer Difference (STD)-NMR was employed as an independent method to measure the protein-ligand interaction. The fit parameters in the STD-NMR equations, the dissociation constant (K_(D)) and a cross-relaxation rate (?_(STD)), were evaluated. KD determined from STD-NMR was consistent with the K_(D) values reported in the literatures, suggesting that STD-NMR data is reliable. ?_(STD) evaluated from STD-NMR was compared with ?_(DNP) evaluated from the DNP-NMR and found to be similar in magnitude.Item Quantitative Determination of Chemical Processes by Dynamic Nuclear Polarization Enhanced Nuclear Magnetic Resonance Spectroscopy(2012-07-16) Zeng, HaifengDissolution dynamic nuclear polarization (DNP) provides several orders of magnitude of NMR signal enhancement by converting the much larger electron spin polarization to nuclear spin polarization. Polarization occurs at low temperature (1.4K) and is followed by quickly dissolving the sample for room temperature NMR detection. DNP is generally applicable to almost any small molecules and can polarize various nuclei including 1H, 19F and 13C. The large signal from DNP enhancement reduces the limit of detection to micromolar or sub-micromolar concentration in a single scan. Since DNP enhancement often provides the only source for the observable signal, it enables tracking of the polarization flow. Therefore, DNP is ideal for studying chemical processes. Here, quantitative tools are developed to separate kinetics and spin relaxation, as well as to obtain structural information from these measurements. Techniques needed for analyzing DNP polarized sample are different from those used in conventional NMR because a large, yet non-renewable hyperpolarization is available. Using small flip angle pulse excitation, the hyperpolarization can still be divided into multiple scans. Based on this principle, a scheme is presented that allows reconstruction of indirect spectral dimensions similarly to conventional 2D NMR. Additionally, small flip angle pulses can be used to obtain a succession of scans separated in time. A model describing the combined effects of the evolution of a chemical process and of spin-lattice relaxation is shown. Applied to a Diels-Alder reaction, it permitted measuring kinetics along with the effects of auto- and cross-relaxation. DNP polarization of small molecules also shows significant promise for studying protein-ligand interaction. The binding of fluorinated ligands to the protease trypsin was studied through the observation of various NMR parameter changes, such as line width, signal intensity and chemical shift of the ligands. Intermolecular polarization transfer from hyperpolarized ligand to protein can further provide information about the binding pocket of the protein. As an alternative to direct observation of protein signal, a model is presented to describe a two-step intermolecular polarization transfer between competitively binding ligands mediated through the common binding pocket of the protein. The solutions of this model relate the evolution of signal intensities to the intermolecular cross relaxation rates, which depend on individual distances in the binding epitope. In summary, DNP provides incomparable sensitivity, speed and selectivity to NMR. Quantitative models such as those discussed here enable taking full advantage of these benefits for the study of chemical processes.