DSpace Repository :: Browsing by Author "Bowman, Michael K."

Browsing by Author "Bowman, Michael K."

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1,2,3-Triazole-Heme Interactions in Cytochrome P450: Functionally Competent Triazole-Water-Heme Complexes
(American Chemical Society, 2012) Conner, Kip P.; Vennam, Preethi; Woods, Caleb M.; Krzyaniak, Matthew D.; Bowman, Michael K.; Atkins, William M.; University of Washington; University of Washington Seattle; University of Alabama Tuscaloosa
In comparison to imidazole (IMZ) and 1,2,4-triazole (1,2,4-TRZ), the isosteric 1,2,3-triazole (1,2,3-TRZ) is unrepresented among cytochrome P450 (CYP) inhibitors. This is surprising because 1,2,3-TRZs are easily obtained via "click" chemistry. To understand this underrepresentation of 1,2,3-TRZs among CYP inhibitors, thermodynamic and density functional theory computational studies were performed with unsubstituted IMZ, 1,2,4-TRZ, and 1,2,3-TRZ. The results indicate that the lower affinity of 1,2,3-TRZ for the heme iron includes a large unfavorable entropy term likely originating in solvent-1,2,3-TRZ interactions; the difference is not solely due to differences in the enthalpy of heme-ligand interactions. In addition, the 1,2,3-TRZ fragment was incorporated into a well-established CYP3A4 substrate and mechanism-based inactivator, 17-alpha-ethynylestradiol (17EE), via click chemistry. This derivative, 17-click, yielded optical spectra consistent with low-spin ferric heme iron (type II) in contrast to 17EE, which yields a high-spin complex (type I). Furthermore, the rate of CYP3A4-mediated metabolism of 17-click was comparable to that of 17EE, with a different regioselectivity. Surprisingly, continuous-wave electron paramagnetic resonance (EPR) and HYSCORE EPR spectroscopy indicate that 17-click does not displace water from the sixth axial ligand position of CYP3A4 as expected for a type II ligand. We propose a binding model in which 17-click pendant 1,2,3-TRZ hydrogen bonds with the sixth axial water ligand. The results demonstrate the potential for 1,2,3-TRZ to form metabolically labile water-bridged low-spin heme complexes, consistent with recent evidence that nitrogenous type II ligands of CYPs can be efficiently metabolized. The specific case of [CYP3A4 center dot 17-click] highlights the risk of interpreting CYP-ligand complex structure on the basis of optical spectra.
Absolute Oxygen R-1e Imaging In Vivo with Pulse Electron Paramagnetic Resonance
(Wiley-Blackwell, 2014) Epel, Boris; Bowman, Michael K.; Mailer, Colin; Halpern, Howard J.; University of Chicago; University of Alabama Tuscaloosa
Purpose: Tissue oxygen (O-2) levels are among the most important and most quantifiable stimuli to which cells and tissues respond through inducible signaling pathways. Tumor O-2 levels are major determinants of the response to cancer therapy. Developing more accurate measurements and images of tissue O-2 partial pressure (pO(2)), assumes enormous practical, biological, and medical importance. Methods: We present a fundamentally new technique to image pO(2) in tumors and tissues with pulse electron paramagnetic resonance (EPR) imaging enabled by an injected, nontoxic, triaryl methyl (trityl) spin probe whose unpaired electron's slow relaxation rates report the tissue pO(2). Heretofore, virtually all in vivo EPR O-2 imaging measures pO(2) with the transverse electron spin relaxation rate, R-2e, which is susceptible to the self-relaxation confounding O-2 sensitivity. Results: We found that the trityl electron longitudinal relaxation rate, R-1e, is an order of magnitude less sensitive to confounding self-relaxation. R-1e imaging has greater accuracy and brings EPR O-2 images to an absolute pO(2) image, within uncertainties. Conclusion: R-1e imaging more accurately determines oxygenation of cancer and normal tissue in animal models than has been available. It will enable enhanced, rapid, noninvasive O-2 images for understanding oxygen biology and the relationship of oxygenation patterns to therapy outcome in living animal systems. (C) 2013 Wiley Periodicals, Inc.
Bloch equations for anisotropic paramagnetic centers with spin of 1/2
(Elsevier, 2013) Maryasov, Alexander G.; Bowman, Michael K.; Russian Academy of Sciences; Voevodsky Institute of Chemical Kinetics & Combustion SB RAS; University of Alabama Tuscaloosa
The Bloch equations are an invaluable tool in magnetic resonance for describing the dynamics of isotropic spin systems. However, when the Bloch equations are reformulated for anisotropic spin systems, much of their utility is lost because the spin evolution they describe is not physically observable. A set of Bloch-like equations are derived for these anisotropic systems in terms of the magnetic moment which is the physical property measured in magnetic resonance and other experiments. The equations describe the dynamics of the magnetic moment including relaxation and only contain parameters that are experimentally measurable. (C) 2013 Elsevier Inc. All rights reserved.
A Caged, Destabilized, Free Radical Intermediate in the Q-Cycle
(Wiley-VCH, 2013) Vennam, Preethi R.; Fisher, Nicholas; Krzyaniak, Matthew D.; Kramer, David M.; Bowman, Michael K.; University of Alabama Tuscaloosa; Michigan State University; United States Department of Energy (DOE)
The Rieske/cytochrome b complexes, also known as cytochrome bc complexes, catalyze a unique oxidant-induced reduction reaction at their quinol oxidase (Q(o)) sites, in which substrate hydroquinone reduces two distinct electron transfer chains, one through a series of high-potential electron carriers, the second through low-potential cytochrome b. This reaction is a critical step in energy storage by the Q-cycle. The semiquinone intermediate in this reaction can reduce O-2 to produce deleterious superoxide. It is yet unknown how the enzyme controls this reaction, though numerous models have been proposed. In previous work, we trapped a Q-cycle semiquinone anion intermediate, termed SQ(o), in bacterial cytochrome bc(1) by rapid freeze-quenching. In this work, we apply pulsed-EPR techniques to determine the location and properties of SQ(o) in the mitochondrial complex. In contrast to semiquinone inter-mediates in other enzymes, SQ(o) is not thermodynamically stabilized, and can even be destabilized with respect to solution. It is trapped in Q(o) at a site that is distinct from previously described inhibitor-binding sites, yet sufficiently close to cytochrome b(L) to allow rapid electron transfer. The binding site and EPR analyses show that SQ(o) is not stabilized by hydrogen bonds to proteins. The formation of SQ(o) involves "stripping" of both substrate -OH protons during the initial oxidation step, as well as conformational changes of the semiquinone and Q(o) proteins. The resulting charged radical is kinetically trapped, rather than thermodynamically stabilized (as in most enzymatic semiquinone species), conserving redox energy to drive electron transfer to cytochrome b(L) while minimizing certain Q-cycle bypass reactions, including oxidation of prereduced cytochrome b and reduction of O-2.
Characterization and catalytic applications of silver nanoparticles supported on hierarchically porous SiO_2 and Co_3O_4 monoliths
(University of Alabama Libraries, 2017) Yildirim, Yasemin; Bakker, Martin G.; University of Alabama Tuscaloosa
Hierarchically porous materials are of great interest in such applications as catalysis, drug delivery, chromatography, and electrochemical sensor arrays due to properties such as high surface area, large void volume, and tunable surface chemistry. In this study, we give the detailed report of the synthesis of supported Ag nanoparticles by nanocasting on to hierarchically porous SiO2 (silica) and Co3O4 (cobalt oxide) monoliths, as well as the use of these materials as heterogeneous catalysts for the hydrogenation and oxidation reactions. In the preliminary work, we investigated the synthesis of Ag supported on hierarchically porous SiO2 and Co3O4 monoliths, and the catalytic activity of these monoliths for the hydrogenation of MB (methylene blue) and MO (methyl orange) dyes in the presence of NaBH4 (sodium borohydride). The SiO2 monoliths were synthesized using a sol-gel technique. The Co3O4 monoliths were prepared by nanocasting, using the SiO2 monoliths as a template. The loading of Ag nanoparticles on the SiO2 and Co3O4 monoliths was done by a solution infiltration method using aqueous AgNO3 (silver nitrate) solution followed by reduction with ethylene glycol and hydrazine hydrate. Such monoliths also were used as continuous flow monolithic microreactors for the catalytic activity and stability studies for the hydrogenation of EO (eosin-Y) dye in the presence of NaBH4. Finally, the use of these monoliths for the oxidation of cyclohexene was investigated including the effects of temperature, oxidant, catalyst loading, and substrate to oxidant ratio. All these studies are presented in different sections depending on the different synthesis, procedures, and catalytic activity occurring in each heterogeneous catalyst.
Characterization and thermoelectric performance of polymer/inorganic hybrid films using spectroscopic and microscopic techniques
(University of Alabama Libraries, 2019) Sutch, Tabitha Patricia; Szulczewski, Greg J.; University of Alabama Tuscaloosa
The main goal of this dissertation was to investigate the thermoelectric performance of films of silver telluride (Ag2Te), bismuth sulfide (Bi2S3) and bismuth telluride (Bi2Te3) nanowires (NWs) dispersed in P(NDI2OD-T2). We hypothesize that the electrical properties of the films made with Ag2Te, Bi2S3, and Bi2Te3 will be n-type and behave like composite materials. Ag2Te was synthesized to make three different lengths of NWs. Powder x-ray diffraction (XRD) and energy dispersive x-ray spectroscopy (EDS) confirmed that β-Ag2Te was synthesized with the anticipated stoichiometry. Scanning electron microscopy (SEM) of Ag2Te NWs in P(NDI2OD-T2) revealed that the longest Ag2Te NWs produced homogeneous composites whereas the shorter Ag2Te NWs phase-separated. Electrical conductivity and Seebeck coefficients for each composite film were determined and theoretical models were used to investigate charge-transport behavior. Seebeck coefficients confirmed that all composites were n-type. The longest Ag2Te NWs produced the highest electrical conductivities with parallel transport behavior and are promising to the field of thermoelectrics. Bi2S3 and Bi2Te3 NWs were synthesized, and their stoichiometry and structures were confirmed using XRD and EDS. SEM images of the NW/P(NDI2OD-T2) films revealed that both Bi2S3 and Bi2Te3 phase-separated. The electrical conductivity for each composite film was determined and theoretical models were applied. The electrical conductivity of the Bi2S3 and Bi2Te3 composites were slightly higher than pristine P(NDI2OD-T2), further indicating that the NWs phase-separated. Seebeck coefficients for both systems confirmed that the composites were n-type. Despite the various strategies for improving the film morphology, composites made with Bi2Te3 and Bi2S3 NWs did not produce promising results. The spin-dynamics of P(NDI2OD-T2) doped with cobaltocene were studied with continuous-wave electron paramagnetic resonance (EPR). Additionally, the electrical conductivity of cobaltocene doped P(NDI2OD-T2) films increased several orders of magnitude compared to pristine P(NDI2OD-T2). A sputter depth profile in conjunction with x-ray photoelectron spectroscopy was used to analyze how the dopant dispersed in the polymer matrix. Temperature-dependent pulsed EPR of two different doping concentrations suggested two different relaxation rates. Overall, this study investigated the temperature-dependent spin dynamics of cobaltocene doped P(NDI2OD-T2) films and lays the foundation for further investigations on n-doped polymer system.
Characterization of the carotenoid cis-bixin
(University of Alabama Libraries, 2018) Tay-Agbozo, Sefadzi; Street, Shane C.; Bowman, Michael K.; University of Alabama Tuscaloosa
Bixin, a carotenoid found in annatto, Bixa orellana, is unique among natural carotenoids by being sparingly water-soluble. Bixin free radicals have been stabilized on the surface of silica alumina and TiO2 and characterized by pulsed electron nuclear double resonance (ENDOR). Least-square fitting of experimental ENDOR spectra calculated from density functional theory (DFT) calculations hyperfine couplings characterized the radicals trapped on silica alumina and TiO2. DFT predicts that the trans bixin radical cation is more stable than the cis bixin radical cation by 1.26 kcal/mol. While this small energy difference is consistent with the 26% trans and 23% cis radical cations in the ENDOR spectrum for silica alumina, the TiO2 spectrum could not be fitted due to poor signal. The ENDOR spectrum for silica alumina shows several neutral radicals formed by loss of a H+ ion from the 9, 9′, 13, or 13′ methyl group, a common occurrence in all water-insoluble carotenoids studied in literature. In addition, the continuous wave (CW) electron paramagnetic resonance (EPR) spectroscopy signal of bixin on silica alumina was intense prior to irradiation. Upon irradiation, the intensity is reduced 4-fold. On the other hand, unlike on TiO2 there was no signal prior to irradiation but signal was observed upon irradiation. The extinction coefficient bixin in chloroform is determined to be 1.1 x 105 ± 0.49 and 1.0 x 105 M-1cm-1 ±0.05 at 472 and 503 nm, respectively, while the redox potential in THF is found to be 0.94 ± 0.01V by cyclic voltammogram (CV) measurements. Based on the irreversibility of the CV, the bixin radical cation is estimated to have a short lifetime and decay rapidly at ambient temperature.
Chromium: binding studies with transferrin and peptide eeeegdd and its effect on colorectal cancer
(University of Alabama Libraries, 2016) Deng, Ge; Vincent, John B.; University of Alabama Tuscaloosa
Chromium as the trivalent ion has been proposed as an essential element for decades. Although that status has recently been discredited, doses of Cr3+ have been shown to generate improvements in insulin sensitivity and blood cholesterol levels in animals that have problems with their glucose and lipid metabolism systems, especially in type 2 diabetic rodent models. The mechanism for these effects at a molecular level is unknown. Transferrins are a class of protein that can reversibly bind 2 equivalents of metal ions. Biologically, transferrins are the main iron transport proteins in plasma. A role for transferrin in the delivery Cr3+ from plasma to tissues has been proposed. Studies have shown that Cr3+ readily binds to the two metal-binding sites in the two lobes of apotransferrin. The Cr3+ binding is accompanied by intense changes in the transferrin’s ultraviolet spectrum. This intense changes arises from chromic ion binding to two tyrosine residues in the two iron-binding sites of transferrin and allow the binding of Cr3+ to transferrin to be monitored. The rate at which Cr3+ binds to transferrin and the stability of Cr-transferrin recently has received considerable attention. In vitro spectroscopic studies previously found that the generation of Cr2-transferrin needs two weeks to guarantee a stoichiometric amount of Cr3+ binding. However, this study indicates that in the presence of 25 mM (bi)carbonate, the concentration in human blood, two Cr3+ ions bind rapidly and tightly to apotransferrin. Glycation of transferrin alters how tightly the protein binds iron and may alter the conformation of diferric transferrin, presumably changing its ability to deliver the iron to tissues. Given that Cr3+ complexes has been proposed as nutritional supplements to improve symptoms of type 2 diabetic subjects, understanding the ability of glycated transferrin to bind and transport Cr is significant, especially for determining the appropriate dose of Cr. This study examined the binding ability of Cr3+ to glycated serum transferrin and the transport of Cr in vivo by glycated transferrin. The results suggest that glycation of transferrin in subjects with elevated blood glucose levels should lower the ability of Cr from pharmacological agents to enter tissues. Additionally, these studies with glycated transferrin also indicate that heat treatment of transferrin makes dramatic change on its conformation and Cr binding ability.
Computational studies of Lewis acidic gas adsorption to transition metal oxide nanoclusters and metal organic frameworks
(University of Alabama Libraries, 2017) Flores, Luis Antonio; Dixon, David A.; University of Alabama Tuscaloosa
Computational studies of the interaction of Lewis acid gases with metal oxide clusters and metal organic frameworks show how these gases interact with and degrade these materials at the molecular level. The calculations were done at the levels of density functional theory and correlated molecular orbital theory ((CCSD(T))). Group VI metal oxides clusters physisorb CO_2 near or below to 298K, and chemisorption of CO_2 by carbonate formation is an endothermic process. SO_2 physisorbs to Group VI clusters near or below 298K. Group VI metal oxides chemisorb SO_2 by forming sulfites with positive free energies of binding at 298K. The formation of sulfates is thermodynamically allowed for Cr clusters because Cr clusters have a higher reducibility than do Mo or W clusters. Group IV metal oxide clusters prefer chemisorption of both gases by carbonate and sulfite formation. Mo and W oxides may function as long lived sorbents for these gases, whereas Cr and Group IV metal oxides would degrade upon exposure to these gases as sulfites, sulfates, or carbonates form on their surfaces. Uranium trioxide clusters are predicted to chemisorb CO_2 by uranyl carbonate formation. The exposure of nuclear waste to CO_2 could cause uranium oxides to degrade leading to ground water contamination. The physisorption of the Lewis acid gases (CO_2, SO_2, H_2O, H_2S, CO, and NO_2) to M-MOF-2 systems (M = Zn, Cu, Co), was investigated. The MOFs are predicted to bind H_2O, H_2S, and SO_2 more strongly than the other gases. The binding energies are larger for Zn and Co than for Cu. Zn-MOF-2 clusters will degrade faster than Cu.
Computational studies of solid state materials for practical applications
(University of Alabama Libraries, 2012) Stott, Amanda C.; Dixon, David A.; University of Alabama Tuscaloosa
The Ni-rich Ni55Ti45 composition of the NiTi alloy is a promising material for aerospace bearing materials. Spiral orbit tribometry friction tests performed on Ni-rich Ni55Ti45 titanium ball bearings indicate that this alloy is a promising candidate for future aerospace bearing applications. Microstructural characterization of the bearing specimens was performed using transmission electron microscopy and energy dispersive spectroscopy, with NiTi, Ni4Ti3, Ni3Ti, and Ni2Ti4Ox phases identified within the microstructure of the alloy. Density functional theory (DFT) was applied to predict the electronic structure of the NixTiy phases, including the band structure and site projected density of states. Ultraviolet photoemission spectroscopy was used to verify the density of states results from the density functional theory calculations, with good agreement observed between experiment and theory. Plane wave ab initio DFT calculations of the B2 NiTi (100), (110), and (111) surfaces, the B2 and B19´ phases of NiTi, and the supercell structures of NiTi, Ni4Ti3 and Ni3Ti are also reported. Electronic energies from the electronic structure calculations are used to assess relative stability of the different surface and supercell geometries. DFT was applied using a plane wave approach for solids to determine the band gap energies in a series of Pb3C6X6 semiconducting extended-network organic structures, to determine the phase stability in the NiTi alloy system, and to study the surface of the (bcc) B2 phase of NiTi. To reveal the molecular structure and optoelectronic properties of these materials, a detailed ab-initio theoretical investigation of the solid-state properties was performed. Density functional theory was applied to predict the electronic structure of the NixTiy phases, including the band structure and site projected density of states. Organo-metallic compounds are also an important class of materials for organic electronic devices due to their semiconducting properties. Ground state geometries, band structure, density of states, and charge density were calculated using density functional theory using the PBE exchange-correlation functional as well as the Heyd-Scuseria-Ernzerhof (HSE06) hybrid functional. The results show that the optical properties and band gap energies can be easily tuned by chemical modifications of the substituent X atom in Pb3(C6X6). Calculations of substituent atoms S, O, Se, and Te are presented. TiV-based alloys are promising candidates for NiMH batteries, as it is well established that the V-based (bcc) solid solution phase acts as the major hydrogen absorbing phase wguke the (hcp) Ti phase acts mainly as a catalyst for electrochemical hydrogenation and dehydrogenation. To achieve improved electrochemical performance a precise knowledge of the microstructure is required. Therefore, the influence of the interfacial energy on the material phase stability was investigated for a series of TiV multi-laminate thin films. Experiments revealed that at a higher layer thickness, the α (hcp) phase is the most stable. As the layer thickness is reduced, a transformation from the α (hcp) phase to the ß (bcc) phase occurs. Atomic-scale characterization of the transformed specimen by atom probe tomography reveals V interfacial diffusion between the layers. Equivalent crystal theory based calculations confirm the V interfacial diffusion mechanism. The predicted segregation profiles match those obtained experimentally.
Computational studies of the fundamental thermodynamic properties of amino acids and small peptides
(University of Alabama Libraries, 2015) Stover, Michele Leigh; Dixon, David A.; University of Alabama Tuscaloosa
In 2011, the Human Proteome Project (HPP) was launched with the main goal of experimentally mapping the entire human proteome. Determining a protein’s sequence is a first step to understanding its structure and function, which are of great importance in biological, biochemical, and medical studies. The sequencing of peptides and proteins by mass spectrometry (MS) has become a major tool in proteomics research because it is a cost effective and highly reproducible analytical technique. The analysis of biomolecules by mass spectrometry requires an understanding of proton transfer reactions because the two most commonly used ionization techniques, electrospray ionization (ESI) and matrix-assisted laser desorption ionization (MALDI), involve the addition and removal of protons. The sites of proton transfer reactions can affect the fragmentation patterns of peptide ions, which consequently impacts the sequence information that can be obtained from mass spectrometry experiments. Thermodynamic values, such as the gas-phase acidity (GA or ∆Gacid), the ΔG for the deprotonation reaction AH → A- + H+, provide valuable information to help in understanding the less studied negative ion peptide fragmentation mode by mass spectrometry. The study of gas-phase proton transfer reactions provides unique insights into the structures and energetics of the peptides. Changing the protonation state can impact the hydrogen bonding in the molecule and result in decreased or increased properties such as solubility, hydrophobicity, and electrostatic interactions. By combining these experimentally obtained results with the results from high level electronic structure theory calculations, an improved understanding of the structures and energetics of polypeptides can be obtained. Herein we describe computational studies using reliable, correlated molecular orbital methods of the gas-phase properties, including acidities and heats of formation, and solution-phase acidities of amino acids (AAs) and small peptides including substituted molecules such as AA amides and phosphorylated AAs. This dissertation will focus on the prediction of fundamental thermodynamic properties of AAs and small peptides that are of interest in the area of biochemistry, proteomics, and the Human Proteome Project.
The computational studies on the chemistry of titanium dioxide nanoparticles
(University of Alabama Libraries, 2010) Wang, Tsang-Hsiu; Dixon, David A.; University of Alabama Tuscaloosa
The chemistry of TiO_2 and SiO_2 nanoclusters is studied using computational methods. The potential energy surfaces (PESs), thermochemistry of the intermediates, and the reaction paths for the initial steps of the hydrolysis of TiCl_4 were calculated. Transition state theory and RRKM unimolecular rate theory are used to predict the rate constants. Clustering energies and heats of formation are calculated for neutral clusters, and the calculated heats of formation were used to study condensation reactions. The reaction energy is substantially endothermic if more than 2 HCl are eliminated. The calculations show that the reported values for ΔH_f^0(TiOCl_2) should be remeasured. Transition metal oxides such as TiO_2 can be used as photocatalysts to control chemical transformations for energy production. An important applications for TiO_2 is its use to photochemically split water to produce H_2 and O_2. The PES for splitting water on the ground and first excited state surfaces of (TiO_2)_n (n=1-4) nanoparticles have been studied up through the coupled cluster CCSD(T)/complete basis set level. Water is readily split to form hydroxyl groups without the need for a photon. Experimental measurements of the photoconversion of ketones (C(O)RR') on the rutile TiO_2 (110) surface show that one can eliminate R or R'. The bond dissociation energies of R=CH_3 and a wide range of R' for the gem-diols CRR'(OH)_2 were calculated at the density functional theory (DFT) and G3(MP2) levels. The calculated bond dissociation energies are in excellent agreement with the experimental values. The calculations show that most of the photodissociation processes are under thermodynamic control except for R'=CF_3. X-ray photoelectron spectroscopy (XPS) and DFT electronic structure calculations were used to study the average formal oxidation state of silicon in fumed silica (CAB-O-SIL®). The results show that the average surface oxidation state of the silicon in fumed silica is predominantly +1 and suggest a notably less hydrophilic character for CAB-O-SIL® than the oxides of silicon with Si in the formal +3 and +4 oxidation states. Once the +3 oxidation state is formed, water on the silica surface facilitates the conversion of the Si^+3 to the Si^+4 oxidation state.
Computational Thermodynamics of Gas and Solution Phase Anions
(University of Alabama Libraries, 2020) McNeill, Ashley Shari; Dixon, David A.; University of Alabama Tuscaloosa
The work in this dissertation focuses on the computational analysis of the thermodynamics of anions in the gas phase and in aqueous solution to provide unique insights into the chemistry of a range of biologically and geochemically relevant chemical species. This often involves calculating properties for these species such as electron affinities and hydration free energies of the anions, which can be difficult or impossible to obtain experimentally. Systems of interest in this work include small peptides, enzyme-catalyzed biological reactions, and the gas phase and solvation energetics of a variety of anionic species including CO2-, H-, X- (halides), OX- (hypohalites), and YH- (chalcogen hydrides). The peptide work, performed largely with the composite correlated molecular orbital theory G3(MP2) method, is compared directly to experiments conducted with low-energy collision-induced dissociation negative ion mode mass spectrometry. Isotope fractionation studies, of significant use in many geochemical applications, are conducted on the overall reaction by the alanine transaminase enzyme (+H3NCH(CH3)COO? + ?OOCCH2CH2C(O)COO? ? CH3C(O)COO? + +H3NCH(CH2CH2COO?)COO?) in order to predict that 13C preferentially collects in the C2 site of pyruvate over alanine by 9‰ at equilibrium. This prediction, calculated from gas phase- and aqueous-optimized clusters with explicit H2O molecules at the MP2/aug-cc-pVDZ with and without the COSMO self-consistent reaction field for implicit solvation, is reflected in simpler models: without explicit solvation, with simpler analogues formaldehyde and methylamine, and from canonical functional group frequencies and reduced masses for R2C=O and R2CH-NH2. Solvation studies of the CO2-, H-, X-, OX-, and YH- anions and corresponding neutrals gave adiabatic electron affinities, reduction potentials, and gas phase and aqueous acidities that are generally in excellent agreement with experiment. These studies used a variety of computational methods, including heavy application of coupled cluster calculations with the Feller-Peterson-Dixon method to obtain high accuracy thermodynamic values. Absolute hydration free energies are determined for neutral and anionic species clustered with 4 to 8 explicit H2O molecules using a supermolecule-continuum approach.
CW EPR parameters reveal cytochrome P450 ligand binding modes
(Elsevier, 2018) Lockart, Molly M.; Rodriguez, Carlo A.; Atkins, William M.; Bowman, Michael K.; University of Alabama Tuscaloosa; University of Washington; University of Washington Seattle
Cytochrome P450 (CYP) monoxygenses utilize heme cofactors to catalyze oxidation reactions. They play a critical role in metabolism of many classes of drugs, are an attractive target for drug development, and mediate several prominent drug interactions. Many substrates and inhibitors alter the spin state of the ferric heme by displacing the heme's axial water ligand in the resting enzyme to yield a five-coordinate iron complex, or they replace the axial water to yield a nitrogen-ligated six-coordinate iron complex, which are traditionally assigned by UV-vis spectroscopy. However, crystal structures and recent pulsed electron paramagnetic resonance (EPR) studies find a few cases where molecules hydrogen bond to the axial water. The water-bridged drug-H2O-heme has UV-vis spectra similar to nitrogen-ligated, six-coordinate complexes, but are closer to "reverse type I" complexes described in older liteature. Here, pulsed and continuous wave (CW) EPR demonstrate that water-bridged complexes are remarkably common among a range of nitrogenous drugs or drug fragments that bind to CYP3A4 or CYP2C9. Principal component analysis reveals a distinct clustering of CW EPR spectral parameters for water-bridged complexes. CW EPR reveals heterogeneous mixtures of ligated states, including multiple directly-coordinated complexes and water-bridged complexes. These results suggest that water-bridged complexes are under-represented in CYP structural databases and can have energies similar to other ligation modes. The data indicates that water-bridged binding modes can be identified and distinguished from directly-coordinated binding by CW EPR.
Development and applications in computational chemistry for inorganic catalysis
(University of Alabama Libraries, 2013) Chen, Mingyang; Dixon, David A.; University of Alabama Tuscaloosa
A robust metadata database called the Collaborative Chemistry Database Tool (CCDBT) for massive amounts of computational chemistry raw data has been designed and implemented. It performs data synchronization and simultaneously extracts the meta data. The indexed meta data can be used for data analysis and data mining. A novel tree growth - hybrid genetic algorithm (TG-HGA) was developed to search the global minimum of small clusters. In the TG algorithm, the clusters grow from a small seed to the size of interest stepwise. New atoms are added to the smaller cluster from the previous step, by analogy to new leaves grown by a tree. The initial structures for the search for the global minimum of TiO_2 nanoclusters were generated by TG-HGA, and new low energy structures that have not been previously reported were found. Low energy isomers of Agn, n = 2 - 99, were studied at different computational level depending on the size of Agn. The geometries of Agn, n = 2 - 8, were optimized using density functional theory (DFT), and the energies were calculated at the CCSD(T)/CBS level. The Agn, n = 9 - 20, were initially generated by the TG-HGA builder with an EAM potential, and optimized using the DFT method. The relative energies and normalized atomization energies for the optimized structures were calculated at the CCSD(T) level with a small basis set. For larger Agn, 20 < n < 100, the low energy structures were generated using TG-HGA with an EAM potential, and the energies were calculated at the DFT level with a small basis set. A range of DFT functionals were benchmarked with the normalized atomization energies at the CCSD(T) level for the small Agn clusters. PW91 and ω-B97XD provided best results for predicting the normalized atomization energies. The normalized atomization energies for Agn start to converge slowly to the bulk at n = 55. At n = 99, the normalized atomization energy is predicted to be ~50 kcal/mol. The low energy isomers of the Irn(CO)m complexes (n=1, 2, 3, 4, and 6) were investigated using electronic structure methods at the density functional theory and coupled cluster (CCSD(T) theory levels. Ir4(CO)12 is predicted to be the most favored complex for reactions of Irn(CO)m with CO at low temperature, and Ir6(CO)16 is predicted to be formed above room temperature. Smaller Irn(CO)m clusters will nucleate to form Ir4(CO)12 spontaneously. Low-lying structures of the small iridum clusters Irn (n = 2 - 8) were optimized using DFT methods. Ir2 and Ir3 were also optimized using the CASSCF method. MRCI-SD (for Ir2) energies and CCSD(T) (for Ir2 and Ir3) energies of the leading configurations from the CASSCF calculations were done to predict the low-lying states. The normalized atomization energies for Irn (n = 2 - 8) were calculated at the CCSD(T) level up to the complete basis set (CBS) limit in some cases using the B3LYP optimized geometries. Inclusion of the spin orbit corrections in the normalized atomization energies for Irn is critical and will decrease the normalized atomization energies by ~ 15 kcal/mol for n ≥ 4. Several molecular models were used to characterize various binding sites of the metal complexes in the zeolites. The calculated structures and energies indicate a metal-oxygen (M(I)-O) coordination number of two for most of the supported complexes but a value of three when the ligands include C2H5 or H. The results characterizing various isomers of supported metal complexes incorporating hydrocarbon ligands indicate that some carbene and carbyne ligands could form. A set of ligand bond dissociation energies is reported to explain reactivity trends. The Pd-L ligand bond dissociation energies (BDEs) of cis- and trans-[L-Pd(PH3)2Cl]+ were predicted using coupled cluster CCSD(T) theory and a variety of density functional theory (DFT) functionals at the B3LYP optimized geometries. For cis-[L-Pd (PH3)2Cl]+ complexes, the Pd-L bond energies are 28 kcal/mol for CO; ~40 kcal/mol for AH3 (A = N, P, As, and Sb), norbornene, and CH3CN; and ~53 kcal/mol for CH3NC, pyrazole, pyridine, and tetrahydrothiophene at the CCSD(T) level. The benchmarks show that the dispersion-corrected hybrid, generalized gradient approximation, DFT functional ω-B97X-D is the best functional to use for this system. Use of the ω-B97X-D/aD functional gives predicted BDEs within 1 kcal/mol of the CCSD(T)/aug-cc-pVTZ BDEs for cis-[L-Pd(PH3)2Cl]+ and 1.5 kcal/mol for trans-[L-Pd(PH3)2Cl]+ . Lanthanide metal atoms, produced by laser ablation, were condensed with CH3F in excess Ar at 8 K. New infrared absorption bands are assigned to the first insertion CH3LnF and oxidative addition methylene lanthanide hydride fluoride CH2LnHF products on the basis of 13C and deuterium substitution and density functional theory calculations of the vibrational frequencies. For Ln = Eu and Yb only CH3LnF is observed. CH3LnF in the Ln formal +2 state is predicted to be more stable than CH2LnHF with the Ln in the formal +3 oxidation state. CH3-LnF forms a single bond between Ln and C and is a substituted methane. The calculated potential energy surface for the CH3F + La → CH3-LaF/CH2-LaHF shows a number of intermediates and transition states on multiple paths. The reaction mechanism involves the potential formation of LaF and LaHF intermediates.
Drug binding characterization of CYPs utilizing CW and pulsed EPR
(University of Alabama Libraries, 2015) Cuce, Alex Andrew; Bowman, Michael K.; University of Alabama Tuscaloosa
Electron paramagnetic resonance (EPR) methods have been used to study drug and ligand interactions with a super family of monooxygenase enzymes known as Cytochrome P450 (CYP). We examined the active-site of four different isoforms of CYPs before and after the addition of drug using EPR. Two of the CYPs studied, CYP3A4 and CYP2C9, play a major role in drug metabolism and the other two, CYP51B1 and CYP125A1, are attractive as therapeutic targets for the pathogen Mycobacterium tuberculosis. EPR has shown to be a quick and highly resolved method, in comparison to current methods such as crystal structure analysis and UV/Vis optical difference spectroscopy, to study CYPs ferric heme active-site before and after drugs bind, which could be very valuable in drug design. Chapter 2 examines pulsed EPR methods for studying the active-site in CYPs. The explanation of practical aspects of experimentation along with data processing provides the EPR background for studying ferric heme-containing enzymes enabling a researcher to extract highly resolved active-site information. The experimental EPR methods described in Chapter 2 are the methods used in Chapter 3 and 4. Chapter 3 examines the resting state active-site of all four isoforms which is structurally described as a single water molecule bound to the distal position of the ferric heme. CW EPR spectroscopy of the isoforms all gave different g-values and MCD showed that water ligands bind at different strengths depending on the CYP isoform which sheds light on substrate specificity in each isoform. An attempt was made at predicting nIR MCD transitions with the EPR parameters but results were unclear. Chapter 4 studied CYP2C9 and CYP125A1 in complex with drugs that had the same binding mode but different optical difference spectra. We showed that the low-spin complex between a drug-metabolizing CYP2C9 variant in complex with a drug PPT retains the water ligand seen in the resting state. Hydrogens from the axial water ligand are observed by pulsed EPR spectroscopy for both drug-free and drug-bound species showing that the drug does not displace the water ligand seen in the resting state. An 15N-label incorporated into PPT is .444 nm from the heme iron indicating that PPT is in the active-site. CYP125A1 gave the same EPR signatures seen for CYP2C9 and PPT along with an X-ray crystal structure of CYP125A1 in complex with LP10 showing a water-bridged complex. The same binding mode was seen in both complexes but optical difference spectra of CYP2C9 and PPT resemble ‘classic’ type II behavior while those of CYP125A1 and LP10 have reverse type I behavior, again providing direct evidence that optical difference spectra are not reliable for characterization of drug binding mode.
Drug Modulation of Water-Heme Interactions in Low-Spin P450 Complexes of CYP2C9d and CYP125A1
(American Chemical Society, 2015) Conner, Kip P.; Cruce, Alex A.; Krzyaniak, Matthew D.; Schimpf, Alina M.; Frank, Daniel J.; de Montellano, Paul Ortiz; Atkins, William M.; Bowman, Michael K.; University of Washington; University of Washington Seattle; University of Alabama Tuscaloosa; University of California San Francisco
Azoles and pyridines are commonly incorporated into small molecule inhibitor scaffolds that target cytochromes P450 (CYPs) as a strategy to increase drug binding affinity, impart isoform-dependent selectivity, and improve metabolic stability. Optical absorbance spectra of the CYP-inhibitor complex are widely used to infer whether these inhibitors are ligated directly to the heme iron as catalytically inert, low-spin (type II) complexes. Here, we show that the low-spin complex between a drug-metabolizing CYP2C9 variant and 4-(3-phenylpropyl)-1H-1,2,3-triazole (PPT) retains an axial water ligand despite exhibiting elements of classic type II optical behavior. Hydrogens of the axial water ligand are observed by pulsed electron paramagnetic resonance (EPR) spectroscopy for both inhibitor-free and inhibitor-bound species and show that inhibitor binding does not displace the axial water. A N-15 label incorporated into PPT is 0.444 nm from the heme iron, showing that PPT is also in the active site. The reverse type I inhibitor, LP10, of CYP125A1 from Mycobacterium tuberculosis, known from X-ray crystal structures to form a low-spin water-bridged complex, is found by EPR and by visible and near-infrared magnetic circular dichroism spectroscopy to retain the axial water ligand in the complex in solution.
Electron paramagnetic resonance investigation of semiquinone intermediate in mitochondrial cytochrome bc1 complex
(University of Alabama Libraries, 2012) Vennam, Preethi Reddy; Bowman, Michael K.; University of Alabama Tuscaloosa
The Rieske/cytochrome b complexes, also known as cytochrome bc complexes, catalyze a unique oxidant-induced reduction reaction at their quinol oxidase (Qo) sites, in which substrate hydroquinone reduces two distinct electron transfer chains, one leading to a series of high-potential electron carriers, the second to low-potential cytochrome b. This reaction is a critical step in energy storage by the Q-cycle. The semiquinone intermediate in this reaction can also reduce O2 to produce deleterious superoxide. It is yet unknown how the enzyme controls this reaction, though numerous models have been proposed. In previous work we were able to trap a Q-cycle semiquinone anion intermediate, termed SQo in bacterial cyt bc1 by rapid freeze-quenching. In this work, we apply pulsed EPR techniques to determine the location of SQo in mictochondrial complex and that mitochondrial SQo has highly unusual properties. In contrast with previous semiquinone intermediates, SQo is not thermodynamically stabilized, or even destabilized with respect to solution. It is localized in the Qo pocket at a niche, which is distinct from previously described inhibitor-binding sites, but is sufficiently close to cytochrome bL to allow rapid electron transfer. Both the location of the binding sites and EPR analysis show that SQo is not stabilized by hydrogen bonds to proteins. These results indicate that the formation of SQo involves "stripping" of both substrate protons during the initial oxidation to the high potential chain, as well as conformational changes of both semiquinone species and Qo site proteins components. The resulting charged radical is kinetically trapped, rather than thermodynamically stabilized (as in most enzymatic semiquinone species), maintaining redox energy to drive electron transfer to cytochrome bL, while minimizing certain Q-cycle bypass reactions including oxidation of pre-reduced cytochrome b and reduction of O2.
Electron paramagnetic resonance studies of drug binding in cytochrome P450 enzymes
(University of Alabama Libraries, 2019) Lockart, Molly Marie; Bowman, Michael K.; University of Alabama Tuscaloosa
Cytochrome P450 enzymes (CYPs) are heme-containing monooxygenase enzymes that exist in nearly every living organism. They are responsible for oxidizing a wide variety of substrates in biosynthetic and detoxification pathways and are a common target for drug design. CYP-drug binding modes are traditionally characterized using optical difference spectra, but these binding assignments oftentimes miss atypical CYP-drug binding and do not account for mixtures of binding modes. Electron paramagnetic resonance (EPR) spectroscopy can shed light on CYP-drug binding. A mixture of continuous wave (CW) EPR and pulsed EPR methods provides some valuable insight into how common drugs and drug fragments interact with the active site heme. First, the two human isoforms that contribute significantly to drug metabolism, CYP3A4 and CYP2C9, are studied in complex with a variety of drugs. The results demonstrate that CW EPR parameters can reveal drug binding modes. Remarkably, this research finds an abundance of water-bridged complexes, and many of them coexist in frozen solution with complexes where the drug directly coordinates to the heme. These mixtures of binding modes have significant ramifications for drug design. Additionally, CYP3A4 is studied in the context of drug-drug interactions, looking at how common drugs, like acetaminophen, caffeine, midazolam, and carbamazepine can simultaneously occupy the active site when combined. These results find that multiple drugs occupy the active site and that they are distinct from any CYP-single drug complex. A method is established for observing human CYP drug-drug interactions with EPR, and it provides evidence of simultaneous drug binding with common drugs. In addition to human CYPs, this research examines drug binding in CYP51B1, a Mycobacterium tuberculosis CYP isoform. The results find that inhibitor-like compounds form mixtures of bound complexes, including some that retain the axial water. Overall, this research provides several new details about CYP-drug interactions. These results and observations highlight the importance of understanding and characterizing CYP-drug binding with more detailed analyses that provide information on the full range of CYP binding modes.
Electron spin dynamics and spin-lattice relaxation of trityl radicals in frozen solutions
(Royal Society of Chemistry, 2016) Chen, Hanjiao; Maryasov, Alexander G.; Rogozhnikova, Olga Yu.; Trukhin, Dmitry V.; Tormyshev, Victor M.; Bowman, Michael K.; University of Alabama Tuscaloosa; Voevodsky Institute of Chemical Kinetics & Combustion SB RAS; Russian Academy of Sciences; Vorozhtsov Novosibirsk Institute of Organic Chemistry; Novosibirsk State University
Electron spin-lattice relaxation of two trityl radicals, d(24)-OX063 and Finland trityl, were studied under conditions relevant to their use in dissolution dynamic nuclear polarization (DNP). The dependence of relaxation kinetics on temperature up to 100 K and on concentration up to 60 mM was obtained at X-and W-bands (0.35 and 3.5 Tesla, respectively). The relaxation is quite similar at both bands and for both trityl radicals. At concentrations typical for DNP, relaxation is mediated by excitation transfer and spin-diffusion to fast-relaxing centers identified as triads of trityl radicals that spontaneously form in the frozen samples. These centers relax by an Orbach-Aminov mechanism and determine the relaxation, saturation and electron spin dynamics during DNP.

Browsing by Author "Bowman, Michael K."

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