Matching Items (70)
Description
Fluoroquinolone antibiotics have been known to cause severe, multisystem adverse side effects, termed fluoroquinolone toxicity (FQT). This toxicity syndrome can present with adverse effects that vary from individual to individual, including effects on the musculoskeletal and nervous systems, among others. The mechanism behind FQT in mammals is not known, although various possibilities have been investigated. Among the hypothesized FQT mechanisms, those that could potentially explain multisystem toxicity include off-target mammalian topoisomerase interactions, increased production of reactive oxygen species, oxidative stress, and oxidative damage, as well as metal chelating properties of FQs. This review presents relevant information on fluoroquinolone antibiotics and FQT and explores the mechanisms that have been proposed. A fluoroquinolone-induced increase in reactive oxygen species and subsequent oxidative stress and damage presents the strongest evidence to explain this multisystem toxicity syndrome. Understanding the mechanism of FQT in mammals is important to aid in the prevention and treatment of this condition.
ContributorsHall, Brooke Ashlyn (Author) / Redding, Kevin (Thesis director) / Wideman, Jeremy (Committee member) / Borges, Chad (Committee member) / School of Molecular Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2021-05
Description
Rhodoferax antarcticus strain ANT.BR, a purple nonsulfur bacterium isolated from a microbial mat in Ross Island, Antarctica, is the first described anoxygenic phototrophic bacterium that is adapted to cold habitats and is the first beta-proteobacterium to undergo complete genome sequencing. R. antarcticus has unique absorption spectra and there are no obvious intracytoplasmic membranes in cells grown phototrophically, even under low light intensity. Analysis of the finished genome sequence reveals a single chromosome (3,809,266 bp) and a large plasmid (198,615 bp) that together harbor 4,262 putative genes. The genome contains two types of Rubiscos, Form IAq and Form II, which are known to exhibit quite different kinetic properties in other bacteria. The presence of multiple Rubisco forms could give R. antarcticus high metabolic flexibility in diverse environments. Annotation of the complete genome sequence along with previous experimental results predict the presence of structural genes for three types of light-harvesting (LH) complexes, LH I (B875), LH II (B800/850), and LH III (B800/820). There is evidence that expression of genes for the LH II complex might be inhibited when R. antarcticus is under low temperature and/or low light intensity. These interesting condition-dependent light-harvesting apparatuses and the control of their expression are very valuable for the further understanding of photosynthesis in cold environments. Finally, R. antarcticus exhibits a highly motile lifestyle. The genome content and organization of all putative polar flagella genes are characterized and discussed.
ContributorsZhao, Tingting, M.S (Author) / Touchman, Jeffrey (Thesis advisor) / Rosenberg, Michael (Committee member) / Redding, Kevin (Committee member) / Stout, Valerie (Committee member) / Arizona State University (Publisher)
Created2011
Description
There is a critical need for the development of clean and efficient energy sources. Hydrogen is being explored as a viable alternative to fuels in current use, many of which have limited availability and detrimental byproducts. Biological photo-production of H2 could provide a potential energy source directly manufactured from water and sunlight. As a part of the photosynthetic electron transport chain (PETC) of the green algae Chlamydomonas reinhardtii, water is split via Photosystem II (PSII) and the electrons flow through a series of electron transfer cofactors in cytochrome b6f, plastocyanin and Photosystem I (PSI). The terminal electron acceptor of PSI is ferredoxin, from which electrons may be used to reduce NADP+ for metabolic purposes. Concomitant production of a H+ gradient allows production of energy for the cell. Under certain conditions and using the endogenous hydrogenase, excess protons and electrons from ferredoxin may be converted to molecular hydrogen. In this work it is demonstrated both that certain mutations near the quinone electron transfer cofactor in PSI can speed up electron transfer through the PETC, and also that a native [FeFe]-hydrogenase can be expressed in the C. reinhardtii chloroplast. Taken together, these research findings form the foundation for the design of a PSI-hydrogenase fusion for the direct and continuous photo-production of hydrogen in vivo.
ContributorsReifschneider, Kiera (Author) / Redding, Kevin (Thesis advisor) / Fromme, Petra (Committee member) / Jones, Anne (Committee member) / Arizona State University (Publisher)
Created2013
Description
Gold-silver alloy nanoparticles (NPs) capped with adenosine 5'-triphosphate were synthesized by borohydride reduction of dilute aqueous metal precursors. High-resolution transmission electron microscopy showed the as-synthesized particles to be spherical with average diameters ~4 nm. Optical properties were measured by UV-Visible spectroscopy (UV-Vis), and the formation of alloy NPs was verified across all gold:silver ratios by a linear shift in the plasmon band maxima against alloy composition. The molar absorptivities of the NPs decreased non-linearly with increasing gold content from 2.0 x 108 M-1 cm-1 (fÉmax = 404 nm) for pure silver to 4.1 x 107 M-1 cm-1 (fÉmax = 511 nm) for pure gold. The NPs were immobilized onto transparent indium-tin oxide composite electrodes using layer-by-layer (LbL) deposition with poly(diallyldimethylammonium) acting as a cationic binder. The UV-Vis absorbance of the LbL film was used to calculate the surface coverage of alloy NPs on the electrode. Typical preparations had average NP surface coverages of 2.8 x 10-13 mol NPs/cm2 (~5% of cubic closest packing) with saturated films reaching ~20% of ccp for single-layer preparations (1.0 ~ 10-12 mol NPs/cm2). X-ray photoelectron spectroscopy confirmed the presence of alloy NPs in the LbL film and showed silver enrichment of the NP surfaces by ~9%. Irreversible oxidative dissolution (dealloying) of the less noble silver atoms from the NPs on LbL electrodes was performed by cyclic voltammetry (CV) in sulfuric acid. Alloy NPs with higher gold content required larger overpotentials for silver dealloying. Dealloying of the more-noble gold atoms from the alloy NPs was also achieved by CV in sodium chloride. The silver was oxidized first to cohesive silver chloride, and then gold dealloyed to soluble HAuCl4- at higher potentials. Silver oxidation was inhibited during the first oxidative scan, but subsequent cycles showed typical, reversible silver-to-silver chloride voltammetry. The potentials for both silver oxidation and gold dealloying also shifted to more oxidizing potentials with increasing gold content, and both processes converged for alloy NPs with >60% gold content. Charge-mediated electrochemistry of silver NPs immobilized in LbL films, using Fc(meOH) as the charge carrier, showed that 67% of the NPs were electrochemically inactive.
ContributorsStarr, Christopher A (Author) / Buttry, Daniel A (Thesis advisor) / Petuskey, William (Committee member) / Jones, Anne (Committee member) / Arizona State University (Publisher)
Created2014
Description
Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is widely accepted as the world's most abundant enzyme and represents the primary entry point for inorganic carbon into the biosphere. Rubisco's slow carboxylation rate of ribulose-1,5-bisphosphate (RuBP) and its susceptibility to inhibition has led some to term it the "bottle neck" of photosynthesis. In order to ensure that Rubisco remains uninhibited, plants require the catalytic chaperone Rubisco activase. Activase is a member of the AAA+ superfamily, ATPases associated with various cellular activities, and uses ATP hydrolysis as the driving force behind a conformational movement that returns activity to inhibited Rubisco active sites. A high resolution activase structure will be an essential tool for examining Rubisco/activase interactions as well as understanding the activase self-association phenomenon. Rubisco activase has long eluded crystallization, likely due to its infamous self-association (polydispersity). Therefore, a limited proteolysis approach was taken to identify soluble activase subdomains as potential crystallization targets. This process involves using proteolytic enzymes to cleave a protein into a few pieces and has previously proven successful in identifying crystallizable protein fragments. Limited proteolysis, utilizing two different proteolytic enzymes (alpha-chymotrypsin and trypsin), identified two tobacco activase products. The fragments that were identified appear to represent most of what is considered to be the AAA+ C-terminal all alpha-domain and some of the AAA+ N-terminal alpha beta alpha-domain. Identified fragments were cloned using the pET151/dTOPO. The project then moved towards cloning and recombinant protein expression in E. coli. NtAbeta(248-383) and NtAbeta(253-354) were successfully cloned, expressed, purified, and characterized through various biophysical techniques. A thermofluor assay of NtAbeta(248-383) revealed a melting temperature of about 30°C, indicating lower thermal stability compared with full-length activase at 43°C. Size exclusion chromatography suggested that NtAbeta(248-383) is monomeric. Circular dichroism was used to identify the secondary structure; a plurality of alpha-helices. NtAbeta(248-383) and NtAbeta(253-354) were subjected to crystallization trials.
ContributorsConrad, Alan (Author) / Wachter, Rebekka (Thesis advisor) / Moore, Thomas (Committee member) / Redding, Kevin (Committee member) / Arizona State University (Publisher)
Created2012
Description
Hydrogenases catalyze the interconversion of protons, electrons, and hydrogen according to the reaction: 2H+ + 2e- <-> H2 while using only earth abundant metals, namely nickel and iron for catalysis. The enzymatic turnover of Clostridium acetobutylicum [FeFe]-hydrogenase has been investigated through the use of electrochemical and scanning probe techniques. Scanning tunneling microscopy (STM) imaging revealed sub-monolayer surface coverage. Cyclic voltammetry yielded a catalytic, cathodic hydrogen production signal similar to that observed for a platinum electrode. From the direct observation of single enzymes and the macroscopic electrochemical measurements obtained from the same electrode, the apparent turnover frequency (TOF) per single enzyme molecule as a function of potential was determined. The TOF at 0.7 V vs. Ag/AgCl for the four SAMs yielded a decay constant for electronic coupling (β) through the SAM of ~ 0.82 Å -1, in excellent agreement with published values for similar SAMs. One mechanism used by plants to protect against damage is called nonphotochemical quenching (NPQ). Triggered by low pH in the thylakoid lumen, NPQ leads to conversion of excess excitation energy in the antenna system to heat before it can initiate production of harmful chemical species by photosynthetic reaction centers. Here a synthetic hexad molecule that functionally mimics the role of the antenna in NPQ is described. When the hexad is dissolved in an organic solvent, five zinc porphyrin antenna moieties absorb light, exchange excitation energy, and ultimately decay by normal photophysical processes. However, when acid is added, a pH-sensitive dye moiety is converted to a form that rapidly quenches the first excited singlet states of all five porphyrins, converting the excitation energy to heat and rendering the porphyrins kinetically incompetent to perform useful photochemistry. Charge transport was also studied in single-molecule junctions formed with a 1,7-pyrrolidine-substituted 3,4,9,10-Perylenetetracarboxylic diimide (PTCDI) molecule. A reduction in the highest occupied (HOMO) and lowest unoccupied (LUMO) molecular orbitals energy gap due to the electronic properties of the substituents is seen when compared to an unsubstituted-PTCDI. The small HOMO-LUMO energy gap allows for switching between electron- and hole-dominated charge transport with a gate voltage, thus demonstrating a single-molecule ambipolar field effect transistor.
ContributorsMadden, Christopher (Author) / Moore, Thomas A. (Thesis advisor) / Jones, Anne (Committee member) / Tao, Nongjian (Committee member) / Arizona State University (Publisher)
Created2012
Description
The heliobacterial reaction center (HbRC) is widely considered the simplest and most primitive photosynthetic reaction center (RC) still in existence. Despite the simplicity of the HbRC, many aspects of the electron transfer mechanism remain unknown or under debate. Improving our understanding of the structure and function of the HbRC is important in determining its role in the evolution of photosynthetic RCs. In this work, the function and properties of the iron-sulfur cluster FX and quinones of the HbRC were investigated, as these are the characteristic terminal electron acceptors used by Type-I and Type-II RCs, respectively. In Chapter 3, I develop a system to directly detect quinone double reduction activity using reverse-phase high pressure liquid chromatography (RP-HPLC), showing that Photosystem I (PSI) can reduce PQ to PQH2. In Chapter 4, I use RP-HPLC to characterize the HbRC, showing a surprisingly small antenna size and confirming the presence of menaquinone (MQ) in the isolated HbRC. The terminal electron acceptor FX was characterized spectroscopically and electrochemically in Chapter 5. I used three new systems to reduce FX in the HbRC, using EPR to confirm a S=3/2 ground-state for the reduced cluster. The midpoint potential of FX determined through thin film voltammetry was -372 mV, showing the cluster is much less reducing than previously expected. In Chapter 7, I show light-driven reduction of menaquinone in heliobacterial membrane samples using only mild chemical reductants. Finally, I discuss the evolutionary implications of these findings in Chapter 7.
ContributorsCowgill, John (Author) / Redding, Kevin (Thesis advisor) / Jones, Anne (Committee member) / Fromme, Petra (Committee member) / Arizona State University (Publisher)
Created2012
Description
Hydrogen has the potential to be a highly efficient fuel source. Its current production via steam reformation of natural gas, however, consumes a large amount of energy and gives off carbon dioxide. A newer method has since surfaced: using a microorganism's metabolism to drive hydrogen production. In this study, the conditions for maximum hydrogen production in Heliobacterium modesticaldum were identified and assessed. The cells were grown under varying conditions and their headspaces were sampled using a gas chromatogram to measure the amount of accumulated hydrogen during each condition. Two cell batches were grown under nitrogen-fixing conditions (-NH4+), while the other two cell batches were grown under non-nitrogen-fixing conditions (+NH4+). The headspaces were then exchanged with either nitrogen (N2) or argon (Ar2). It was found that the condition for which the most hydrogen was produced was when the cells were grown under nitrogen-fixing conditions and the headspace was exchanged with argon. These results suggest that most of Heliobacteria modesticaldum's hydrogen production is due to nitrogenase activity rather than hydrogenase activity. Further research is recommended to quantify the roles of nitrogenase, [NiFe] hydrogenase, and [FeFe] hydrogenase.
ContributorsMcmahon, Savanah Dior (Author) / Redding, Kevin (Thesis director) / Ghirlanda, Giovanna (Committee member) / Barrett, The Honors College (Contributor) / Department of Chemistry and Biochemistry (Contributor) / School of Music (Contributor)
Created2015-05
Description
F2-isoprostanes are a series of prostaglandin-like compounds derived from the free radical-mediated lipid peroxidation of arachidonic acid, a polyunsaturated fatty acid that is ubiquitously expressed in cell membranes. F2-isoprostanes are biomarkers of oxidative stress, an imbalance between oxidants and antioxidants that can cause damage to DNA, proteins, lipids, and carbohydrates. Increased production of lipid peroxidation products have been implicated in the pathology of a number of conditions and diseases in humans. The objective of this thesis was to (1) optimize the LC/MS/MS F2-isoprostane method currently used in human samples for use in research animals and veterinary medicine, including the use of solid phase extraction, and (2) validate the optimized method in rodent and canine experimental studies. Our optimized method showed that Lyprinol treatment in dogs with osteoarthritis decreases F2-isoprostane levels nearly 2-fold. In addition, adjuvant alpha-tocopherol prevented tumor-induced increased F2-isoprostane levels. Finally, contrary to earlier studies using less specific ELISA F2-isoprostane methods, we demonstrate that unconditioned dogs benefit from low intensity exercise. Our data demonstrate successful optimization of the human LC/MS/MS F2-isoprostane method in rats and canines. Importantly, our results emphasize the need to use the more sensitive and specific LC/MS/MS method as compared to ELISA-based assays in order to distinguish the 15- and 5-series F2-isoprostanes, evidenced in particular by the two canine studies.
ContributorsCorrigan, Devin Connell (Author) / Redding, Kevin (Thesis director) / Anderson, Karen (Committee member) / Mustacich, Debbie (Committee member) / Barrett, The Honors College (Contributor) / School of Life Sciences (Contributor) / Department of Chemistry and Biochemistry (Contributor)
Created2015-05
Description
With a quantum efficiency of nearly 100%, the electron transfer process that occurs within the reaction center protein of the photosynthetic bacteria Rhodobacter (Rh.) sphaeroides is a paragon for understanding the complexities, intricacies, and overall systemization of energy conversion and storage in natural systems. To better understand the way in which photons of light are captured, converted into chemically useful forms, and stored for biological use, an investigation into the reaction center protein, specifically into its cascade of cofactors, was undertaken. The purpose of this experimentation was to advance our knowledge and understanding of how differing protein environments and variant cofactors affect the spectroscopic aspects of and electron transfer kinetics within the reaction of Rh. sphaeroides. The native quinone, ubiquinone, was extracted from its pocket within the reaction center protein and replaced by non-native quinones having different reduction/oxidation potentials. It was determined that, of the two non-native quinones tested—1,2-naphthaquinone and 9,10- anthraquinone—the substitution of the anthraquinone (lower redox potential) resulted in an increased rate of recombination from the P+QA- charge-separated state, while the substitution of the napthaquinone (higher redox potential) resulted in a decreased rate of recombination.
ContributorsSussman, Hallie Rebecca (Author) / Woodbury, Neal (Thesis director) / Redding, Kevin (Committee member) / Lin, Su (Committee member) / School of Molecular Sciences (Contributor) / Barrett, The Honors College (Contributor)
Created2015-12