Matching Items (43)

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Characterization of a Polyclonal Antibody Specific for Synechococcus WH8102 Plastoquinol Terminal Oxidase.

Description

Photosynthesis is a critical process that fixes the carbon utilized in cellular respiration. In higher plants, the immutans gene codes for a protein that is both involved in carotenoid biosynthesis

Photosynthesis is a critical process that fixes the carbon utilized in cellular respiration. In higher plants, the immutans gene codes for a protein that is both involved in carotenoid biosynthesis and plastoquinol oxidation (Carol et al 1999, Josse et al 2003). This plastoquinol terminal oxidase (PTOX) is of great interest in understanding electron flow in the plastoquinol pool. In order to characterize this PTOX, polyclonal antibodies were developed. Expression of Synechococcus WH8102 PTOX in E. coli provided a useful means to harvest the protein required for antibody production. Once developed, the antibody was tested for limit of concentration, effectiveness in whole cell lysate, and overall specificity. The antibody raised against PTOX was able to detect as low as 10 pg of PTOX in SDS-PAGE, and could detect PTOX extracted from lysed Synechococcus WH8102. The production of this antibody could determine the localization of the PTOX in Synechococcus.

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  • 2014-05

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The Neurochemical Consequences of Music Therapy on Dementia Patients

Description

As the incidence of dementia continues to rise, the need for an effective and non-invasive method of intervention has become increasingly imperative. Music therapy has exhibited these qualities in addition

As the incidence of dementia continues to rise, the need for an effective and non-invasive method of intervention has become increasingly imperative. Music therapy has exhibited these qualities in addition to relatively low implementation costs, therefore establishing itself as a promising means of therapeutic intervention. In this review, current research was investigated in order to determine its effectiveness and uncover the neurochemical mechanisms that lead to positive manifestations such as improved memory recall, increased social affiliation, increased motivation, and decreased anxiety. Music therapy has been found to improve several aspects of memory recall. One proposed mechanism involves temporal entrainment, during which the melodic structures present in music provide a framework for chunking information. Although entrainment's role in the treatment of motor defects has been thoroughly studied, its role in treating cognitive disorders is still relatively new. Musicians have also been shown to demonstrate extensive plastic changes; therefore, it is hypothesized that non-musicians may also glean some benefits from engaging in music. Social affiliation has been found to increase due to increases in endogenous oxytocin. Oxytocin has also been shown to strengthen hippocampal spike transmission, a promising outcome for Alzheimer's patients. An increase in motivation has also been found to occur due to music's ability to tap into the reward center of the brain. Dopaminergic transmission between the VTA, NAc and higher functioning regions such as the OFC and hypothalamus has been revealed. Additionally, relaxing music decreases stress levels and modifies associated autonomic processes, i.e. heart rate, blood pressure, and respiratory rate. On the contrary, stimulating music has been found to initiate sympathetic nervous system activity. This is thought to occur by either a reflexive brainstem response or stimulus interpretation by the amygdala.

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Date Created
  • 2016-05

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F2-isoprostanes and F2-isoprostane Metabolites: Biomarkers for Oxidative Stress and Therapeutic Efficacy

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

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.

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Date Created
  • 2015-05

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Quinone Removal and Replacement within the Reaction Center Protein of Rhodobacter sphaeroides

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

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.

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Date Created
  • 2015-12

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Examining the Effect of Vinegar on Glucose Response

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The objective of this randomized, single-blind crossover study was to examine the effect of vinegar on the blood glucose response to meal ingestion. This study was associated with a

The objective of this randomized, single-blind crossover study was to examine the effect of vinegar on the blood glucose response to meal ingestion. This study was associated with a companion study Is Apple Cider Vinegar Effective for Reducing Heartburn Symptoms Related to Gastroesophageal Reflux Disease. Glucose meters were utilized to measure blood glucose levels immediately prior to, and at four ½ hour intervals following meal ingestion. Previous studies have demonstrated that vinegar modulates the meal-time glucose response. Hence an alternative hypothesis was used: that a significant difference will be observed between the control and the vinegar groups. The results from the study were not significant likely due to a small sample size. The test meal eaten with a drink composed of vinegar diluted in water appeared to be most effective at decreasing the overall change in postprandial blood glucose. The vinegar drink also played a role in decreasing the peak glucose level at 30 minutes post-meal.

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Date Created
  • 2015-05

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ELECTRON TRANSFER PROCESS BETWEEN COFACTORS OF HELIOBACTERIA'S REACTION CENTER

Description

ABSTRACT:
The experiment was conducted to analyze the role of menaquinone (MQ) in heliobacteria’s reaction center (HbRC). Their photosynthetic apparatus is a homodimeric of type I reaction center (1). HbRC

ABSTRACT:
The experiment was conducted to analyze the role of menaquinone (MQ) in heliobacteria’s reaction center (HbRC). Their photosynthetic apparatus is a homodimeric of type I reaction center (1). HbRC contains these cofactors: P800 (special pair cholorphyll), A0 (8-hydroxy-chlorophyll [Chl] a), and FX (iron-sulfur cluster). The MQ factor is bypassed during the electron transfer process in HbRC. Electrons from the excited state of P800 (P800*) are transported to A0 and then directly to Fx. The hypothesis is that when electrons are photoaccumulated at Fx, and without the presence of any electron acceptors to the cluster, they would be transferred to MQ, and reduce it to MQH2 (quinol). Experiments conducted in the past with HbRC within the cell membranes yielded data that supported this hypothesis (Figures 4 and 5). We conducted a new experiment based on that foundation with HbRC, isolated from cell membrane. Two protein assays were prepared with cyt c553 and ascorbate in order to observe this phenomenon. The two samples were left in the glove box for several days for equilibration and then exposed to light in different intensity and periods. Their absorption was monitored at 800 nm for P800 or 554 nm for cyt c553 to observe their oxidation and reduction processes. The measurements were performed with the JTS-10 spectrophotometer. The data obtained from these experiments support the theory that P800+ reduced by the charge recombination of P800+Fx-. However, it did not confirm the reduction of P800+ done by cyt c553¬ which eventually lead to a net accumulation of oxidized cyt c553; instead it revealed another factor that could reduce P800+ faster and more efficient than cyt c553 (0.5 seconds vs several seconds), which could be MQ. More experiments need to be done in order to confirm this result. Hence, the data collected from this experiment have yet to support the theory of MQ being reduced to MQH2 outside the bacterial membranes.

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Date Created
  • 2015-05

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The Optimization of Conditions for Maximum Hydrogen Production in Heliobacterium modesticaldum

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

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.

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Date Created
  • 2015-05

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Analysis of Acyl Carrier Protein in the cyanobacterium Synechocystis sp. PCC 6803

Description

Acyl Carrier Protein (ACP) is a small, acidic protein that plays an essential role in fatty acid synthesis by elongating fatty acid chains. ACP was isolated from an extract of

Acyl Carrier Protein (ACP) is a small, acidic protein that plays an essential role in fatty acid synthesis by elongating fatty acid chains. ACP was isolated from an extract of a modified strain of Synechocystis sp. PCC 6803 that contains a thioesterase and from which the acyl-ACP synthetase has been deleted. Using ammonium sulfate precipitation to isolate a crude protein fraction containing ACP, immunoblot analysis was performed to determine relative amounts of free and acylated-ACP in the cell. The nature of fatty acids attached to ACP was determined by creating butylamide derivatives that were analyzed using GC/MS. Immunoblot analysis showed a roughly 1:1 ratio of acylated ACP to free ACP in the cell depending on the nutritional state of the cell. From GC/MS data it was determined that palmitic acid was the predominate component of acyl groups attached to ACP. The results indicate that there is a significant amount of acyl-ACP, a feedback inhibitor of early steps in the fatty acid biosynthesis pathway, in the cell. Moreover, the availability of free ACP may also limit fatty acid biosynthesis. Most likely it is necessary for ACP to be overexpressed or to have the palmitic acid cleaved off in order to synthesize optimal amounts of lauric acid to be used for cyanobacterial biofuel production.

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Date Created
  • 2016-05

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Correlational Analysis of Intelligence Mindset, Motivation Backgrounds, and Significance of Gender Identity

Description

The goal of this investigation was to perform a correlational analysis of the intelligence mindsets, motivational background, and significance of gender identity as factors driving student success. 42 students enrolled

The goal of this investigation was to perform a correlational analysis of the intelligence mindsets, motivational background, and significance of gender identity as factors driving student success. 42 students enrolled in Computer Science and Engineering (CSE) 110: Principles of Programming with Java completed a modified Scientific Measurement Questionnaire (SMQ), a survey instrument designed to study the previously mentioned factors. This survey was modeled on a similar survey administered by Dr. Ian Gould to students enrolled in his Organic Chemistry course at Arizona State University. Following the development of a scoring system to generate quantifiable data, it was determined that students in this course displayed a greater inclination towards beliefs in malleable intelligence and in an intrinsic locus of control as opposed to a belief in static intelligence and an external locus of control. Students exhibited a multi-faceted approach in responding to the questions in the motivational background section, indicating that there were no distinctively dominating factors driving student motivation. Instead, it was observed that students generally derived motivation from these factors in a synergistic fashion. Responses to questions regarding gender indicated that while students believed that the way they were perceived by others was significantly influenced by their gender, the notion of gender identity played little to no role in their overall personal identity and self-schema. As the study was designed to offer insight into the role of gender identity and the population discrepancies within the course, it is important to note that the findings suggest gender identity is not a primary factor of concern with regard to student performance. While the data acquired suggested potential trends in student mindsets, a notable limitation of the scope of the project was the undersized sample population.

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  • 2016-05

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Elucidating Structural and Functional Information on the Human Cold-Sensing Protein TRPM8 via Isolating the Pore Domain and Cross-Chimeric Studies

Description

Transient Receptor Potential (TRP) ion channels are a diverse family of nonselective, polymodal sensors in uni- and multicellular eukaryotes that are implicated in an assortment of biological contexts and human

Transient Receptor Potential (TRP) ion channels are a diverse family of nonselective, polymodal sensors in uni- and multicellular eukaryotes that are implicated in an assortment of biological contexts and human disease. The cold-activated TRP Melastatin-8 (TRPM8) channel, also recognized as the human body's primary cold sensor, is among the few TRP channels responsible for thermosensing. Despite sustained interest in the channel, the mechanisms underlying TRPM8 activation, modulation, and gating have proved challenging to study and remain poorly understood. In this thesis, I offer data collected on various expression, extraction, and purification conditions tested in E. Coli expression systems with the aim to optimize the generation of a structurally stable and functional human TRPM8 pore domain (S5 and S6) construct for application in structural biology studies. These studies, including the biophysical technique nuclear magnetic spectroscopy (NMR), among others, will be essential for elucidating the role of the TRPM8 pore domain in in regulating ligand binding, channel gating, ion selectively, and thermal sensitivity. Moreover, in the second half of this thesis, I discuss the ligation-independent megaprimer PCR of whole-plasmids (MEGAWHOP PCR) cloning technique, and how it was used to generate chimeras between TRPM8 and its nearest analog TRPM2. I review steps taken to optimize the efficiency of MEGAWHOP PCR and the implications and unique applications of this novel methodology for advancing recombinant DNA technology. I lastly present preliminary electrophysiological data on the chimeras, employed to isolate and study the functional contributions of each individual transmembrane helix (S1-S6) to TRPM8 menthol activation. These studies show the utility of the TRPM8\u2014TRPM2 chimeras for dissecting function of TRP channels. The average current traces analyzed thus far indicate that the S2 and S3 helices appear to play an important role in TRPM8 menthol modulation because the TRPM8[M2S2] and TRPM8[M2S3] chimeras significantly reduce channel conductance in the presence of menthol. The TRPM8[M2S4] chimera, oppositely, increases channel conductance, implying that the S4 helix in native TRPM8 may suppress menthol modulation. Overall, these findings show that there is promise in the techniques chosen to identify specific regions of TRPM8 crucial to menthol activation, though the methods chosen to study the TRPM8 pore independent from the whole channel may need to be reevaluated. Further experiments will be necessary to refine TRPM8 pore solubilization and purification before structural studies can proceed, and the electrophysiology traces observed for the chimeras will need to be further verified and evaluated for consistency and physiological significance.

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Date Created
  • 2016-05