Biochemistry, Biophysics, and Structural Biology Commons^™

Evolutionary And Molecular Analysis Of Conserved Vertebrate Immunity To Fungi, Erin Carter

Honors College

The innate immune system is highly conserved amongst all multicellular organisms. Yet a constant battle exists between host cells and pathogens due to the rapid evolution of immune system components. Functional genomics and in silico methods can be employed to elucidate the evolutionary patterns of vertebrate immunity to pathogenic fungi such as Candida albicans, an opportunistic fungal pathogen that can cause lethal candidiasis in the immunocompromised. Mammals such as humans and mice possess conserved C-type lectin receptors that recognize the C. albicans cell wall. However, these receptors have not been identified in fish. Here I describe how we identified potential …

Fast Pyrolysis Of Muconic Acid And Formic Acid Salts, Laura Duran

Honors College

Lignocellulosic biomass is emerging as a sustainable resource for the production of alternative liquid fuels. As the need to lessen dependence on petroleum sources grows, lignocellulosic feedstocks are being investigated as a renewable, abundant source of energy. Chemical pulping processes include a high-lignin by-product, black liquor, which is already used for fuel in industry. Black liquor is burned to generate steam and electricity and to recover pulping chemicals. Currently, the thermochemical conversion of black liquor to liquid fuel is being researched at The University of Maine. In this black liquor research, an intermediate lignin-derived acid, muconic acid, and formic acid …

Reverse Genetic Analysis Of A Cysteine Protease-Encoding Gene (Rd19a) Of Arabidopsis Thaliana In Relation To The Mechanism Of Resistance To The Piercing/Sucking Insect Myzus Persicae, Siobhan A. Cusack

Honors College

A recent study in Solanum bulbocastanum (a wild relative of the cultivated potato) aiming to identify potential genes involved in aphid and pathogen resistance mechanisms found that a homolog of the Arabidopsis thaliana cysteine protease gene RD19a is upregulated during aphid infestation. RD19a is upregulated in response to abiotic stresses such as drought and high salinity, and rd19a mutants show increased susceptibility to bacterial infection. In this study, Arabidopsis rd19a mutants and wild-type plants were subjected to aphid feeding to observe and compare the molecular, physiological and phenotypic responses. The aim was to further establish the proof of concept regarding …

Characterizing The Role Of Cortactin In Actin Pedestal Assembly By Enterohemorrhagic Escherichia Coli (Ehec), Sarah E. Grout

Honors Scholar Theses

Enterohemorrhagic Escherichia coli (EHEC) is a major foodborne cause of bloody diarrhea and renal failure. During colonization of the intestine, EHEC injects the transmembrane receptor protein Tir and the cytoplasmic effector protein EspFU into host cells to reorganize the actin cytoskeleton into adhesion “pedestals.” EspFU has been shown to bind and activate the actin nucleation factor N-WASP to drive actin polymerization into pedestals. However, EspFU can still assemble pedestals in cells lacking N-WASP, suggesting that this effector protein is able to also trigger N-WASP-independent pathways of actin polymerization during infection. Cortactin is an atypical nucleation factor that localizes to pedestals, …

Patatin-Related Phospholipase Pplaiiiδ Increases Seed Oil Content With Long-Chain Fatty Acids In Arabidopsis, Maoyin Li, Sung Bahn, Chuchuan Fan, Jia Li, Tien Phan, Michael Ortiz, Mary Roth, Ruth Welti, Jan Jaworski, Xuemin Wang

Biology Department Faculty Works

The release of fatty acids from membrane lipids has been implicated in various metabolic and physiological processes, but in many cases, the enzymes involved and their functions in plants remain unclear. Patatin-related phospholipase As (pPLAs) constitute a major family of acyl-hydrolyzing enzymes in plants. Here, we show that pPLAIIId promotes the production of triacylglycerols with 20- and 22-carbon fatty acids in Arabidopsis (Arabidopsis thaliana). Of the four pPLAIIIs (a, b, g, d), only pPLAIIId gene knockout results in a decrease in seed oil content, and pPLAIIId is most highly expressed in developing embryos. The overexpression of pPLAIIId increases the content …

The Role Of Oswrky71 And Its Interacting Proteins In Seed Germination And Early Growth Of Cereal Grains, Margaret Ja Shin

UNLV Theses, Dissertations, Professional Papers, and Capstones

During seed germination and early seedling growth, complex molecular and physiological events occur in rice (Oryza sativa) and other cereal grains. As the seed transitions to vegetative tissue, it responds to both favorable and unfavorable environmental conditions and is vulnerable to attack by predation and disease. Although seeds are relatively small and tender in size, extensive and sophisticated molecular networks enables the immobile seed to grow, survive and adapt in its environment. One of the networks I am interested in is in the crosstalk between the gibberellin (GA) and abscisic acid (ABA) signaling pathways. These pathways are interesting because they …

High-Throughput Approaches For The Assessment Of Factors Influencing Bioavailability Of Small Molecules In Pre-Clinical Drug Development, Megan Marie Mccallum

Theses and Dissertations

A bioactive molecule must pass many hurdles to be designated as a "good" pharmaceutical lead or hit compound. It should have a significant activity, selectivity, bioavailability, and metabolic half-life. Many factors have been identified that influence the free drug concentration or bioavailability of orally administered drugs in the earliest development stages. In vitro pre-clinical assays have been developed to measure these parameters. The small molecule properties that are investigated here include aqueous solubility, permeability, reactivity (electrophilicity), small molecule-protein binding, and displacement of protein-bound molecules (drug-drug interactions). The development of rapid and miniaturized assays to quantify these factors is presented herein. …

Methods In Metallomics, Proteomics, And Toxicology: Development And Applications Of Laser Ablation Inductively Coupled Plasma Mass Spectrometry And Native Sds-Page, William John Wobig

Theses and Dissertations

Polyacrylamide gel electrophoresis (PAGE) is a bio-analytical method used to separate proteins in solution into an array of individual bands of proteins in a gel matrix. Current PAGE methods, however, have severe limitations in simultaneously maintaining a protein's native structure and association with transition metals while providing adequate resolution. Laser Ablation Inductively Coupled Plasma Mass Spectrometry (LA-ICP-MS) provides a means to perform trace to ultra-trace level inorganic analysis of solid samples such as dried PAGE gels containing metallo-protein arrays. Current LA-ICP-MS methods involving the analysis of PAGE gels, however, have been limited in their effective use by inadequate limits of …

Assessing Tissue Characterization Of Abdominal Organs Using Fuzzy C-Means Cluster Analysis Of Color- Fusion Mr Images, Terrance D. Weeden, H. Keith Brown

Terrance Weeden

Structural And Functional Characterization Of Sortase A, Vishaka Santosh

Theses and Dissertations

Sortases have been known to be essential in Gram-positive bacteria for attaching proteins onto the peptidoglycan layer of the bacterium. Sortase A has been found to be useful as a “molecular stapler”, although; in vivo, the enzyme is responsible for attaching proteins to the peptidoglycan layer of Gram-positive bacteria. It accomplishes both of these tasks by joining two proteins together via an LPXTG sorting sequence. The enzyme has been proven to be very useful in attaching any two proteins together without worrying about recombinant techniques to generate the fusion protein. The problem with this enzyme is that the catalytic diad, …

Nuclear Localization Of Cpi-17, A Protein Phosphatase-1 Inhibitor Protein, Affects Histone H3 Phosphorylation And Corresponds To Proliferation Of Cancer And Smooth Muscle Cells., Masumi Eto, Jason A Kirkbride, Rishika Chugh, Nana Kofi Karikari, Jee In Kim

Department of Molecular Physiology and Biophysics Faculty Papers

CPI-17 (C-kinase-activated protein phosphatase-1 (PP1) inhibitor, 17kDa) is a cytoplasmic protein predominantly expressed in mature smooth muscle (SM) that regulates the myosin-associated PP1 holoenzyme (MLCP). Here, we show CPI-17 expression in proliferating cells, such as pancreatic cancer and hyperplastic SM cells. Immunofluorescence showed that CPI-17 was concentrated in nuclei of human pancreatic cancer (Panc1) cells. Nuclear accumulation of CPI-17 was also detected in the proliferating vascular SM cell culture and cells at neointima of rat vascular injury model. The N-terminal 21-residue tail domain of CPI-17 was necessary for the nuclear localization. Phospho-mimetic Asp-substitution of CPI-17 at Ser12 attenuated the nuclear …

The Activation, Receptor Complexing And Endogenous Regulation Of The Type-I Interferon Response As It Pertains To Innate Immunity, James D. Marion Jr.

Theses and Dissertations

To defend against pathogen challenge, multi-cellular organisms mount an immune response that recognizes, sequesters and eradicates invading infectious agents. Critical to this safeguard is the receptor-mediated detection of pathogens. Pathogen recognition then initiates a variety of signaling cascades that lead to the modulation of genes orchestrating an immune response. Toll-like receptor 3 (TLR3), a transmembrane receptor found in endosomes, is vital to the innate immune response against viruses. Double-stranded RNA (dsRNA) stimulation of TLR3 initiates a signaling cascade that leads to the production of type-I interferons and proinflammatory cytokines necessary to trigger the protective defenses of the immune system. Critical …

Serine Proteolytic Pathway Activation Reveals An Expanded Ensemble Of Wound Response Genes In Drosophila, Rachel A. Patterson, Michelle T. Juarez, Anita Hermann, Roman Sasik, Gary Hardiman, William Mcginnis

Publications and Research

After injury to the animal epidermis, a variety of genes are transcriptionally activated in nearby cells to regenerate the missing cells and facilitate barrier repair. The range and types of diffusible wound signals that are produced by damaged epidermis and function to activate repair genes during epidermal regeneration remains a subject of very active study in many animals. In Drosophila embryos, we have discovered that serine protease function is locally activated around wound sites, and is also required for localized activation of epidermal repair genes. The serine protease trypsin is sufficient to induce a striking global epidermal wound response without …

Genetic Parameters For Concentrations Of Minerals In Longissimus Muscle And Their Associations With Palatability Traits In Angus Cattle, R. G. Mateescu, A. J. Garmyn, Richard G. Tait Jr., Qing Duan, Q. Liu, Mary S. Mayes, Dorian J. Garrick, A. L. Van Eenennaam, D. L. Vanoverbeke, G. G. Hilton, Donald C. Beitz, James M. Reecy

Richard G. Tait Jr.

The objective of this study was to estimate genetic parameters for concentrations of minerals in LM and to evaluate their associations with beef palatability traits. Samples of LM from 2,285 Angus cattle were obtained and fabricated into steaks for analysis of mineral concentrations and for trained sensory panel assessments. Nine minerals, including calcium, copper, iron, magnesium, manganese, phosphorus, potassium, sodium, and zinc, were quantified. Restricted maximum likelihood procedures were used to obtain estimates of variance and covariance components under a multiple-trait animal model. Estimates of heritability for mineral concentrations in LM varied from 0.01 to 0.54. Iron and sodium were …

Genetic Parameters For Carnitine, Creatine, Creatinine, Carnosine, And Anserine Concentration In Longissimus Muscle And Their Association With Palatability Traits In Angus Cattle, R. G. Mateescu, A. J. Garmyn, M. A. O'Neil, Richard G. Tait Jr., Almass A. Abuzaid, Mary S. Mayes, Dorian J. Garrick, A. L. Van Eenennaam, D. L. Vanoverbeke, G. G. Hilton, Donald C. Beitz, James M. Reecy

Richard G. Tait Jr.

The objective of this study was to estimate genetic parameters for carnitine, creatine, creatinine, carnosine, and anserine concentration in LM and to evaluate their associations with Warner-Bratzler shear force (WBSF) and beef palatability traits. Longissimus muscle samples from 2,285 Angus cattle were obtained and fabricated into steaks for analysis of carnitine, creatine, creatinine, carnosine, anserine, and other nutrients, and for trained sensory panel and WBSF assessments. Restricted maximum likelihood procedures were used to obtain estimates of variance and covariance components under a multiple-trait animal model. Estimates of heritability for carnitine, creatine, creatinine, carnosine, and anserine concentrations in LM from Angus …

Mesobiliverdin Ixα Enhances Rat Pancreatic Islet Yield And Function, Taihei Ito, Dong Chen, Cheng-Wei Tom Chang, Takashi Kenmochi, Tomonori Saito, Satoshi Suzuki, Jon Y. Takemoto

Chemistry and Biochemistry Faculty Publications

The aims of this study were to produce mesobiliverdin IXα, an analog of anti-inflammatory biliverdin IXα, and to test its ability to enhance rat pancreatic islet yield for allograft transplantation into diabetic recipients. Mesobiliverdin IXα was synthesized from phycocyanobilin derived from cyanobacteria, and its identity and purity were analyzed by chromatographic and spectroscopic methods. Mesobiliverdin IXα was a substrate for human NADPH biliverdin reductase. Excised Lewis rat pancreata infused with mesobiliverdin IXα and biliverdin IXα-HCl (1–100 μM) yielded islet equivalents as high as 86.7 and 36.5%, respectively, above those from non-treated controls, and the islets showed a high degree of …

Divergent Protein Motifs Direct Ef-P Mediated Translational Regulation In Salmonella And Escherichia Coli, Steven J. Hersch, Mengchi Wang, S. Betty Zou, Kyung-Mee Moon, Leonard J. Foster, Michael Ibba, William Wiley Navarre

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Elongation factor P (EF-P) is a universally conserved bacterial translation factor homologous to eukaryotic/archaeal initiation factor 5A. In Salmonella, deletion of the efp gene results in pleiotropic phenotypes, including increased susceptibility to numerous cellular stressors. Only a limited number of proteins are affected by the loss of EF-P, and it has recently been determined that EF-P plays a critical role in rescuing ribosomes stalled at PPP and PPG peptide sequences. Here we present an unbiased in vivo investigation of the specific targets of EF-P by employing stable isotope labeling of amino acids in cell culture (SILAC) to compare the …

Metabolism And Transcriptional Responses To Asparagine In Arabidopsis Thaliana, Qianyi Zhang

Electronic Thesis and Dissertation Repository

Asparagine aminotransferase transforms asparagine into α-ketosuccinamate, which is further deamidated by an ω-amidase. Serine:glyoxylate aminotransferase, encoded by AGT1 in Arabidopsis, was identified as asparagine aminotransferase. In the roots of 10-day-old Arabidopsis seedlings treated with 20 mM asparagine, AGT1 transcript levels increased by 2-fold while ω-amidase transcripts were decreased by 30%. Recombinant AGT1 had a substrate preference for asparagine when compared with alanine and serine as amino group donors. An ω-amidase candidate gene, AT5G12040, was identified based on amino acid sequence identity with mammalian gene Nitrilase 2. RT-PCR of a T-DNA insertion mutant line showed that ω-amidase expression was abolished …

Cellular Adaptation Of Macrophages To Anthrax Lethal Toxin-Induced Pyroptosis Via Epigenetic Mechanisms, Chae Young Han

Electronic Thesis and Dissertation Repository

Cellular adaptation to microbial stresses has been demonstrated in several cell types. Macrophages (MФ) are sentinel immune cells fending off invading microbes. Anthrax lethal toxin (LeTx) is a key virulence factor released by Bacillus anthracis that causes rapid cell death, pyroptosis. A small number of RAW246.7 macrophages (~4%) exposed to a non-lethal dose of LeTx become resistant to LeTx-induced pyroptosis for ~ 4 weeks, termed “toxin-induced resistance (TIR)”. Here, I showed that high levels of DNA methyl transferase1 (DNMT1) expression were maintained although global genomic methylation levels were not high in TIR. TIR cells treated with the DNMT inhibitor 5-azacitidine …

Effects Of Beta Amyloid On The Dna Methylation Status Of An In Vitro Model Of Alzheimer’S Disease, Noor Taher

Senior Honors Theses

Available evidence points toward an epigenetic process in Alzheimer’s disease. This thesis describes the research that was done to investigate changes in DNA methylation using an in vitro model of the disease. Although the results indicated no global changes in methylation levels after treating differentiated IMR-32 cells with beta amyloid, there were several regions of the genome that changed their methylation status. Gene ontology studies revealed that these regions are associated with neuronal differentiation and cell fate genes, thus providing a possible model for the contribution of beta amyloid to the development of Alzheimer’s disease. This study provides incentive to …