Biochemistry Commons^™

Evaluation Of The Protein Recognition Properties Of Peptide Nucleic Acids, Crystal Cox Serrano

Master's Theses

The objective is to evaluate the ability of aminoacyl-tRNA synthetases (aaRS) to recognize the non-standard nucleic acid, PNA (peptide nucleic acid). PNA has immense potential in biomedical applications due to its increased thermostability and nuclease resistance over natural nucleic acids. PNA represents a superior alternative to natural nucleic acids in many biomedical applications due to its specificity, strong binding, and nuclease resistance. This study is the initial data set that indicates PNA are recognized by translation enzymes.

Nmr-Based Structural Analysis Of Threonylcarbamoyl-Amp Synthase And Its Substrate Interactions, Kimberly A. Harris, Benjamin G. Bobay, Kathryn L. Sarachan, Alexis F. Sims, Yann Bilbille, Christopher Deutsch, Dirk Iwata-Reuyl, Paul F. Agris

Chemistry Faculty Publications and Presentations

The hypermodified nucleoside N(6)-threonylcarbamoyladenosine (t(6)A37) is present in many distinct tRNA species and has been found in organisms in all domains of life. This post-transcriptional modification enhances translation fidelity by stabilizing the anticodon/codon interaction in the ribosomal decoding site. The biosynthetic pathway of t(6)A37 is complex and not well understood. In bacteria, the following four proteins have been discovered to be both required and sufficient for t(6)A37 modification: TsaC, TsaD, TsaB, and TsaE. Of these, TsaC and TsaD are members of universally conserved protein families. Although TsaC has been shown to catalyze the formation of L-threonylcarbamoyl-AMP, a key intermediate in …

Investigating The Interaction Of Aurka And Ube2c In Colorectal Cancer Cells, Apurva M. Hegde

Dissertations and Theses (Open Access)

Colorectal cancer (CRC) is the third leading cause of cancer-related deaths in the US. Among the many genomic aberrations previously implicated in colorectal cancer, recurrent amplification of chromosome 20q is frequently associated with liver metastasis. Previous research in our lab identified a gene signature on chromosome 20q associated with colorectal cancer progression. In this study, one of the genes in the signature, the ubiquitin conjugating enzyme UBE2C, was identified through preliminary bioinformatics analysis as a candidate for further examination of its role in CRC progression. Co-expression analysis of UBE2C in tumor-normal datasets from the public database Oncomine revealed all the …

Histone H3 K4 Methylation Regulates The Spindle Assembly Checkpoint Through Direct Binding Of Multiple Checkpoint Components And Cdc20, Andria C. Schibler

Dissertations and Theses (Open Access)

Histone H3K4 methylation is conserved across species and is associated with active transcription. By using Saccharomyces cerevisiae, we found histone H3K4 methylation has a previously unknown role in regulating mitosis through the Spindle Assembly Checkpoint. The Spindle Assembly Checkpoint ensures duplicated chromosomes are segregated correctly and each daughter cell receives one full copy of the genome. Our data show SET1 mutants and histone H3K4 mutants display a resistance to the mitotic poison, benomyl. Moreover methylated histone H3 directly binds to Spindle Assembly Checkpoint proteins Bub3 and Mad2 as well as the activator of the Anaphase Promoting Complex (APC) protein …

Paenibacillus Larvae Spore Germination And American Foulbrood Disease Development In Honey Bee Larvae, Israel Alvarado

UNLV Theses, Dissertations, Professional Papers, and Capstones

Honey bee (Apis mellifera) pollination plays an important role in meeting our agricultural needs, yet we are incapable of completely preventing a decline in bee health, partially due to bacterial diseases. American Foulbrood (AFB), a bacterial disease of honey bee larvae, is particularly troublesome because the infectious agent is the bacterial spore of Paenibacillus larvae. Bacterial spores are resistant to extreme temperatures, unaffected by antibiotics, withstand exposure to disinfectants, and can remain dormant for years until they can revert back to vegetative cells.

Our research investigated P. larvae spore germination at the functional and pathogen-host level. We have found that …

Coordinate Regulation Of The Wnt And Hippo Pathways By The Novel Ras Effector Nore1a., Marvin Lee Schmidt Jr.

Electronic Theses and Dissertations

Ras is the most frequently activated oncogene found in human cancer, but its mechanisms of action remain only partially understood. Ras activates multiple signaling pathways in order to promote transformation, but can also exhibit a potent ability to induce growth arrest and death. NORE1A (RASSF5) is a direct Ras effector that acts as a tumor suppressor promoting senescence and apoptosis. Its expression is frequently lost in human tumors and its mechanism of action remains unclear. Here I show that NORE1A forms a direct, Ras regulated, complex with β-TrCP, the substrate recognition component of the SCF-β-TrCP ubiquitin ligase complex. NORE1A also …

Repsa Directed Assessment Of Native Cleavage Resistance Of Dna To Type Iis Restriction Endonucleases And Modification Of Repsa For High Temperature Application, Matthew Beyer

Master of Science in Chemical Sciences Theses

We have modified the combinatorial selection method Restriction Endonuclease Protection and Selection Assay (REPSA) to work in high temperature conditions for the discovery of new DNA-binding proteins in thermophiles (HT-REPSA). We utilized Thermus thermophilus (HB-8/ATCC 27634/DSM 579) as a test organism due to its amenable nature in a laboratory setting and current status as a model thermophilic organism. We used a TetR Family (TFR) transcription factor SbtR as the model protein for optimization of HT-REPSA protocols, as data had previously been obtained regarding SbtR physical characteristics and DNA-binding properties. REPSA was conducted until a cleavage resistant species arose after 7 …

Poly(N-Isopropylacrylamide) Hydrogels For Storage And Delivery Of Reagents To Paper-Based Analytical Devices, Haydn T. Mitchell, Spencer Schultz, Philip Costanzo, Andres W. Martinez

Chemistry and Biochemistry

The thermally responsive hydrogel N,N'-methylenebisacrylamide-cross-linked poly(N-isopropylacrylamide) (PNIPAM) was developed and evaluated as a reagent storage and delivery system for microfluidic paper-based analytical devices (microPADs). PNIPAM was shown to successfully deliver multiple solutions to microPADs in specific sequences or simultaneously in laminar-flow configuration and was found to be suitable for delivering four classes of reagents to the devices: Small molecules, enzymes, antibodies and DNA. PNIPAM was also able to successfully deliver a series of standard glucose solutions to microPADs equipped to perform a colorimetric glucose assay. The results of these tests were used to produce an external calibration …

Analytical Approaches In Investigating The Kinetics Of Water-Molecule Complexes In Tropospheric Reactions, William J. Keeton

Theses and Dissertations

Ozone is a heavily monitored pollutant. Ozone is not directly emitted into the atmosphere, but rather the product of chemical reactions. Ground level ozone occurs when nitrogen oxides (NOx) and volatile organic compounds (VOCs) react with each other in the presence of sunlight. The primary precursors of ozone are anthropogenically emitted, and as a result, tropospheric ozone has cost millions of dollars in damages and has hurt the health of countless people. This dissertation is a collection of work that aims to provide insight into atmospheric reactions that result in tropospheric ozone and the instrumentation to study such reactions. While …

Histone Deacetylase 6 (Hdac6) Is Critical For Tumor Cell Survival And Promotes The Pro-Survival Activity Of 14-3-3Ζ Viadeacetylation Of Lysines Within The14-3-3Ζ Binding Pocket, Jeffrey Benjamin Mortenson

Theses and Dissertations

Our understanding of non-histone acetylation as a means of cellular regulation is in its infancy. Using a mass spectrometry approach we identified acetylated lysine residues and monitored acetylation changes across the proteome as a consequence of metabolic stress (hypoxia). We observed changes in acetylation status of non-histone lysines in tumor cells. Through the use of small molecule inhibitors of histone deacetylase enzymes (HDACs) and siRNA screening identified HDAC6 as a pro-survival regulator of lysine acetylation during hypoxia. The phospho-binding protein 14-3-3ζ acts as a signaling hub controlling a network of interacting partners and oncogenic pathways. We show here that lysines …

Production And Characterization Of Islet Amyloid Polypeptide Using Recombinant Technology, Emilio Duverna

Graduate Theses and Dissertations

Islet amyloid polypeptide (IAPP) also called amylin is an amyloid-forming protein; IAPP is a proteinaceous hormone that comprises 37 amino acid residues. It is secreted along with insulin from the pancreatic β-cells to help it regulate the uptake and removal of glucose in the bloodstream. IAPP has been observed in the amyloid deposits found in pancreatic β-cells of most patients suffering from type II diabetes mellitus. This research project aims at producing recombinant amylin peptide. To achieve this goal, we used the pBAD plasmid vector which we introduced into Escherichia coli to express the peptide. Although the vector was successfully …

Using Peptoids To Build Robust, Efficient Microarray Systems, Dhaval Sunil Shah

Graduate Theses and Dissertations

Recent studies have shown microarrays to be indispensable for various biological applications, allowing for high-throughput processing and screening of biological samples such as RNA, DNA, proteins and peptides using a small sample volume (< 1 µL). Peptoids (poly-N-substituted glycine oligomers) can be used as a substitute for antibodies as capture molecules, as well as coatings for slides in antibody microarrays. The ease of synthesis of peptoids, high customizability with desired bioactivity, and speed of synthesis allows us to build a diagnostic system with a large dynamic range that can detect biomolecules from a minimal sample size. In this study, peptoid-based antibody mimics are designed to have both structural and functional features similar to those of antibodies, including a stable constant region (scaffolding) and a variable region for protein recognition. Peptoids previously screened via combinatorial library synthesis to be specific to bind Mdm-2 (mouse double minute 2 homolog) and GST (gluthathione S-transferase), have been synthesized. The protein recognition peptoids have been conjugated to PEG (polyethylene glycol) molecules with modified end groups; an amine group on one end that allows for immobilization and orientation on the slide, and an azide group on the other end that will allow for attachment to the peptoid through “click chemistry”. The number of capture molecules printed on the slides can be increased by making the available surface area of the slide larger via coating with microspheres. We have determined that partially water soluble peptoids that are also helical, can self-assemble into microspheres. Sequences have been developed that can consistently produce uniform microsphere coatings on slides that increase the overall surface area. A high surface area corresponds to a higher number of binding sites, and therefore a more sensitive system. The work done has shown that slides may be successfully coated in order to potentially improve the detection system.

Designing Fret Assays To Study Electrostatic Interactions Pertaining To The Binding Of Intrinsically Disordered Proteins, Ashley Ann Howard

Graduate Theses and Dissertations

Fibroblast growth factor receptor plays a major role in several biological processes. Without FGFR, a human cannot live. FGFR is involved in cell differentiation and wound healing. Of course, if FGFR signaling becomes unregulated, it causes severe distress in the body. Several cancers are contributed to high signaling levels, as well as developmental conditions like rickets and Kallmann’s syndrome. FGFR is thought to undergo an auto-inhibition (or self-regulatory) process in order to try to facilitate regulation. The exact method of this inhibition is currently unknown, but is proposed to involve the unstructured acid box region of FGFR. We developed a …

Rerouting Cellular Electron Flux To Increase The Rate Of Biological Methane Production, Jennie L. Catlett, Alicia M. Ortiz, Nicole R. Buan

Department of Biochemistry: Faculty Publications

Methanogens are anaerobic archaea that grow by producing methane, a gas that is both an efficient renewable fuel and a potent greenhouse gas. We observed that overexpression of the cyoplasmic heterodisulfide reductase enzyme HdrABC increased the rate of methane production from methanol by 30% without affecting the growth rate relative to the parent strain. Hdr enzymes are essential in all known methane-producing archaea. They function as the terminal oxidases in the methanogen electron transport system by reducing the coenzyme M (2-mercaptoethane sulfonate) and coenxyme B (7-mercaptoheptaonylthreonine sulfonate) heterodisulfide, CoM-S-S-CoB, to regenerate the thiol-coenzymes for reuse. In Methanosarcina acetivorans, HdrABC expression …

Ganoderma Lucidum Reduces Obesity In Mice By Modulating The Composition Of The Gut Microbiota, Chih-Jung Chang, Chuan-Sheng Lin, Chia-Chen Lu, Jan Martel, Yun-Fei Ko, David M. Ojcius, Shun-Fu Tseng, Tsung-Ru Wu, Yi-Yuan Margaret Chen, John Ding-E. Young, Hsin-Chih Lai

All Dugoni School of Dentistry Faculty Articles

Obesity is associated with low-grade chronic inflammation and intestinal dysbiosis. Ganoderma lucidum is a medicinal mushroom used in traditional Chinese medicine with putative anti-diabetic effects. Here, we show that a water extract of Ganoderma lucidum mycelium (WEGL) reduces body weight, inflammation and insulin resistance in mice fed a high-fat diet (HFD). Our data indicate that WEGL not only reverses HFD-induced gut dysbiosis—as indicated by the decreased Firmicutes-to-Bacteroidetes ratios and endotoxin-bearing Proteobacteria levels—but also maintains intestinal barrier integrity and reduces metabolic endotoxemia. The anti-obesity and microbiota-modulating effects are transmissible via horizontal faeces transfer from WEGL-treated mice to HFD-fed mice. We further …

Investigating The Links Between Ozone And Organic Aerosol Chemistry In A Biomass Burning Plume From A Prescribed Fire In California Chaparral, M. J. Alvarado, C. R. Lonsdale, Robert Yokelson, Sheryl Kashi Akagi, H. Coe, J. S. Craven, E. V. Fischer, G. R. Mcmeeking, J. H. Seinfeld, T. Soni, J. W. Taylor, D. R. Weise, C. E. Wold

Chemistry and Biochemistry Faculty Publications

Within minutes after emission, complex photochemistry in biomass burning smoke plumes can cause large changes in the concentrations of ozone (O3) and organic aerosol (OA). Being able to understand and simulate this rapid chemical evolution under a wide variety of conditions is a critical part of forecasting the impact of these fires on air quality, atmospheric composition, and climate. Here we use version 2.1 of the Aerosol Simulation Program (ASP) to simulate the evolution of O₃ and secondary organic aerosol (SOA) within a young biomass burning smoke plume from the Williams prescribed fire in chaparral, which was sampled over …

An Approach To An Agave Problem: The Bioremediation Of Agricultural Waste By Yeast Fermentations, Joshua Michael Jarodsky

Theses and Dissertations

The agricultural waste products created by industries, such as the Tequila manufacturing industry, create thousands of tons of waste material every year. This agricultural waste often becomes an environmental and ecological problem for fields and surrounding areas. The purpose of this study is to determine the potential of using the yeast Kluyveromyces marxianus 7-1 and Kluyveromyces marxianus 8-1 to ferment the leaf waste from the harvesting of the Agave tequilana as a form of bioremediation. Fermentations (anaerobic, under constant shaking, and at room temperature) were monitored for colony forming units, pH of fermentation broth, concentration of reducing sugars, and protein …

Cyclic Rhamnosylated Elongation Factor P Establishes Antibiotic Resistance In Pseudomonas Aeruginosa, Andrei Rajkovic, Sarah Erickson, Anne Witzky, Owen E. Branson, Jin Seo, Philip R. Gafken, Michael A. Frietas, Julian P. Whitelegge, Kym F. Faull, William Wiley Navarre, Andrew J. Darwin, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Elongation factor P (EF-P) is a ubiquitous bacterial protein that is required for the synthesis of poly-proline motifs during translation. In Escherichia coli and Salmonella enterica, the posttranslational β-lysylation of Lys34 by the PoxA protein is critical for EF-P activity. PoxA is absent from many bacterial species such as Pseudomonas aeruginosa, prompting a search for alternative EF-P posttranslation modification pathways. Structural analyses of P. aeruginosa EF-P revealed the attachment of a single cyclic rhamnose moiety to an Arg residue at a position equivalent to that at which β-Lys is attached to E. coli EF-P. Analysis of the genomes …

Expression, Localization, And Kinetic Characterization Of The Phospholipid Biosynthesis Enzyme Ctp: Phosphocholine Cytidylyltransferase From The Protozoan Parasite Leishmania Major, Justin Daniel Theodore Lange

Theses and Dissertations

The eukaryotic parasite Leishmania is the causative agent of the disease leishmaniasis. L. major is the most common of 21 species that causes visceral leishmaniasis in humans, and 30 that cause the same disease in other mammals. Visceral leishmaniasis causes fever, weight loss, and over a short amount of time, multiple organ failure, and has a 100% mortality rate within 2 years. This makes it the second largest parasitic killer in the world behind malaria. Over 90% of the worldâ??s cases of visceral leishmaniasis have been reported in underdeveloped countries of India, Bangladesh, Nepal, Sudan, Ethiopia and Brazil, with 500,000 …

Thyroxine Replacement In Patients On Parenteral Nutrition: A Clinical Conundrum, Abdul Rehman, Noor Ul-Ain Baloch, Muhammad Awais

Department of Biological & Biomedical Sciences

No abstract provided.