Biochemistry Commons^™

Studies Toward The Synthesis Of Lyconadin A And Cranomycin, Brad M. Loertscher

Theses and Dissertations

Lyconadin A is a pentacyclic Lycopodium alkaloid isolated from the club moss Lycopodium companatum with anticancer activity. Our approach sought to incorporate a 7-exo–6-exo acyl radical cyclization cascade to access the bicyclo[5.4.0]undecane framework of lyconadin A. Our studies created methodology for the synthesis of 5-alkyl and 3,5-dialkyl-6-carbomethoxy-2-pyridones and sterically demanding epoxide substrates. These epoxide substrates underwent an unanticipated Payne rearrangement.Cranomycin is a potent antibiotic with antiprotozoal activity. Structurally it is a cyclopentane ring system with substitution at each carbon in the ring. Another interesting structural aspect is the existence of three contiguous quaternary stereocenters including two tertiary alcohols and a …

Developmental Signaling Requires Inwardly Rectifying K+ Channels In Drosophila Melanogaster, Giri Raj Dahal

Theses and Dissertations

Inwardly rectifying potassium (IRK/Kir) channels regulate intracellular K+ concentrations and membrane potential. Disruption of Kir2.1 causes dominantly inherited Andersen Tawil Syndrome (ATS). ATS patients suffer from cardiac arrhythmias, periodic paralysis, and cognitive impairment. These symptoms are consistent with current understanding of the role of ion channels in muscle cells and neurons. However, ATS symptoms also include craniofacial and digital deformities such as cleft palate, dental defects, wide set eyes, low set ears, and crooked or fused digits. These developmental defects were not consistent with current understanding of developmental signaling or previously described roles for ion channels. We found that phenotypes …

The Role Of Smpb In Licensing Tmrna Entry Into Stalled Ribosomes, Mickey R. Miller

Theses and Dissertations

Ribosomes translate the genetic information contained in mRNAs into protein by linking together amino acids with the help of aminoacyl-tRNAs. In bacteria, protein synthesis stalls when the ribosome reaches the 3'-end of truncated mRNA transcripts lacking a stop codon. Trans-translation is a conserved bacterial quality control process that rescues stalled ribosomes. Transfer-messenger RNA (tmRNA) and its protein partner SmpB mimic a tRNA by entering the A site of the ribosome and accepting the growing peptide chain. The ribosome releases the truncated mRNA and resumes translation on the tmRNA template. The open reading frame found on tmRNA encodes a peptide tag …

An Ethnobotanical Approach To Finding Antimicrobial Compounds In Wooly Blue Curls (Trichostema Lanatum) Using A Kirby-Bauer Disc Diffusion Assay, Matthew C. Fleming, P. Matthew Joyner

Featured Research

Plants can be an important source of creativity and production of new drugs. In this study, extracts of wooly blue curls (Trichostema lanatum) were made using DMSO and tested for antimicrobial activity on a panel of bacteria commonly found in separate ecological niches. Wooly blue curls (WBC) was chosen due to its being recorded as a strong disinfectant by the Chumash people. It was found that WBC does exhibit antimicrobial activity against gram positive bacteria and not against gram negative bacteria. However, gram negative bacteria with reduced drug efflux function became susceptible to the WBC extract.

Immunomodulatory Activity Of Sambucus Mexicana And Trichostema Lanatum On Lps Stimulated Raw 264.7 Macrophage Cells, Victoria Hester, P. Matthew Joyner

Featured Research

Chumash medicinal plants Sambucus mexicana (Mexican elderberry) and Trichostema lanatum (woolly blue curls) were tested for immunomodulatory activity. Anti-inflammatory effects were determined by treating LPS induced RAW 264.7 macrophage cells with plant extracts and measuring the levels of cytokines: tumor necrosis factor alpha (TNF-alpha) and interleukin 10 (IL-10). We hypothesized that both plants would exert immunomodulatory activity by reducing the pro-inflammatory production of TNF-alpha or by promoting M2 polarization with a concurrent increase in IL-10 production. At concentration 0.01 mg/mL woolly blue curls and Mexican elderberry demonstrated anti-inflammatory activity by reducing the concentration of TNF-alpha in vitro, while levels of …

Purification And Characterization Of Enzymes From Yeast: An Extended Undergraduate Laboratory Sequence For Large Classes, Kelly E. Johanson, Terry J. Watt, Neil R. Mcintyre, Marleesa Thompson

Faculty and Staff Publications

Providing a project-based experience in an undergraduate biochemistry laboratory class can be complex with large class sizes and limited resources. We have designed a 6-week curriculum during which students purify and characterize the enzymes invertase and phosphatase from bakers yeast. Purification is performed in two stages via ethanol precipitation and anion exchange chromatography, and students perform both direct and coupled enzyme assays. By completion of the experimental series, students are able to identify which enzymes they have purified and have obtained kinetic parameters for one. This experimental series requires minimal instructor preparation time, is cost effective, and works with multiple …

Role Of Nucleation In Cortical Microtubule Array Organization: Variations On A Theme, Erica A. Fishel, Ram Dixit

Biology Faculty Research

The interphase cortical microtubules (CMTs) of plant cells form strikingly ordered arrays in the absence of a dedicated microtubule-organizing center. Considerable research effort has focused on activities such as bundling and severing that occur after CMT nucleation and are thought to be important for generating and maintaining ordered arrays. In this review, we focus on how nucleation affects CMT array organization. The bulk of CMTs are initiated from γ-tubulin-containing nucleation complexes localized to the lateral walls of pre-existing CMTs. These CMTs grow either at an acute angle or parallel to the pre-existing CMT. Although the impact of microtubule-dependent nucleation is …

Carcinogenic Adducts Induce Distinct Dna Polymerase Binding Orientations, Kyle B. Vrtis, Radoslaw P. Markiewicz, Louis J. Romano, David Rueda

Chemistry Faculty Research Publications

DNA polymerases must accurately replicate DNA to maintain genome integrity. Carcinogenic adducts, such as 2-aminofluorene (AF) and N-acetyl-2-aminofluorene (AAF), covalently bind DNA bases and promote mutagenesis near the adduct site. The mechanism by which carcinogenic adducts inhibit DNA synthesis and cause mutagenesis remains unclear. Here, we measure interactions between a DNA polymerase and carcinogenic DNA adducts in real-time by single-molecule fluorescence. We find the degree to which an adduct affects polymerase binding to the DNA depends on the adduct location with respect to the primer terminus, the adduct structure and the nucleotides present in the solution. Not only do the …

Investigation Of Water-Molecule Complexes And Their Catalytic Effect On Important Atmospheric Reactions, Taylor Scott Cline

Theses and Dissertations

This dissertation is a collection of works that investigates issues related to environmental chemistry. The first portion of this research explores the role of water vapor on the kinetics of important atmospheric reactions. Work is presented on the self-reaction of β-hydroxyethyl peroxy radical (β-HEP) and the catalytic increase in reaction rate by water vapor. β-HEP serves as a model system for investigating the possible role of water vapor in perturbing the kinetics and product branching ratio of atmospheric reactions of other alkyl peroxy radicals. The self-reaction rate coefficient of β-HEP was investigated between 276-296 K with 1.0 × 10^15 to …

Influence Of Media Composition On The Growth Rate Of Chlorella Vulgaris And Scenedesmus Acutus Utilized For Co2 Mitigation, Czarena L. Crofcheck, Xinyi E, Aubrey Shea, Michael D. Montross, Mark Crocker, Rodney Andrews

Biosystems and Agricultural Engineering Faculty Publications

Atmospheric carbon dioxide levels have increased since the industrial revolution due to increasing combustion of fossil fuels. One possible CO₂ mitigation strategy is the use of microalgae for mitigation of CO₂. This paper focuses on the influence of media composition on the growth rate of two microalgae strains, Chlorella vulgaris and Scenedesmus actus. A KNO₃ based medium was found to work better for Chlorella, while a urea based equivalent worked better for Scenedesmus. The urea based media investigated here resulted in growth similar to that found with previously reported KNO₃ based media. …

Miniaturized Electrostatic Ion Beam Trap Mass Analyzer, Junting Wang

Theses and Dissertations

The electrostatic ion beam trap (EIBT) was designed by D. Zajfman during the previous decade. This ion trap combines many properties of the Fourier-transform ion cyclotron resonance (FTICR) mass analyzer and time-of-flight (TOF) mass analyzer. There are several advantages for the electrostatic ion beam trap. First, large mass-to-charge particles in an electrostatic field could be easier to analyze. Second, there is a folded flight path, which could make the mass analyzer smaller compared to conventional TOF mass analyzer. This principle of operation of this ion trap is analogous to an optical resonator. The ions are trapped in a voltage valley …

Identification And Differentiation Of Tier 1 Bacterial Agents Using Gas Chromatography-Mass Spectrometry, Dan Li

Theses and Dissertations

A simple method was developed for detection and differentiation of five Tier 1 bacterial agents, including Bacillus anthracis, Francisella tularensis, Yersinia pestis, Burkholderia pseudomallei and Burkholderia mallei as well as their closely related near neighbors by gas chromatography-mass spectrometry (GC-MS). Generally, different classes of compounds can be used as biomarkers for biowarfare agent detection, including nucleic acids (i.e., DNA or RNA), proteins (i.e., antibodies), carbohydrates (i.e., sugars), lipopolysaccharides, lipids (i.e., fatty acids) and small molecules. One-step thermochemolysis (TCM) was developed to provide GC-MS detectable biomarker signatures, including sugars, fatty acids and small molecules. Solid phase micro-extraction (SPME) was used for …

Bacterial Spores Remain Viable After Electrospray Charging And Desolvation, Sara Nielson Pratt

Theses and Dissertations

The electrospray survivability of B. subtilis spores and E. coli was tested in atmospheric mobility experiments. E. coli did not survive electrospray charging and desolvation, but B. subtilis did. Experimental conditions ensured that any surviving bacteria were charged, desolvated, and de-agglomerated. B. subtilis was also found to survive both positive and negative electrospray and subsequent introduction into vacuum conditions. Attempts were made to measure the charge distribution of viable B. subtilis spores using electrostatic deflection. From those experiments, it was found that either the spores do not become highly charged under the electrospray conditions used or only spores in a …

Diversity Of Plant Methionine Sulfoxide Reductases B And Evolution Of A Form Specific For Free Methionine Sulfoxide, Dung Tien Le, Lionel Tarrago, Yasuko Watanabe, Alaattin Kaya, Byung Cheon Lee, Uyen Tran, Rie Nishiyama, Dmitri E. Fomenko, Vadim N. Gladyshev, Lam-Son Phan Tran

Department of Biochemistry: Faculty Publications

Methionine can be reversibly oxidized to methionine sulfoxide (MetO) under physiological conditions. Organisms evolved two distinct methionine sulfoxide reductase families (MSRA & MSRB) to repair oxidized methionine residues. We found that 5 MSRB genes exist in the soybean genome, including GmMSRB1 and two segmentally duplicated gene pairs (GmMSRB2 and GmMSRB5, GmMSRB3 and GmMSRB4). GmMSRB2 and GmMSRB4 proteins showed MSRB activity toward protein-based MetO with either DTT or thioredoxin (TRX) as reductants, whereas GmMSRB1 was active only with DTT. GmMSRB2 had a typical MSRB mechanism with Cys121 and Cys 68 as catalytic and resolving residues, respectively. Surprisingly, this enzyme also possessed …

Raman Spectroscopic Analysis Of Human Skin Tissue Sections Ex-Vivo: Evaluation Of The Effects Of Tissue Processing And Dewaxing, Syed Mehmood Ali, Franck Bonnier, Ali Tfayli, Helen Lambkin, Kathleen Flynn, Vincent Mcdonagh, Claragh Healy, Thomas Lee, Fiona Lyng, Hugh Byrne

Articles

Raman spectroscopy coupled with K-means clustering analysis (KMCA) is employed to elucidate the biochemical structure of human skin tissue sections, and the effects of tissue processing. Both hand and thigh sections of human cadavers were analysed in their unprocessed and formalin fixed paraffin processed (FFPP) and subsequently dewaxed forms. In unprocessed sections, KMCA reveals clear differentiation of the stratum corneum, intermediate underlying epithelium and dermal layers for sections from both anatomical sites. The stratum corneum is seen to be relatively rich in lipidic content; the spectrum of the subjacent layers is strongly influenced by the presence of melanin, while that …

Purification And Characterization Of Oxidation-Resistant Ribonuclease Inhibitor Variants, Alec W. Uebersohn

Lawrence University Honors Projects

Ribonuclease inhibitor (RI) is an intracellular mammalian protein which binds vertebrate-specific ribonucleases; this interaction is one of the tightest non-covalent interactions yet discovered. The biological activity of RI is poorly understood, but it is thought to regulate the biological functions of ribonucleases, which include initiating blood vessel growth, maintaining neuron viability, attacking pathogens, and mediating cell stress responses. RI is also involved in pathways unrelated to ribonucleases, including interactions with Drosha and PTEN, an anti-tumor protein.

One of the defining characteristics of RI is its oxidation sensitivity, a result of its unusually high cysteine content. The oxidation of RI is …

Isolation, Characterization And Synthesis Of Asthma Inducing Fungal Glycolipid And Analytical Method Development For Novel Antimicrobial Peptide Mimics, Vinod Chaudhary

Theses and Dissertations

NKT cells are an important part of human immune system and recognize a specific set of antigens called glycolipids. Only a handful of "natural" NKT cell antigens are known till date. Although NKT cells play a protective role against pathogenic organisms, imbalances in NKT cell functions are implicated in many diseases including asthma. Allergic asthma, a Th2 driven inflammation of airways, is primarily caused by inhalation of environmental allergens. In the last decade, inhaled allergen Aspergillus fumigatus has been under scrutiny for the presence of NKT cell antigens that might trigger asthma. We successfully isolated, characterized and synthesized a "natural" …

Validation Of Antibodies Used To Study Hypoxia Inducible Factors In Two Species Of Fundulus, Jenna D. Hill

LSU New Orleans Theses and Dissertations

Hypoxia inducible factors (HIFs) are transcription factors and the master regulators of oxygen-dependent gene expression in animals. The focus of this thesis is the distribution of HIF protein in tissues of the fish Fundulus heteroclitus and F. grandis, two widespread species that occur in naturally hypoxic waters. Polyclonal antibodies against HIF-1α, HIF-2α, and HIF-3α were tested on proteins made in vitro and on extracts made from several tissues of normoxic and hypoxic fish. Antibodies against HIF-1α and 3α bound specifically to full length protein made in vitro, and produced bands on western blots of nuclear extracts of near …

Characterization Of Cpey And Cpez Mutants In Fremyella Diplosiphon Strain Utex 481, Christina M. Kronfel

LSU New Orleans Theses and Dissertations

Phycoerythrin (PE) present on the outer phycobilisome (PBS) rods in Fremyella diplosiphon contains covalently attached phycoerythrobilin (PEB) chromophores for efficient photosynthetic light capture. Chromophore ligation on phycobiliprotein subunits occurs through bilin lyase catalyzed reactions. The cpeY and cpeZ genes in F. diplosiphon were shown to attach PEB on alph-82 of PE. To better understand the individual functions of cpeY and cpeZ in native cyanobacteria, we characterized PBS and PE purified from cpeY and cpeZ deletion mutants and compared them with wild type (WT). Both cpeY and cpeZ mutants generated much less PE than WT as well as assembling much less …

Regulation Of The Target Of Rapamycin Signaling Pathway In Saccharomyces Cerevisiae, Tammy Pracheil

LSU New Orleans Theses and Dissertations

An integrative, biochemical, genetic, and molecular biology approach utilizing gene manipulation, gene knock outs, plasmid based protein expression, and in vivo protein localization of fluorescence tagged proteins was employed to determine the function of an essential protein, Lst8, in TORC1 and TORC2 signaling and a previously uncharacterized complex, the Far3-7-8-9-10-11 complex (Far complex) in the budding yeast, Saccharomyces cerevisiae. Mutations in SAC7 and FAR11 suppressed lethality of both lst8 and tor2-21 mutations but not TORC1 inactivation, suggesting that the essential function of Lst8 is linked only to TORC2.

Far11, a component of a six-member complex, was found to interact …