Molecular Biology Commons^™

Muc4/Muc4 Functions And Regulation In Cancer., Goldi Kozloski

Goldi A Kozloski

Genetic Effect Of The Dwarfing Genes On Some Culm Characteristics Associatcd With Lodging Resistance In Bread Wheat, Md. Mahbub Hasan

Md. Mahbub Hasan

Due to the challenge of screening traits related to lodging resistance under natural field conditions, selection for lodging resistant varieties in wheat breeding programs is difficult. The identification of easily measurable culm anatomical traits related to lodging resistance would simplify the selection process. The present study was conducted to determine the effect of dwarfing genes on culm anatomical traits related to lodging resistance in our of basal internode 1. Field and laboratory study was conducted in Shahjalal University of Science and Technology, Sylhet, Bangladesh with eight wheat genotypes having Rhr1, Rht2 dwarfing genes in them and a local land race …

Characterization And Functional Regulation Of Bioactive Peptides In Avian Macrophages And Heterophils, Lakshmi Kannan

Graduate Theses and Dissertations

Oligopeptides and low molecular weight polypeptides play central roles as effectors and signal transducers acting as hormones, neurotransmitters, growth factors, toxins, and antimicrobial factors that are important for the survival of the organism. Owing to the ubiquitous involvement of peptides in many key regulatory processes, we have been interested to identify native peptides in different cells and tissues and understand their functions. To conduct our studies, we used avian macrophages and heterophils as models of specialized cells which constitute central components of innate immunity. These studies involved (a) qualitative identification and characterization of the peptides associated with high intensity mass …

Trnas: Cellular Barcodes For Amino Acids, Ranat Banerjee, Shawn Chen, Kiley Dare, Marla Gilreath, Mette Praetorius-Ibba, Medha Raina, Noah M. Reynolds, Theresa E. Rogers, Hervé Roy, Srujana S. Yadavalli, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

The role of tRNA in translating the genetic code has received considerable attention over the last 50 years, and we now know in great detail how particular amino acids are specifically selected and brought to the ribosome in response to the corresponding mRNA codon. Over the same period, it has also become increasingly clear that the ribosome is not the only destination to which tRNAs deliver amino acids, with processes ranging from lipid modification to antibiotic biosynthesis all using aminoacyl‐tRNAs as substrates. Here we review examples of alternative functions for tRNA beyond translation, which together suggest that the role of …

Toc Receptor Dimerization Participates In The Initiation Of Membrane Translocation During Protein Import Into Chloroplasts, Jeonghwa Lee, Fei Wang Wang, Danny Schnell

Danny Schnell

The post-translational import of nucleus-encoded preproteins into chloroplasts occurs through multimeric translocons in the outer (Toc) and inner (Tic) membranes. The high fidelity of the protein import process is maintained by specific recognition of the transit peptide of preproteins by the coordinate activities of two homologous GTPase Toc receptors, Toc34 and Toc159. Structural and biochemical studies suggest that dimerization of the Toc receptors functions as a component of the mechanism to control access of preproteins to the membrane translocation channel of the translocon. We show that specific mutations that disrupted receptor dimerization in vitro reduced the rate of protein import …

Conformational Changes In Receptor Tyrosine Kinase Signaling: An Erbb Garden Of Delights., Goldi Kozloski

Goldi A Kozloski

Broad Range Amino Acid Specificity Of Rna-Dependent Lipid Remodelling By Multiple Peptide Resistance Factors, Hervé Roy, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacylphosphatidylglycerol synthases (aaPGSs) are multiple peptide resistance factors that transfer amino acids from aminoacyl-tRNAs to phosphatidylglycerol (PG) in the cytoplasmic membrane. Aminoacylation of PG is used by bacteria to decrease the net negative charge of the cell envelope, diminishing affinity for charged molecules and allowing for adaptation to environmental changes. Lys-PGS, which transfers lysine to PG, is essential for the virulence of certain pathogens, providing resistance to both host cationic antimicrobial peptides and therapeutic antibiotics. Ala-PGS was also recently described, but little is known about the possible activities of other members of the highly diverse aaPGS family of proteins. Systematic …

Self-Assembly And Disassembly Of The Snare Complex: Examined Using Circular Dichroism And Atomic Force Microscopy, Jeremy D. Cook, Won Jin Cho, Timothy L. Stemmler, Bhanu P. Jena

Biochemistry and Molecular Biology Faculty Publications

In this study, we report for the first time that both t-SNAREs and v-SNARE and their complexes in buffered suspension, exhibit defined peaks at CD signals of 208 and 222 nm wavelengths, consistent with a higher degree of helical secondary structure. Surprisingly, when incorporated in lipid membrane, both SNAREs and their complexes exhibit reduced folding. In presence of NSF-ATP, the SNARE complex disassembles, as reflected from the CD signals demonstrating elimination of α-helices within the structure.

The Cca Anticodon Specifies Separate Functions Inside And Outside Translation In Bacillus Cereus, Sandro F. Ataide, Theresa E. Rogers, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Bacillus cereus 14579 encodes two tRNAs with the CCA anticodon, tRNA^Trp and tRNA^Other. tRNA^Trp was separately aminoacylated by two enzymes, TrpRS1 and TrpRS2, which share only 34% similarity and display different catalytic capacities and specificities. TrpRS1 was 18-fold more proficient at aminoacylating tRNA^Trp with Trp, while TrpRS2 more efficiently utilizes the Trp analog 5-hydroxy Trp. tRNA^Other was not aminoacylated by either TrpRS but instead by the combined activity of LysRS1 and LysRS2, which recognized sequence elements absent from tRNA^Trp. Polysomes were found to contain tRNA^Trp, consistent with its role in …

Sinorhizobium Meliloti Cpdr1 Is Critical For Co-Ordinating Cell Cycle Progression And The Symbiotic Chronic Infection, H. Kobayashi, N. J. Kobayashi, Peter Chien, L. A. Simmons, G. C. Walker

Peter Chien

ATP-driven proteolysis plays a major role in regulating the bacterial cell cycle, development and stress responses. In the nitro -fixing symbiosis with host plants, Sinorhizobium meliloti undergoes a profound cellular differentiation, including endoreduplication of the ome. The regulatory mechanisms governing the alterations of the S. meliloti cell cycle in planta are largely unknown. Here, we report the characterization of two cpdR homologues, cpdR1 and cpdR2, of S. meliloti that encode single-domain response regulators. In Caulobacter crescentus, CpdR controls the polar localization of the ClpXP protease, thereby mediating the regulated proteolysis of key protein(s), such as CtrA, involved in cell cycle …

Geometric Build-Up Solutions For Protein Determination Via Distance Geometry, Robert Tucker Davis

Masters Theses & Specialist Projects

Proteins carry out an almost innumerable amount of biological processes that are absolutely necessary to life and as a result proteins and their structures are very often the objects of study in research. As such, this thesis will begin with a description of protein function and structure, followed by brief discussions of the two major experimental structure determination methods. Another problem that often arises in molecular modeling is referred to as the Molecular Distance Geometry Problem (MDGP). This problem seeks to find coordinates for the atoms of a protein or molecule when given only a set of pair-wise distances between …

Genomic Analysis Of The Basal Lineage Fungus Rhizopus Oryzae Reveals A Whole-Genome Duplication, Li-Jun Ma, Ashraf S. Ibrahim, Christopher Skory, Manfred G. Grabherr, Gertraud Burger, Margi Butler, Marek Elias, Alexander Idnurm, B. Franz Lang, Teruo Sone, Ayumi Abe, Sarah E. Calvo, Luis M. Corrochano, Reinhard Engels, Jianmin Fu, Wilhelm Hansberg, Jung-Mi Kim, Chinnappa D. Kodira, Michael J. Koehrsen, Bo Liu, Diego Miranda-Saavedra, Sinead O'Leary, Lucila Ortiz-Castellanos, Russell Poulter, Julio Rodriguez-Romero, José Ruiz-Herrera, Yao-Qing Shen, Qiandong Zeng, James Galagan, Bruce W. Birren, Christina A. Cuomo, Brian L. Wickes

Li-Jun Ma

Rhizopus oryzae is the primary cause of mucormycosis, an emerging, life-threatening infection characterized by rapid angioinvasive growth with an overall mortality rate that exceeds 50%. As a representative of the paraphyletic basal group of the fungal kingdom called “zygomycetes,” R. oryzae is also used as a model to study fungal evolution. Here we report the genome sequence of R. oryzae strain 99–880, isolated from a fatal case of mucormycosis. The highly repetitive 45.3 Mb genome assembly contains abundant transposable elements (TEs), comprising approximately 20% of the genome. We predicted 13,895 protein-coding genes not overlapping TEs, many of which are paralogous …

Fluorescence Correlation Spectroscopy Of Phosphatidylinositolspecific Phospholipase C Monitors The Interplay Of Substrate And Activator Lipid Binding, Mingming Pu, Mary F. Roberts, Anne Gershenson

Anne Gershenson

Phosphatidylinositol-specific phospholipase C (PI-PLC) enzymes simultaneously interact with the substrate, PI, and with non-substrate lipids such as phosphatidylcholine (PC). For Bacillus thuringiensis PI-PLC these interactions are synergistic with maximal catalytic activity observed at low to moderate mole fractions of PC (XPC) and maximal binding occurring at low mole fractions of anionic lipids. It has been proposed that residues in α helix B help modulate membrane binding and that dimerization on the membrane surface both increases affinity for PC and activates PI-PLC yielding the observed PI/PC synergy. Vesicle binding and activity measurements using a variety of PI-PLC mutants support many aspects …

Effect Of Chronic Treatment With A Cyclooxygenase Inhibitor On Reproductive Parameters In Male Rat, Sheikh A Saeed, Naheed Anwar, Khalid M Khan, Noorjehan Sarfraz

Department of Biological & Biomedical Sciences

BACKGROUND: Indomethacin is a member of non-steroidal anti-inflammatory drugs (NSAIDs) commonly used for treatment of gout, arthritis, and other inflammatory conditions. It has been shown to inhibit ovarian prostaglandins synthesis in mammals, birds, fish and reptiles. However, the effects of its chronic administration on male reproductive functions remain largely unknown. Using rat as a model, we studied the effect of chronic treatment with indomethacin on the male reproductive system.

METHODS: Testosterone was measured in the serum, testicular tissue, and testicular interstitial fluid by radioimmunoassay. Moreover, we also studied the direct effect of indomethacin in vitro on luteinizing hormone stimulated testosterone …

Inhibition Of Yeast Hexokinase By The Antimalarial Drug Artemisinin: Probing Mechanism Of Action With A Model Enzyme, Jennifer S. Spence

Biological Sciences Theses & Dissertations

A leading infectious cause of death, malaria threatens approximately half of the world's population, and drug-resistant strains of Plasmodium falciparum have created immense difficulty in chemotherapy of the disease. The artemisinin (ART) class of antimalarials may represent a powerful solution. In addition to their safety, effectiveness, and moderate cost, they are the only drugs in use for which there has been no widespread evidence of clinical resistance. The exact parasiticidal mechanism of ART is highly contested, but evidence suggests that protein alkylation may play a role in cytotoxicity. in vitro essays were performed using yeast hexokinase (HK) to demonstrate a …

Identifying Biomarkers For Resistance To Novel Cisplatin Analogues In Human Lung, Breast And Prostate Cancers, Becky Michelle Hess

UNLV Theses, Dissertations, Professional Papers, and Capstones

Cisplatin is a common therapeutic agent used in cancer treatment. Unfortunately, resistance to cisplatin in addition to severe side effects limits its use in cancer treatment. Two novel cisplatin analogues, 4DB and 4TB were shown to have varying cytotoxicity in lung, breast and prostate cancer cells. The hypothesis for this study states that the differences in 4DB and 4TB cytotoxicity among different tissue types is due to the type and efficiency of DNA repair mechanisms involved in response to these drugs.

To test the hypothesis, proteins involved in the rate limiting step of nucleotide excision repair (NER) and mismatch repair …

Characterization Of The Catalytic Mechanism Of The Carboxyltransferase Component Of Escherichia Coli Acetyl-Coa Carboxylase Leading To Rational Inhibitor Identification And The Development Of Enzymatic Mimics, Nabil K. Thalji

Honors Theses

No abstract provided.

Human Cerebral Neuropathology Of Type 2 Diabetes Mellitus, Peter T. Nelson, Charles D. Smith, Erin L. Abner, Frederick A. Schmitt, Stephen W. Scheff, Gregory J. Davis, Jeffrey N. Keller, Gregory A. Jicha, Daron Davis, Wang-Xia Wang, Adria Hartman, Douglas G. Katz, William R. Markesbery

Pathology and Laboratory Medicine Faculty Publications

The cerebral neuropathology of Type 2 diabetes (CNDM2) has not been positively defined. This review includes a description of CNDM2 research from before the ‘Pubmed Era’. Recent neuroimaging studies have focused on cerebrovascular and white matter pathology. These and prior studies about cerebrovascular histopathology in diabetes are reviewed. Evidence is also described for and against the link between CNDM2 and Alzheimer's disease pathogenesis. To study this matter directly, we evaluated data from University of Kentucky Alzheimer's Disease Center (UK ADC) patients recruited while non-demented and followed longitudinally. Of patients who had come to autopsy (N = 234), 139 met …

A Toc159 Import Receptor Mutant, Defective In Hydrolysis Of Gtp, Supports Preprotein Import Into Chloroplasts, Birgit Agne, Sibylle Infanger, Fei Wang, Valère Hofstetter, Gwendoline Rahim, Meryll Martin, Dong Wook Lee, Inhwan Hwang, Danny Schnell, Felix Kessler

Danny Schnell

The heterotrimeric Toc core complex of the chloroplast protein import apparatus contains two GTPases, Toc159 and Toc34, together with the protein-conducting channel Toc75. Toc159 and Toc34 are exposed at the chloroplast surface and function in preprotein recognition. Together, they have been shown to facilitate the import of photosynthetic proteins into chloroplasts in Arabidopsis. Consequently, the ppi2 mutant lacking atToc159 has a non-photosynthetic albino phenotype. Previous mutations in the conserved G1 and G3 GTPase motifs abolished the function of Toc159 in vivo by disrupting targeting of the receptor to chloroplasts. Here, we demonstrate that a mutant in a conserved G1 lysine …

Resampling And Editing Of Mischarged Trna Prior To Translation Elongation, Jiqiang Ling, Byung Ran So, Srujana S. Yadavalli, Hervé Roy, Shinichiro Shoji, Kurt Fredrick, Karin Musier-Forsyth, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Faithful translation of the genetic code depends on the GTPase EF-Tu delivering correctly charged aminoacyl-tRNAs to the ribosome for pairing with cognate codons. The accurate coupling of cognate amino acids and tRNAs by the aminoacyl-tRNA synthetases is achieved through a combination of substrate specificity and product editing. Once released by aminoacyl-tRNA synthetases, both cognate and near-cognate aminoacyl-tRNAs were considered to be committed to ribosomal protein synthesis through their association with EF-Tu. Here we show instead that aminoacyl-tRNAs in ternary complex with EF-Tu•GTP can readily dissociate and rebind to aminoacyl-tRNA synthetases. For mischarged species, this allows resampling by the product editing …

Adaptation Of The Bacterial Membrane To Changing Environments Using Aminoacylated Phospholipids, Hervé Roy, Kiley Dare, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Fine‐tuning of the biophysical properties of biological membranes is essential for adaptation of cells to changing environments. For instance, to lower the negative charge of the lipid bilayer, certain bacteria add lysine to phosphatidylglycerol (PG) converting the net negative charge of PG (−1) to a net positive charge in Lys‐PG (+1). Reducing the net negative charge of the bacterial cell wall is a common strategy used by bacteria to resist cationic antimicrobial peptides (CAMPs) secreted by other microbes or produced by the innate immune system of a host organism. The article by Klein et al. in the current issue of …

Dopamine Controls Locomotion By Modulating The Activity Of The Cholinergic Motor Neurons In C. Elegans, Andrew T. Allen

Masters Theses 1911 - February 2014

Dopamine is an important neurotransmitter in the brain, where it plays a regulatory role in the coordination of movement and cognition by acting through two classes of G protein-coupled receptors to modulate synaptic activity. In addition, it has been shown these two receptor classes can exhibit synergistic or antagonistic effects on neurotransmission. However, while the pharmacology of the mammalian dopamine receptors have been characterized in some detail, less is known about the molecular pathways that act downstream of the receptors. As in mammals, the soil nematode Caenorhabditis elegans uses two classes of dopamine receptors to control neural activity and thus …

Functional Analysis Of Receptor-Like Kinases In Pollen-Pistil Interactions In Arabidopsis Thaliana, Mini Aggarwal

Masters Theses 1911 - February 2014

Rac/Rop GTPases are molecular switches in plants that control the growth of polarized cells such as pollen tubes and root hairs, differentiation, development, actin dynamics, production of reactive oxygen species (ROS) and disease resistance. These small GTPases are activated by guanine nucleotide exchange factors (GEFs) that replace GDP for GTP and are referred to as RopGEFs in plants. To identify upstream components of the RopGEF regulated signaling pathways, GEF1 from Arabidopsis thaliana was used as a bait to screen a seedling cDNA library in a yeast two-hybrid system. This yielded members of a small family of the Catharanthus roseus Receptor-like …

Quaternary Dynamics And Plasticity Underlie Small Heat Shock Protein Chaperone Function, Florian Stengel, Andrew J. Baldwin, Alexander J. Painter, Nomalie Jaya, Eman Basha, Lewis E. Kay, Elizabeth Vierling, Carol V. Robinson, Justin L. P. Benesch

Elizabeth Vierling

Small Heat Shock Proteins (sHSPs) are a diverse family of molecular chaperones that prevent protein aggregation by binding clients destabilized during cellular stress. Here we probe the architecture and dynamics of complexes formed between an oligomeric sHSP and client by employing unique mass spectrometry strategies. We observe over 300 different stoichiometries of interaction, demonstrating that an ensemble of structures underlies the protection these chaperones confer to unfolding clients. This astonishing heterogeneity not only makes the system quite distinct in behavior to ATP-dependent chaperones, but also renders it intractable by conventional structural biology approaches. We find that thermally regulated quaternary dynamics …

The Signal Peptide Peptidase Is Required For Pollen Function In Arabidopsis, Danny Schnell, S. Han, L. Green

Danny Schnell

The Signal Peptide Peptidases (SPP) are members of the Intramembrane Cleaving Proteases, which are involved in an array of protein-processing and intracellular signaling events in animals. Arabidopsis (Arabidopsis thaliana) has six genes encoding SPP-like proteins, the physiological functions of which are unknown. As a first step in defining the roles of the SPPs in plants, we examined the distribution and activities of Arabidopsis SPP (AtSPP; accession no. At2g03120), the SPP-like gene with the highest degree of similarity to human SPP. The protease is expressed at low levels throughout the plant, with the highest levels in emerging leaves, roots, and floral …

Substrate Binding Site Flexibility Of The Small Heat Shock Protein Molecular Chaperones, Nomalie Jaya, Victor Garcia, Elizabeth Vierling

Elizabeth Vierling

Small heat shock proteins (sHSPs) serve as a first line of defense against stress-induced cell damage by binding and maintaining denaturing proteins in a folding-competent state. In contrast to the well-defined substrate binding regions of ATP-dependent chaperones, interactions between sHSPs and substrates are poorly understood. Defining substrate-binding sites of sHSPs is key to understanding their cellular functions and to harnessing their aggregation-prevention properties for controlling damage due to stress and disease. We incorporated a photoactivatable cross-linker at 32 positions throughout a well-characterized sHSP, dodecameric PsHsp18.1 from pea, and identified direct interaction sites between sHSPs and substrates. Model substrates firefly luciferase …

Cholesterol Exposure At The Membrane Surface Is Necessary And Sufficient To Trigger Perfringolysin O Binding, John J. Flanagan, Rodney K. Tweten, Arthur E. Johnson, Alejandro P. Heuck

Alejandro P. Heuck

Perfringolysin O (PFO) is the prototype for the cholesterol-dependent cytolysins, a family of bacterial pore-forming toxins that act on eukaryotic membranes. The pore-forming mechanism of PFO exhibits an absolute requirement for membrane cholesterol, but the complex interplay between the structural arrangement of the PFO C-terminal domain and the distribution of cholesterol in the target membrane is poorly understood. Herein we show that PFO binding to the bilayer and the initiation of the sequence of events that culminate in the formation of a transmembrane pore depend on the availability of free cholesterol at the membrane surface, while changes in the acyl …

Hmgb1 Mediates Endogenous Tlr2 Activation And Brain Tumor Regression, James Curtin, Naiyou Liu, Marianela Candolfi, Weidong Xiong, Hikmat Assi, Kader Yagiz, Matthew Edwards, Kathrin Michelsen, Kurt Kroeger, Chunyan Liu, Akm Ghulam Muhammad, Mary Clark, Moshe Arditi, Begonya Comin-Anduix, Antoni Ribas, Pedro Lowenstein, Maria Castro

Articles

BACKGROUND: Glioblastoma multiforme (GBM) is the most aggressive primary brain tumor that carries a 5-y survival rate of 5%. Attempts at eliciting a clinically relevant anti-GBM immune response in brain tumor patients have met with limited success, which is due to brain immune privilege, tumor immune evasion, and a paucity of dendritic cells (DCs) within the central nervous system. Herein we uncovered a novel pathway for the activation of an effective anti-GBM immune response mediated by high-mobility-group box 1 (HMGB1), an alarmin protein released from dying tumor cells, which acts as an endogenous ligand for Toll-like receptor 2 (TLR2) signaling …

Analysis Of N-Linked Oligosaccharides Of Prostate-Specific Antigen And Prostatic Acid Phosphatase In Prostatic Fluids, Krista Yaudes White

Theses and Dissertations in Biomedical Sciences

Presently, prostate cancer is the most common cancer afflicting men in the United States, with serum PSA being the "gold standard" protein biomarker used in the clinic for detecting and diagnosing prostate cancer. Nonetheless, serum PSA levels can also be elevated in non-cancerous conditions as well, such as benign prostatic hyperplasia (BPH). Due to this overlap, many unnecessary biopsies and radical prostatectomies occur, leading to patient distress. Despite recent advances to clinical assays which consider other clinical parameters, there is still a great need for improved clinical detection methodologies for prostate cancer, including improved biomarkers. Therefore, this research project aims …

Regulation Of Intracellular Calcium Concentration By Nanosecond Pulsed Electric Fields, Shaka S. Scarlett, Jody A. White, Peter F. Blackmore, Karl H. Schoenbach, Juergen Kolb

Bioelectrics Publications

Changes in [Ca²⁺]_i response of individual Jurkat cells to nanosecond pulsed electric fields (nsPEFs) of 60 ns and field strengths of 25, 50, and 100 kV/cm were investigated. The magnitude of the nsPEF-induced rise in [Ca²⁺]_i was dependent on the electric field strength. With 25 and 50 kV/cm, the [Ca²⁺]_i response was due to the release of Ca²⁺ from intracellular stores and occurred in less than 18 ms. With 100 kV/cm, the increase in [Ca²⁺]_i was due to both internal release and to influx across the plasma …