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Pyrrolysine And Selenocysteine Use Dissimilar Decoding Strategies, Yan Zhang, Pavel V. Baranov, John F. Atkins, Vadim N. Gladyshev

Vadim Gladyshev Publications

Selenocysteine (Sec) and pyrrolysine (Pyl) are known as the 21st and 22nd amino acids in protein. Both are encoded by codons that normally function as stop signals. Sec specification by UGA codons requires the presence of a cis-acting selenocysteine insertion sequence (SECIS) element. Similarly, it is thought that Pyl is inserted by UAG codons with the help of a putative pyrrolysine insertion sequence (PYLIS) element. Herein, we analyzed the occurrence of Pyl-utilizing organisms, Pyl-associated genes, and Pyl-containing proteins. The Pyl trait is restricted to several microbes, and only one organism has both Pyl and Sec. We found that methanogenic …

Association Between Archaeal Prolyl- And Leucyl-Trna Synthetases Enhances TrnaPro Aminoacylation, Mette Praetorius-Ibba, Theresa E. Rogers, Rachel Samson, Zvi Kelman, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacyl-tRNA synthetase-containing complexes have been identified in different eukaryotes, and their existence has also been suggested in some Archaea. To investigate interactions involving aminoacyl-tRNA synthetases in Archaea, we undertook a yeast two-hybrid screen for interactions between Methanothermobacter thermautotrophicus proteins using prolyl-tRNA synthetase (ProRS) as the bait. Interacting proteins identified included components of methanogenesis, protein-modifying factors, and leucyl-tRNA synthetase (LeuRS). The association of ProRS with LeuRS was confirmed in vitro by native gel electrophoresis and size exclusion chromatography. Determination of the steady-state kinetics of tRNA^Pro charging showed that the catalytic efficiency (k_cat/K_m) of ProRS increased 5-fold …

Preparation, Structural Characterization, And Dynamic Properties Investigation Of Permalloy Antidot Arrays, Andriy Vovk, Leszek M. Malkinski, Scott L. Whittenburg, Charles O'Connor, Jin-Seung Jung, Suk-Hong Min

Chemistry and Biochemistry Faculty Publications

Regular nanosized structures are considered to be promising materials for magnetic information storage media with high density of information. Recently attention was paid to static and dynamic magnetic properties arising from dimensional confinement in such nanostructures. Here we present an investigation of permalloy antidot arrays of different thicknesses. Thin permalloyfilms of thickness ranging from were deposited on nanoporous membranes with a pore size of . It was found that additional ferromagnetic resonance peaks appear for film thicknesses below , while films with larger thicknesses show resonanceproperties similar to continuous films. A comparison between the filmsdeposited onto Si wafers and porous …

Alternative Assay Techniques For Protein Kinase A, Justin Brink

Honors Capstones

Capstone submitted as a graduation requirement for the BSU Honors Program.

Primary Sex Reversal In Female Betta Splendens Following Treatments With Temperature And Trenbolone Acetate., Tracy S. Smith

Electronic Theses and Dissertations

Sex reversal in fish can occur as a result of disturbances in natural conditions or laboratory manipulations. Trenbolone Acetate (TBA), a synthetic anabolic-androgenic steroid used mainly in the cattle industry, was applied to female Betta splendens, a sexually dimorphic fish. Previously, females were analyzed for secondary sex characteristics after treatment with TBA, increased temperature, or a combination of both. This study focused on primary sex characteristics by histological examination. Gonads were surgically removed and prepared following basic histological techniques. Following H&E staining, sections were observed for signs of ovarian regression, as well as signs of testicular formation. The presence …

The Epigenomic Viewpoint On Cellular Differentiation Of Myeloid Progenitor Cells As It Pertains To Leukemogenesis, Vincent E. Sollars

Biochemistry and Microbiology

The new millennium has brought with it a surge of research in the field of epigenetics. This has included advances in our understanding of stem cell characteristics and mechanisms of commitment to cell lineages prior to differentiation. The nature of stem cells is similar to that of malignant cells in that they have unlimited self-renewal and protection from apoptosis, leading researchers to suspect that stem cells are the target of oncogenesis. This review will explore the idea of how epigenetic control of gene expression may contribute to mechanisms controlling differentiation of myeloid progenitor cells and its importance to our understanding …

Potential Microcin Production By Helicobacter Pylori Hpm8, Sarah R. Null

All-Inclusive List of Electronic Theses and Dissertations

Helicobacter pylori undergoes significant mutation during chronic infection. The organism adapts to its specific host allowing a selective advantage even after repeated exposure to new competing strains. Genetic drift does not fully explain the ability of some strains to outcompete pre-existing strains or newly introduced strains. The hypothesis behind this research is that H. pylori HPM8 produces a peptide encoded on a plasmid, pHPM8, which has antibacterial activity against other H. pylori strains. Reverse Transcriptase Polymerase Chain Reaction (RT-PCR) was used to determine the stage of the growth cycle the peptide is being expressed. RT-PCR demonstrated peptide expression throughout the …

Myod Targets Chromatin Remodeling Complexes To The Myogenin Locus Prior To Forming A Stable Dna-Bound Complex, Ivana L. De La Serna, Yasuyuki Ohkawa, Charlotte A. Berkes, Donald A. Bergstrom, Caroline S. Dacwag, Stephen J. Tapscott Fred Hutchinson Cancer Research Center, Anthony N. Imbalzano

Biology Faculty Publications

The activation of muscle-specific gene expression requires the coordinated action of muscle regulatory proteins and chromatin-remodeling enzymes. Microarray analysis performed in the presence or absence of a dominant-negative BRG1 ATPase demonstrated that approximately one-third of MyoD-induced genes were highly dependent on SWI/SNF enzymes. To understand the mechanism of activation, we performed chromatin immunoprecipitations analyzing the myogenin promoter. We found that H4 hyperacetylation preceded Brg1 binding in a MyoD-dependent manner but that MyoD binding occurred subsequent to H4 modification and Brg1 interaction. In the absence of functional SWI/SNF enzymes, muscle regulatory proteins did not bind to the myogenin promoter, thereby providing …

In Vivo Characterization Of The Integrin Beta(3) As A Receptor For Hantaan Virus Cellular Entry, Jin-Won Song, Ki-Joon Song, Luck-Ju Baek, Blaise Frost

Chemistry Faculty Publications

No abstract provided.

Biomolecule Detection Via Target Mediated Nanoparticle Aggregation And Dielectrophoretic Impedance Measurement, Philip J. Costanzo, Enzhu Liang, Timothy E. Patten, Scott D. Collins, Rosemary L. Smith

Chemistry and Biochemistry

A new biosensing system is described that is based on the aggregation of nanoparticles by a target biological molecule and dielectrophoretic impedance measurement of these aggregates. The aggregation process was verified within a microchannel via fluorescence microscopy, demonstrating that this process can be used in a real time sensor application. Positive dielectrophoresis is employed to capture the nanoparticle aggregates at the edge of thin film electrodes, where their presence is detected either by optical imaging via fluorescence microscopy or by measuring the change in electrical impedance between adjacent electrodes. The electrical detection mechanism demonstrates the potential for this method as …

Selenocysteine Insertion Directed By The 3’-Utr Secis Element In Escherichia Coli, Dan Su, Yehua Li, Vadim Gladyshev

Vadim Gladyshev Publications

Co-translational insertion of selenocysteine (Sec) into proteins in response to UGA codons is directed by selenocysteine insertion sequence (SECIS) elements. In known bacterial selenoprotein genes, SECIS elements are located in the coding regions immediately downstream of UGA codons. Here, we report that a distant SECIS element can also function in Sec insertion in bacteria provided that it is spatially close to the UGA codon. We expressed a mammalian phospholipid hydroperoxide glutathione peroxidase in Escherichia coli from a construct in which a natural E.coli SECIS element was located in the 3’-untranslated region (3’-UTR) and adjacent to a sequence complementary to the …

The Effect Of Rooibos Tea (Aspalathus Linearis) On The Survival And Transformation Of C3h-Bt1 Mouse Fibroblasts, Krista M. Beach, Paul J. Smith

Honors Capstones

Capstone submitted as a graduation requirement for the BSU Honors Program.

Nematode Selenoproteome: The Use Of The Selenocysteine Insertion System To Decode One Codon In An Animal Genome?, Kalin Taskov, Charles Chapple, Gregory V. Kryukov, Sergi Castenello, Alexei V. Lobanov, Konstantin V. Korotkov, Roderic Guigo, Vadim Gladyshev

Vadim Gladyshev Publications

Selenocysteine (Sec) is co-translationally inserted into selenoproteins in response to codon UGA with the help of the selenocysteine insertion sequence (SECIS) element.The number of selenoproteinsinanimals varies, with humans having 25 and mice having 24 selenoproteins. To date, however, only one selenoprotein, thioredoxin reductase, has been detected in Caenorhabditis elegans, and this enzyme contains only one Sec. Here, we characterize the selenoproteomes of C.elegans and Caenorhabditis briggsae with three independent algorithms,one searching for pairs of homologous nematode SECIS elements, another searching for Cys- or Sec-containing homologs of potential nematode selenoprotein genes and the third identifying Sec-containing homologs of annotated nematode …

Identification And Expansion Of Pancreatic Stem/Progenitor Cells., You-Qing Zhang, Marcie Kritzik, Nora Sarvetnick

Journal Articles: Regenerative Medicine

Pancreatic islet transplantation represents an attractive approach for the treatment of diabetes. However, the limited availability of donor islets has largely hampered this approach. In this respect, the use of alternative sources of islets such as the ex vivo expansion and differentiation of functional endocrine cells for treating diabetes has become the major focus of diabetes research. Adult pancreatic stem cells /progenitor cells have yet to be recognized because limited markers exist for their identification. While the pancreas has the capacity to regenerate under certain circumstances, questions where adult pancreatic stem/progenitor cells are localized, how they are regulated, and even …

Defensin In The Ticks Dermacentor Variabilis And Ixodes Scapularis, Shane Michael Ceraul

Theses and Dissertations in Biomedical Sciences

This dissertation presents the identification, molecular structure and expression of defensin, an antimicrobial peptide, in the ticks, Dermacentor variabilis and Ixodes scapularis. A 624 bp cDNA molecule for defensin was cloned and sequenced from hemocytes, fat body and midgut of unstimulated D. variabilis ticks. Defensin peptide was detected only in the hemocyte lysates. These results suggest that the defensin from D. variabilis is synthesized and stored in the hemocytes, although the role of other tissues is not excluded.

The ability of the juvenile stage ticks to express the defensin gene was confirmed. Detection of the defensin transcript in all …

The Antitumor Agent, Arglabin-Dma, Preferentially Induces Apoptosis In Human Colon Tumor Cells, Sung Wook Kwon

Theses and Dissertations in Biomedical Sciences

Arglabin-DMA, an analog of farnesyl pyrophosphate (FPP), reportedly inhibits farnesyltransferase (FTase) directly by competitively blocking the binding of Ras protein and its posttranslational modification, as suggested in previous studies. But, the mechanisms by which Arglabin-DMA inhibits tumor growth in vivo and in vitro are still relatively poorly characterized. To determine the mechanism by which this drug inhibits tumor growth, the effects of Arglabin-DMA in two human colon tumor cell lines (mutant K-ras HCT 116 and wild-type ras HT-29) were explored on cell proliferation, apoptosis, and cell cycle kinetics in vitro. In cell viability studies, we showed that Arglabin-DMA …

Mechanisms Of Cell Death Initiated In Herpes Simplex Virus Thymidine Kinase Expressing Colon Tumor Cells Treated With Ganciclovir And Ucn-01, Christina Elizabeth Ahn

Theses and Dissertations in Biomedical Sciences

Metastatic colon carcinoma is the second leading cause of death from malignancy in the United States, and development of more effective treatments is essential. Heterologous expression of Herpes Simplex Virus Thymidine Kinase (HSVtk) in combination with the prodrug, ganciclovir (GCV), has shown great promise for the genetic therapy of many cancers, but most patients have had only a partial or minimal response to the therapy. After screening a panel of two drug combinations, our laboratory has shown that the combination of GCV and the protein kinase inhibitor UCN-01 (7-hydroxystaurosporine) enhances tumor cell death more effectively than either drug alone. However …

The Glucose And Insulin Responsiveness Of The Rat Glucose-6-Phosphate Dehydrogenase Promoter, Daryl Arkwright-Keeler

Dissertations

The regulation of expression of some enzymes involved in glucose homeostasis and fat metabolism requires both insulin and glucose to elicit an effect, while the regulation of others requires only one. Glucose-6-phosphate dehydrogenase (G6PDH), an important enzyme involved in glucose homeostasis, has been shown to be transcriptionally regulated by carbohydrate feeding in rat liver, a diet regime that increases the levels of both glucose and insulin. Our lab had previously shown that insulin induces expression of G6PDH, thus we wanted to determine if glucose also induced G6PDH expression and if so, are the glucose and insulin responses mediated through identical …

Bufadienolides In The Chemical Defenses Of The Toads, Bufo Americanus And Bufo Fowleri, Catherine E. Merovich

Dissertations

I investigated the steroidal chemical defenses (bufadienolides) of Bufo americanus and Bufo fowleri. By the nature of their complex, biphasic life cycles, toads, like other amphibians are important components of aquatic and terrestrial habitats and are prey to numerous invertebrates and vertebrates. Bufadienolides are presumed to be important anti-predatory compounds although much of their chemical ecology is poorly understood. I investigated (1) ontogenetic variation in bufadienolides, (2) bufadienolides from adult parotoid secretion, (3) effectiveness of bufadienolides against a terrestrial predator, and (4) effectiveness of bufadienolides against an aquatic predator. I hypothesized that B. americanuswould have a more extensive …

Alterations In Uterine And Placental Sodium Pump Abundance May Contribute To The Onset Of Mouse Labor, Carlos Jacob Vance

Theses and Dissertations

Objective: Reductions in sodium pump (SP) abundance can give rise to increases in contractile force in uterine and vascular smooth muscle as well as an increased secretion in secretory cells, including potentially those of the placenta. To determine whether the mouse might serve as a model for human pregnancy in terms of the SP and to determine whether changes in SP abundance anticipate or follow labor, we studied pregnant mice over the final trimester of their pregnancy.

Study Design: C57Bl6 dams (n=46) were bred and studied during their pregnancy. Animals (n=4) were sacrificed at specific gestational time points. Other mice …

The Microbial Selenoproteome Of The Sargasso Sea, Yan Zhang, Dmitri E. Fomenko, Vadim Gladyshev

Vadim Gladyshev Publications

Background: Selenocysteine (Sec) is a rare amino acid which occurs in proteins in major domains of life. It is encoded by TGA, which also serves as the signal for termination of translation, precluding identification of selenoprotein genes by available annotation tools. Information on full sets of selenoproteins (selenoproteomes) is essential for understanding the biology of selenium. Herein, we characterized the selenoproteome of the largest microbial sequence dataset, the Sargasso Sea environmental genome project.
Results: We identified 310 selenoprotein genes that clustered into 25 families, including 101 new selenoprotein genes that belonged to 15 families. Most of these proteins were predicted …

Ion Transmission In The First Vacuum Stage Of An Inductively Coupled Plasma Mass Spectrometer, Jeffrey Harris Macedone

Theses and Dissertations

The inductively coupled plasma mass spectrometer (ICP-MS) is the instrument of choice for trace and ultra-trace elemental analysis. However, the ICP-MS suffers from matrix effects. Matrix effects occur when instrument response varies as the composition of the sample matrix is changed. Matrix effects, or non-spectroscopic interferences, limit the accuracy of routine analysis. Identification of the sources of matrix effects provide a basis for reducing or eliminating them. As inaccuracies in the ICP-MS are more severe than those in the ICP atomic emission spectrometer, the problem may be due, at least in part, to the vacuum interface used to couple the …

Nucleic Acid Sequencing Using Charge-Switch Nucleotides, John G. K. Williams, Gregory R. Bashford

Biomedical Imaging and Biosignal Analysis Laboratory

The present invention provides compounds, methods and systems for sequencing nucleic acid using single molecule detection. Using labeled NPs that exhibit charge-switching behavior, single-molecule DNA sequencing in a microchannel sorting system is realized. In operation, sequencing products are detected enabling real-time sequencing as successive detectable moieties flow through a detection channel. By electrically sorting charged molecules, the cleaved product molecules are detected in isolation without interference from unincorporated NPs and without illuminating the polymerase-DNA complex.

Clean, High-Yield Preparation Of S,S- And R,S- Amino Acid Isosteres, Aslam A. Malik, Todd E. Clement, Hasan Palandoken, James Robinson Iii, Joy A. Stringer

Chemistry and Biochemistry

The present invention provides compounds and methods that can be used to convert the intermediate halomethyl ketones (HMKs), e.g., chloromethyl ketones, to the corresponding S,S- and R,S-diastereomers. More particularly, the present invention provides: (1) reduction methods; (2) inversion methods; and (3) methods involving the epoxidation of alkenes. Using the various methods of the present invention, the R,S-epoxide and the intermediary compounds can be prepared reliably, in high yields and in high purity.

Um34 In Selenocysteine Trna Is Required For The Expression Of Stress-Related Selenoproteins In Mammals, Bradley A. Carlson, Xue-Ming Xu, Vadim N. Gladyshev, Dolph L. Hatfield

Vadim Gladyshev Publications

Selenium is an essential micronutrient in the diet of mammals and has many health benefits. Selenium-containing proteins are responsible for most, if not all, of these benefits. This element is incorporated into protein as selenocysteine (Sec), the 21^st amino acid in the genetic code. There are two species of Sec tRNA in mammalian cells that differ by a single 2’-O-hydroxymethyl group on the ribosyl moiety at position 34 (Um34). The relationship between this modification and selenoprotein synthesis was examined in mice in which the wild type Sec tRNA gene was replaced with a mutant Sec tRNA transgene incapable of …

Variability In Lipid Profile Before And After Coronary Artery Bypass Grafting (Cabg), M Perwaiz Iqbal, Hasanat Mohammad Sharif, Naseema Mehboobali

Department of Biological & Biomedical Sciences

Objective: To investigate changes in total cholesterol, low density lipoprotein (LDL) cholesterol, high density lipoprotein (HDL) cholesterol, and triglycerides in serum of Pakistani patients before, immediately after and 5 days post CABG.
Method: Serum samples from 31 consecutive Pakistani angina patients undergoing CABG at the Aga Khan University Hospital were analyzed for total cholesterol, LDL cholesterol, HDL cholesterol and triglycerides using kit methods.
Results: Immediately after CABG, there is a significant decline in the mean levels of serum cholesterol, LDL cholesterol, HDL cholesterol and triglycerides. However, 5 days post CABG, there is a significant increase in the concentrations of total …

Arabidopsis Atspl14, A Plant-Specific Sbp-Domain Transcription Factor, Participates In Plant Development And Sensitivity To Fumonisin B1, Julie M. Stone, Xinwen Liang, Emily R. Nekl, Justin J. Stiers

Department of Biochemistry: Faculty Publications

The recessive Arabidopsis thaliana fumonisin B1-resistant (fbr6) mutant was identified by its ability to survive in the presence of a programmed cell death (PCD)-inducing fungal toxin FB1. The fbr6 mutant also displays altered plant architecture in the absence of FB1, most notably elongated petioles and enhanced leaf margin serration. These phenotypes are a result of a T-DNA insertion in the SQUAMOSA promoter binding protein (SBP) domain gene, AtSPL14. AtSPL14 encodes a plant-specific protein with features characteristic of a transcriptional regulator, including a nuclear localization signal sequence, a plant-specific DNA binding domain (the SBP box), and a protein …

Selective Rescue Of Selenoprotein Expression In Mice Lacking A Highly Specialized Methyl Group In Selenocysteine Trna, Bradley A. Carlson, Xue-Ming Xu, Vadim N. Gladyshev, Dolph L. Hatfield

Vadim Gladyshev Publications

Selenocysteine (Sec) is the 21st amino acid in the genetic code. Its tRNA is variably methylated on the 2’-Ohydroxyl site of the ribosyl moiety at position 34 (Um34). Herein, we identified a role of Um34 in regulating the expression of some, but not all, selenoproteins. A strain of knock-out transgenic mice was generated, wherein the Sec tRNA gene was replaced with either wild type or mutant Sec tRNA transgenes. The mutant transgene yielded a tRNA that lacked two base modifications, N⁶- isopentenyladenosine at position 37 (i⁶A37) and Um34. Several selenoproteins, including glutathione peroxidases 1 and 3, …

5-Ht1b Receptor-Mediated Presynaptic Inhibition Of Gaba Release In The Suprachiasmatic Nucleus, Jayne R. Bramley, Patricia J. Sollars, Gary E. Pickard, F. Edward Dudek

School of Veterinary and Biomedical Sciences: Faculty Publications

The suprachiasmatic nucleus (SCN) receives a dense serotonergic innervation that modulates photic input to the SCN via serotonin 1B (5-HT_1B) presynaptic receptors on retinal glutamatergic terminals. However, the majority of 5-HT_1B binding sites in the SCN are located on nonretinal terminals and most axonal terminals in the SCN are GABAergic. We therefore tested the hypothesis that 5-HT_1B receptors might also be located on SCN GABAergic terminals by examining the effects of the highly selective 5-HT_1B receptor agonist CP-93,129 on SCN miniature inhibitory postsynaptic currents (mIPSCs). Whole cell patch-clamp recordings of mIPSCs were obtained from rat …

Temperature Dependence Of The Double Layer Capacitance For The Restricted Primitive Model Of An Electrolyte Solution From A Density Functional Approach, Douglas Henderson, J. Reszko-Zygmunt, Stefan Sokolowski, Dezso Boda

Faculty Publications

We apply a different version of the density functional theory, given by Pizio, Patrykiejew, and Sokolowski [J. Chem. Phys. 121, 11957 (2004)], for a nonuniform restricted primitive model of an electrolyte solution to evaluate the temperature dependence of the capacitance of an electric double layer. We show that this theory is capable of reproducing the computer simulation data at a quantitative level. In particular, the reversal of the temperature dependence of the capacitance at low temperatures is predicted. This phenomenon has been difficult to predict from theory. Further, this theory also leads to an accurate description of the double layer …