Digital Commons Network^™

Molecular Mechanisms Involved In Human Platelet Aggregation By Synergistic Interaction Of Platelet-Activating Factor And 5-Hydroxytryptamine, B H. Shah, H Rasheed, I H. Rahman, A H. Shariff, F L. Khan, H B. Rahman, S Hanif, S A. Saeed

Department of Biological & Biomedical Sciences

Our recent studies have shown that co-activation of Gq and Gi proteins by 5-hydroxytryptamine (5-HT) and adrenaline show synergism in human platelet aggregation. This study was conducted to examine the mechanism(s) of synergistic interaction of 5-HT and platelet activating factor (PAF) in human platelets. We show that PAF, but not 5-HT, increased platelet aggregation in a concentration-dependent manner. However, low concentrations of 5-HT (2 microM) potentiated platelet aggregation induced by subthreshold concentration of PAF (40 nM) indicating a synergistic interaction between the two agonists and this synergism was blocked by receptor antagonists to either 5-HT or PAF. 5-HT also potentiated …

Aerobic Biodegradation Of Methyl Tert-Butyl Ether By Aquifer Bacteria From Leaking Underground Storage Tank Sites, S. R. Kane, H. R. Beller, T. C. Legler, C. J. Koester, Holly C. Pinkart, R. U. Halden, A. M. Happel

All Faculty Scholarship for the College of the Sciences

The potential for aerobic methyl tert-butyl ether (MTBE) degradation was investigated with microcosms containing aquifer sediment and groundwater from four MTBE-contaminated sites characterized by oxygen-limited in situ conditions. MTBE depletion was observed for sediments from two sites (e.g., 4.5 mg/liter degraded in 15 days after a 4-day lag period), whereas no consumption of MTBE was observed for sediments from the other sites after 75 days. For sediments in which MTBE was consumed, 43 to 54% of added [U-¹⁴C]MTBE was mineralized to¹⁴CO₂. Molecular phylogenetic analyses of these sediments indicated the enrichment of species closely …

Formation Of Multistranded Structures From The Self-Assembly Of G-Rich Dna Oligomers By Various Sequence And Environmental Influences, Bryan D. Vi

Seton Hall University Dissertations and Theses (ETDs)

No abstract provided.

Cold-Temperature Adaptation Of Muscle Creatine Kinase From An Antartic Teleost (Chaenocephalus Aceratus), Paul Winnard Jr.

Electronic Theses and Dissertations

The white muscle of Chuenocephulus aceratus, an Antarctic teleost of the Channicthyidae family, has a compromised glycoiytic capacity and this fish cannot depend on glycolysis for rapid ATP generation For C. aceratus, creatine kinase (CK) and phosphocreatine (PCr) reserves comprise the metabolic pathway that may supplement and overcome this deficiency in energy transduction. Two conditions, low glycolytic capacity and evolution in a chronically cold habitat (-1.86°C), give us reason to believe that C. aceratus muscle CK (MMCK) has been subjected to strong selective pressure. Thus, the hypothesis of this thesis is that MMCK fiom C. aceratus white muscle …

Autoimmunity, Immune Deficiency And Cancer: Multiple Roles Of The Protein Tyrosine Phosphate Shp-1, Melissa J. Joliat

Electronic Theses and Dissertations

One of a large number of mutant mice used in immunological research, the "motheaten" mouse was the first model of a specific protein tyrosine phosphatase deficiency. Mice carrying one of two allelic mutations at the "motheaten" locus have severe systemic autoimmunity and immune dysfunction as a result of mutations in the hematopoietic-cell phosphatase (Hcph) gene, which encodes the protein tyrosine phosphatase SHP-1. Studies using "motheaten" (me/me) and "viable motheaten" (me^v/me^v) mice have increased the understanding of numerous signaling pathways in immune and hematopoietic cells. A number of studies on SHP-1 function …

Detection Of Insulin Receptor, Epidermal Growth Factor Receptor, And Interleukin-6 On Individual Mouse Embryos By Immuno-Polymerase Chain Reaction, Kun Xu

Electronic Theses and Dissertations

Three series of experiments were conducted to: 1) optimize the conditions for the production of pUC19 plasmid and of biotinylated pUC19 fragments; 2) optimize the conditions for the production of protein A-streptavidin chimera (chimeric protein); and 3) detect soluble antigens [bovine serum albumin (BSA) and interleukin-6 (IL-6)] and membrane-bound antigens [insulin receptor and epidermal growth factor receptor (EGFr)] of mouse embryos by immuno-polymerase chain reaction (I-PCR). The first experimental series, which included bacterial culture, chimeric protein purification, and chimeric protein functional experiments, was performed to investigate the effects of IPTG (isopropyl-β-D-thiogalactopyranoside) induction time and temperature, bacterial culture medium, and protein …

Ultrasound Based Quantitative Motion Measurement Using Speckle Size Estimation, James D. Hamilton, Larry Y. L. Mo, Gregory R. Bashford

Biomedical Imaging and Biosignal Analysis Laboratory

An ultrasound system determines the relative movement in a first direction (F1) of first matter, such as blood flow, and second matter, such as an artery wall, in a subject under study (S). A beam (B1) of ultrasound waves defining a plurality of beam positions (BP1 and BP2) and beam axes (A1 and A2) are moved in scan direction having components parallel to direction F1. First and second blocks of data representing the first and second matter, respectively, are generated. A processor (20) performs an estimation of speckle size on first data to obtain a first result, and performs analysis …

Transitions In Histone Acetylation Reveal Boundaries Of Three Separately Regulated Neighboring Loci, Michael D. Litt, Melanie A. Simpson, Felix Recillas-Targa, Marie-Noelle Prioleau, Gary Felsenfeld

Department of Biochemistry: Faculty Publications

We have studied developmentally regulated patterns of histone acetylation at high resolution across ~54 kb of DNA containing three independently regulated but neighboring genetic loci. These include a folate receptor gene, a 16 kb condensed chromatin region, the chicken β-globin domain and an adjacent olfactory receptor gene. Within these regions the relative levels of acetylation appear to fall into three classes. The condensed chromatin region maintains the lowest acetylation at every developmental stage. Genes that are inactive show similarly low levels, but activation results in a dramatic increase in acetylation. The highest levels of acetylation are seen at regulatory sites …

Map Kinases In Cynomolgus Monkey Sperm Hyperactivation, Emily Nicole Haynes

Biological Sciences Theses & Dissertations

Mammalian sperm exhibit characteristic motility patterns, termed hyperactivated (HA) motility, associated with completion of capacitation. In cynomolgus monkey (Macaca fascicularis) sperm, this HA motility is dependent in vitro upon the addition of exogenous cyclic nucleotide mediators, caffeine and dibutyryl-cyclic adenosine monophosphate (dbcAMP). Previous reports have shown protein tyrosine phosphorylation to be an integral component of this caffeine- and cAMP-stimulated HA motility. This study investigated the involvement of the mitogen-activated protein (MAP) kinase-signaling cascade. Semen specimens were collected in Talp-HEPES medium from proven breeders via electroejaculation. After washing, sperm were incubated in the presence and absence of the MAP …

Correlation Between Histone Lysine Methylation And Developmental Changes At The Chicken Β-Globin Locus, Michael D. Litt, Melanie A. Simpson, Miklos Gaszner, D. David Allis, Gary Felsenfeld

Department of Biochemistry: Faculty Publications

Methylation of histones at specific residues plays an important role in transcriptional regulation. Chromatin immunoprecipitation of dimethylated lysine 9 on histone H3 across 53 kilobases of the chicken β-globin locus during erythropoiesis shows an almost complete anticorrelation between regions of elevated lysine 9 methylation and acetylation. Lysine 9 is methylated most over constitutive condensed chromatin and developmentally inactive globin genes. In contrast, lysine 4 methylation of histone H3 correlates with H3 acetylation. These results lead us to propose a mechanism by which the insulator in the β-globin locus can protect the globin genes from being silenced by adjacent condensed chromatin.

Inferring And Testing Hypotheses Of Cladistic Character Dependence By Using Character Compatibility, F. Robin O’Keefe, Peter J. Wagner

Biological Sciences Faculty Research

The notion that two characters evolve independently is of interest for two reasons. First, theories of biological integration often predict that change in one character requires complementary change in another. Second, character independence is a basic assumption of most phylogenetic inference methods, and dependent characters might confound attempts at phylogenetic inference. Previously proposed tests of correlated character evolution require a model phylogeny and therefore assume that nonphylogenetic correlation has a negligible effect on initial tree construction. This paper develops “tree-free” methods for testing the independence of cladistic characters. These methods can test the character independence model as a hypothesis before …

Selenium Metabolism In Drosophila, F. Javier Martin-Romero, Gregory V. Kryukov, Alexei V. Lobanov, Bradley A. Carlson, Byeong Jae Lee, Vadim N. Gladyshev, Dolph L. Hatfield

Vadim Gladyshev Publications

Selenocysteine is a rare amino acid in protein that is encoded by UGA with the requirement of a downstream mRNA stem-loop structure, the selenocysteine insertion sequence element. To detect selenoproteins in Drosophila, the entire genome was analyzed with a novel program that searches for selenocysteine insertion sequence elements, followed by selenoprotein gene signature analyses. This computational screen and subsequent metabolic labeling with ⁷⁵Se and characterization of selenoprotein mRNA expression resulted in identification of three selenoproteins: selenophosphate synthetase 2 and novel G-rich and BthD selenoproteins that had no homology to known proteins. To assess a biological role for these …

Identification And Characterization Of A New Mammalian Glutaredoxin (Thioltransferase), Grx2, Vadim N. Gladyshev, Aimin Liu, Sergey V. Novoselov, Konstantin Krysan, Qi-An Sun, Valentin M. Kryukov, Gregory V. Kryukov, Marjorie F. Lou

Vadim Gladyshev Publications

A thiol/disulfide oxidoreductase component of the GSH system, glutaredoxin (Grx), is involved in the reduction of GSH-based mixed disulfides and participates in a variety of cellular redox pathways. A single cytosolic Grx (Grx1) was previously described in mammals. We now report identification and characterization of a second mammalian Grx, designated Grx2. Grx2 exhibited 36% identity with Grx1 and had a disulfide active center containing the Cys-Ser-Tyr-Cys motif. Grx2 was encoded in the genomes of mammals and birds and expressed in a variety of cell types. The gene for human Grx2 consisted of four exons and three introns, spanned 10 kilobase …

Quinic Acid-Mediated Induction Of Hypovirulence And A Hypovirulence-Associated Double-Stranded Rna In Rhizoctonia Solani, Chunyu Liu

Electronic Theses and Dissertations

This study is a part of a project focused on the relationship between dsRNA and hypovirulence in R. solani. Hypovirulence refers to a condition in which a pathogen has a drastically reduced ability to cause disease. Rhizoctonia solani is a soil-borne pathogen causing diseases in numerous plants. Phenyl acetic acid (PAA), a metabolite of phenylalanine, causes Rhizoctonia disease symptoms on potato in the absence of the pathogen itself. The amount of PAA produced by a hypovirulent isolate is 10% of that produced by virulent isolates. A 3.6 kb dsRNA (M2) has been shown to be associated with hypovirulence in …

Analysis Of Events Governing The Meiotic Division In Mouse Spermatocytes, Shannon Stewart Eaker

Doctoral Dissertations

The meiotic division is essential for successful gametogenesis. However, many events occurring during male and female meiotic development remain poorly understood. While it is known that chromosomes must pair, recombine, and segregate to form gametes, critical questions remain. How and when do these events occur with respect to each other? What mechanisms monitor their developmental success? Insight into these questions is provided in this dissertation, using the mouse spermatocyte as a model. The purpose of this work is to aid in the overall understanding of mammalian meiosis.

After an introduction into mammalian meiosis in Part I, a temporal order of …

Problem-Based Learning Variant: Transition Phase For A Large Institution, I Khan, A Fareed

Department of Biological & Biomedical Sciences

Objective: To compare students' test scores and perceptions of problem-based learning (PBL) and lecture-based learning (LBL) by applying a PBL-variant.
Method: For the transition from LBL to PBL, PBL was varied for one discipline only and for a large group of students. Two hundred forty nine second year medical students were taught a topic of Biochemistry by the LBL method and then 141 of these were taught another topic by the PBL-variant. At the conclusion of each topic an MCQ test was given. One week later a 9 item questionnaire was given to the 50 students now attending classes to …

Sir3-Dependent Assembly Of Supramolecular Chromatin Structures In Vitro, Philippe T. Georgel, Madeleine A. Palacios Debeer, Gregory Pietz, Catherine A. Fox, Jeffrey C. Hansen

Biological Sciences Faculty Research

Baculovirus-expressed recombinant Sir3p (rSir3p) has been purified to near homogeneity, and its binding to naked DNA, mononucleosomes, and nucleosomal arrays has been characterized in vitro. At stoichiometric levels rSir3p interacts with intact nucleosomal arrays, mononucleosomes, and naked DNA, as evidenced by formation of supershifted species on native agarose gels. Proteolytic removal of the core histone tail domains inhibits but does not completely abolish rSir3p binding to nucleosomal arrays. The linker DNA in the supershifted complexes remains freely accessible to restriction endonuclease digestion, suggesting that both the tail domains and nucleosomal DNA contribute to rSir3p–chromatin interactions. Together these data indicate …

Reversible Phosphorylation Of Photosynthetic Pep Carboxylase: Studies On C4-Leaf Pp2a And Recombinant Pepc-Kinase From Cam-Induced Mesembryanthemum Crystallinum, L. Dong, N. V. Ermolova, M. A. Cushman, T. Taybi, J. C. Cushman, Raymond Chollet

Department of Biochemistry: Faculty Publications

The activity and allosteric properties of plant PEPC are controlled posttranslationally by specific reversible phosphorylation of a strictly conserved Ser residue near the N-terminus. This up/down-modulation is catalyzed by a transcriptionally regulated, seemingly dedicated Ser/Thr kinase (PpcK) and an opposing Ser/Thr phosphatase (PP2A). We have now partially purified and characterized the native form of this largely “neglected” PP2A from maize leaves using various chromatographic and affinity matrices, and C₄ [³²P]PEPC as substrate (Dong et al., 2001, Planta [in press]). The results indicate that the C₄-leaf holoenzyme is analogous to yeast and mammalian PP2As in regards …

Unexpected Post-Translational Modifications And Novel Sequence-Structure Relationships Revealed By The Crystal Structure Of Chlamydomonas Reinhardtii Rubisco, T. C. Taylor, A. Backlund, R. J. Spreitzer, I. Andersson

Department of Biochemistry: Faculty Publications

The crystal structure of Rubisco from Chlamydomonas reinhardtii has been solved to 1.4 Å resolution. In addition to clear electron density for the carbamate on lys-201, the active site magnesium ion and the inhibitor 2-CABP, there was unexpected but unambiguous electron density for further specific post-translational modification. The N-terminus of the mature small subunit is methylated. The large subunit contains two methylcysteine residues (256 & 369) and two hydroxyproline residues (104 & 151). The small subunit from Chlamydomonas shows a structure more similar to that of the higher green plants than that from other algae of known structure. An extensive …

Can Negligible Levels Of Rubisco Activase, Be The Reason For Decreased Rubisco Activity Levels In Guard Cells?, S. Madhavan, P. Echtenkamp

Department of Biochemistry: Faculty Publications

Rubisco in guard cell chloroplasts has been a subject of controversy for decades. Several studies have unequivocally confirmed not only the presence of Rubisco in guard cells but also found it to be catalytically active. Based on a cell basis, the amount of Rubisco in guard cells is suggested to be in the range of 0.6 to 1.4 % whereas its activity is reported to be in the range of 0.35 to 0.5%, of a mesophyll cell. This low activity may be due a) to a low Rubisco content, and/or, b) to either the absence or an inactive functional activation …

Intrasteric Inhibition Of Atp Binding Is Not Required To Prevent Unregulated Autophosphorylation Or Signaling By The Insulin Receptor, Mark Frankel, Ararat J. Ablooglu, Joseph W. Leone, Elena Rusinova, J. B. A. Ross, Robert L. Heinrikson, Ronald A. Kohanski

Chemistry and Biochemistry Faculty Publications

Receptor tyrosine kinases may use intrasteric inhibition to suppress autophosphorylation prior to growth factor stimulation. To test this hypothesis we made an Asp1161Ala mutant in the activation loop that relieved intrasteric inhibition of the unphosphorylated insulin receptor (IR) and its recombinant cytoplasmic kinase domain (IRKD) without affecting the activated state. Solution studies with the unphosphorylated mutant IRKD demonstrated conformational changes and greater catalytic efficiency from a 10-fold increase in kcat and a 15-fold-lower K_{m ATP} although K_{m peptide} was unchanged. Kinetic parameters of the autophosphorylated mutant and wild-type kinase domains were virtually identical. The Asp1161Ala mutation increased the …

Regulation Of The Hcmv Ul98 Promoter By Cooperation Of The Cyclic Amp Response Element And Gamma Interferon Response Element, Tabmitha Patricia Young Jervey

Theses and Dissertations in Biomedical Sciences

The HCMV UL98 early alkaline exonuclease gene promoter was examined to determine the DNA sequences as well as viral and/or cellular proteins functional in the regulation of this early gene. To assess promoter activation, UL98 promoter sequences were first cloned upstream of the chloramphenicol acetyltransferase (CAT) reporter gene and cotransfected with expression plasmids which express the HCMV major immediate early (IE) proteins 1E72 and 1E86. To more specifically determine the importance of individual cis-acting elements in UL98 promoter activation, the promoter region underwent mutagenesis to delete or alter sequences. The variant promoters were again cloned into a reporter-CAT construct and …

Interactions Among Murine Cytomegalovirus Us22 Family Gene Products That Influence Viral Pathogenesis, Zaruhi Karabekian

Theses and Dissertations in Biomedical Sciences

Cytomegalovirus (CMV) is a complex, ubiquitous herpesvirus that is characterized by acute, chronic, and latent infections. Monocytes-macrophages are the key target cell type involved in pathogenesis, which is most effectively studied using the murine model of CMV infection. Previously three murine CMV (MCMV) genes (M139, M140, and M141) were identified to regulate viral expression in cultured macrophages and in mice. These genes are members of the US22 gene family with respect to HCMV homology. There is no function assigned to the proteins encoded by these genes. However, deletion of M139, M140, and M141 significantly curtails growth of MCMV in macrophages …

Positive Regulation Of Pka On Human Gonadotropin-Releasing Hormone (Hgnrh) Gene Expression In Human Placental Jeg-3 Cells, Zhaoyang Wen

Chemistry & Biochemistry Theses & Dissertations

Using the human placental choriocarcinoma JEG-3 cell line as an in vitro human placental model, we studied the mechanisms of the PKA positive regulation of the hGnRH gene expression in the human placenta. Studies in JEG3 cells showed that through the PKA catalytic subunit a, human GnRH upstream promoter activity was stimulated by PKA signaling pathway in a cAMP dependent mechanism. The sequence between —202 (Afl II) and —554 (BamH I) base pair in the human GnRH upstream promoter region appeared to be responsible for the PKA positive regulation of the gene expression. Furthermore, Western blot analysis demonstrated the involvement …

An Enzymatic Method For The Detection Of Human Serum Albumin, M U. Javed, S N. Waqar

Department of Biological & Biomedical Sciences

Albumin is the most abundant protein in human serum. A dye-binding method is commonly used in clinical laboratories for its estimation using different types of dyes. However, all these dye methods were interfered by a variety of compounds. Here we present a method for the detection of albumin in human serum and other biological fluids. The principle is based on the fact that lactate dehydrogenase isoenzyme-5 (LDH-5) binds specifically to Dextran-Blue (DB). Albumin inhibits the binding of LDH-5 with DB. Absence of LDH activity in DB fraction after gel filtration indicates the presence of albumin in sample and vice versa.

Complex Effects Arising In Smoke Plume Simulations Due To Inclusion Of Direct Emissions Of Oxygenated Organic Species From Biomass Combustion, Sherri A. Mason, Richard J. Field, Robert J. Yokelson, Michael A. Kochivar, Mark R. Tinsley, Darold E. Ward, Wei Min Hao

Chemistry and Biochemistry Faculty Publications

Oxygenated volatile organic species (oxygenates), including HCOOH, H₂CO, CH₃OH, HOCH₂CHO (hydroxyacetaldehyde), CH₃COOH, and C₆H₅OH, have recently been identified by Fourier transform infrared measurements as a significant component of the direct emissions from biomass combustion. These oxygenates have not generally been included in the hydrocarbon-based initial emission profiles used in previous photochemical simulations of biomass combustion smoke plumes. We explore the effects of oxygenates on this photochemistry by using an established initial emission hydrocarbon profile and comparing simulation results obtained both with and without addition of the above six …

Shape Imprinting Due To Variable Disulfide Bonds In Polyacrylamide Gels, Andrew B. Greytak, Alexander Y. Grosberg, Toyoichi Tanaka

Faculty Publications

Through the use of variable disulfide crosslinkers, we have created polyacrylamide gels whose shape can be altered after polymerization. N,N'-bisacryloylcystamine is incorporated as a crosslinker, along with a smaller amount of a permanent crosslinker. After polymerization, the disulfide bonds are cleaved into thiols through reduction. By reoxidizing the thiols with the gel held in a new macroscopic shape, a new set of disulfide bonds is formed, and the gel is forced to adopt the new shape. Retension of the new shape improves with greater distortion from the original shape, as well as with increased concentration of variable …

Selective Inhibition Of Selenocysteine Trna Maturation And Selenoprotein Synthesis In Transgenic Mice Expressing Isopentenyladenosine-Deficient Selenocysteine Trna, Mohamed E. Moustafa, Bradley A. Carlson, Muhammed A. El-Saadani, Gregory V. Kryukov, Qi-An Sun, John W. Harney, Kristina E. Hill, Gerald F. Combs, Lionel Feigenbaum, David B. Mansur, Raymond F. Burk, Marla J. Berry, Alan M. Diamond, Byeong Jae Lee, Vadim Gladyshev, Dolph A. Hatfield

Vadim Gladyshev Publications

Selenocysteine (Sec) tRNA (tRNA ^[Ser]Sec) serves as both the site of Sec biosynthesis and the adapter molecule for donation of this amino acid to protein. The consequences on selenoprotein biosynthesis of overexpressing either the wild type or a mutant tRNA ^[Ser]Sec lacking the modified base, isopentenyladenosine, in its anticodon loop were examined by introducing multiple copies of the corresponding tRNA ^[Ser]Sec genes into the mouse genome. Overexpression of wild-type tRNA ^[Ser]Sec did not affect selenoprotein synthesis. In contrast, the levels of numerous selenoproteins decreased in mice expressing isopentenyladenosine-deficient (i⁶A^-) tRNA ^[Ser]Sec in a protein- and …

Nonsymbiotic Hemoglobins In Rice Are Synthesized During Germination And In Differentiating Cell Types, E. J. H. Ross, L. Shearman, M. Mathiesen, Y.J. Zhou, R. Arredondo-Peter, Gautam Sarath, R. V. Klucas

Gautam Sarath Publications

Nonsymbiotic hemoglobins (ns-Hbs) previously have been found in monocots and dicots; however, very little is known about the tissue and cell type localization as well as the physiological function(s) of these oxygen-binding proteins. We report the immunodetection and immunolocalization of ns-Hbs in rice (Oryza sativa L.) by Western blotting and in situ confocal laser scanning techniques. Ns-Hbs were detected in soluble extracts of different tissues from the developing rice seedling by immunoblotting. Levels of ns-Hbs increased in the germinating seed for the first six days following imbibition and remained relatively constant thereafter. In contrast, ns-Hb levels decreased during leaf …

Chapter 13. The 15 Kda Selenoprotein (Sepl5): Functional Studies And A Role In Cancer Etiology, Vadim Gladyshev, Alan M. Diamond, Dolph L. Hatfield

Vadim Gladyshev Publications

The 15 kDa selenoprotein (Sepl5) is one of several recently identified selenoproteins. It contains a single selenocysteine residue in the middle of a 162-amino acid open reading frame and has no detectable homology to known proteins. The human Sepl5 gene spans 5 1 kb, has 5 exons and is located on chromosome 1 at position p3 1. The gene contains two single nucleotide polymorphisms in the 3'-untranslated region (3'-UTR) including one in the SECIS element, that are distributed differently between Caucasians and African Americans. Sep15 localizes to the endoplasmic reticulum where it is tightly bound to UDP-glucose:glycoprotein glucosyltransferase, a protein …