Life Sciences Commons^™

Singlet-Oxygen-Mediated Inactivation Of Acetylcholinesterase: A Comparison Of Purified Enzyme In Solution And Enzyme Bound To K562 Leukemia Cells, Gail D. Deadwyler

Dissertations

No abstract provided.

Molecular Cloning And Analysis Of The Promoter For Rat Hepatic Neutral Cholesterol Ester Hydrolase, Ramesh Natarajan

Theses and Dissertations

Neutral cholesterol ester hydrolase (CEH) is a key enzyme in regulating hepatic free cholesterol. Using the CEH specific cDNA sequence in the 5'-untranslated region as a primer, 1.3 kb of sequence upstream of the ATG initiation codon was amplified and cloned. Primer extension analysis with total RNA from rat primary hepatocytes identified a transcription initiation site, 60 bases upstream from the initiation codon. No typical TATA-box sequences were found upstream from the transcription start site. However, a consensus GC-box, which can bind the positive transcription factor SP1, was found 35 bases upstream from the transcription start site. In addition the …

Small Miracles: Propagation Efforts Using Plants In The Family Hamamelidaceae, Martha Sasser Joiner

College of Graduate Studies: Theses & Dissertations (1964–2006)

Attempts were made to propagate five species in the family Hamamelidaceae by softwood cuttings or seeds. Local populations of witch-hazel Hamamelis virginiana L. and sweetgum Liquidambar styraciflua L. supplied cuttings and seeds of these native American species. Cultivated plants of dwarf witch-alder Fothergilla gardenii Murray were used initially for cuttings. Later, three native populations of this species were located and cuttings were obtained from them. Also, for seed propagation studies, seeds of F. gardenii were obtained from the North Carolina Botanical Garden and seeds were obtained from cultivated plants of two Asian species, winter hazel Corylopsis sinensis Hemsley and Chinese …

Acute And Chronic Exposure To Lead: Cellular And Behavioral Effects On The Pond Snail Helisoma Trivolvis, Virginia T. Bennett

College of Graduate Studies: Theses & Dissertations (1964–2006)

The focus of this study is on the effects of lead on Helisoma trivolvis, a pulmonate pond snail. Many species of molluscs are being used as bioindicators of environmental toxins. To provide a background for the reader, a brief history and description of the Phylum Mollusca (at the class level) is presented in Chapter 1. A second chapter (Ch. 2) is concerned with lead and its effects. A synopsis of past and present studies that show the interaction between lead and molluscs is given in Chapter 3, then two experimental chapters follow. The first experimental chapter (Ch. 4) is …

Nest Box Use, Reproductive Success, And Nestling Growth Of American Kestrels (Falco Sparverius) Nesting Along The Coastal Plain Of Georgia, Walter Roy Shuford

College of Graduate Studies: Theses & Dissertations (1964–2006)

In an attempt to increase breeding populations of American Kestrels (Falco sparverius) in Georgia, 402 nest boxes were erected in the Coastal Plain of Georgia beginning in 1994 through the spring of 1996. Nest box occupancy averaged 8% in 1996 and 9.7% in 1997, two fold the average in 1995. Occupancy at Fort Gordon military base averaged 27.5% (28 of 102) in 1996 and 29 4% (30 of 102) in 1997 Nest box use in all other areas averaged 13% (4 of 300) in 1996 and increased to 3% (9 of 300) in 1997. Clutch sizes for nests …

Abnormalities In Post-Translational Processing Of Platelet Rap 1b In Niddm: A Possible Cause Of Platelet Hyperactivity And Cardiovascular Disease In Diabetes, Elizabeth Ann Hall

Theses and Dissertations in Biomedical Sciences

Post-translational processing is critical for the appropriate subcellular localization and function of platelet G-proteins. The majority of the platelet responses to agonists are mediated through specific receptor/G-protein complexes. Therefore, G-protein activity is central to "normal" platelet activity (i.e. aggregation). We have shown that Simvastatin, the in vivo inhibitor of HMG CoA Reductase and therefore isoprenoid synthesis, inhibits the post-translational processing of specific platelet G-proteins and alters platelet responses to agonists. These results show the importance of post-translational processing of G-proteins to platelet activity. Altered post-translational processing of specific G-proteins may explain platelet hyperactivity and the increased incidence of cardiovascular disease …

The Measurement Of Angiotensin-Converting Enzyme In Human Serum, Andrew P. Kenny

Theses

The measurement of angiotensin converting enzyme using the substrate FAPGG was investigated with a view to optimising its performance. It was found that a 100 mnol/l triethanolamine-HCl buffer gave a significantly higher absorbance change then the existing TRIS-HCl in the method. As this buffer has not been previously described in the measurement of ACE it was decided to introduce this buffer into a kinetic procedure and evaluate the modified method. The reference interval for the triethanolamine Method was determined by measuring the ACE activity on 200 serum from blood donors and a significant difference (P < 0.05) in the SACE activity of male and female donors was observed.

The modified method was found …

Genetic Analysis Of Chlorophyll Biosynthesis, David Bollivar

Scholarship

No abstract provided.

A Study Of Biochemical And Haematological Variables With Performance In Showjumping Horses, Darren B. Simmons

Theses : Honours

This honours project examined the biochemical and haematological changes induced by exercise in the showjumping horse to determine whether a relationship exists between these values and performance. To complete this study 7 geldings and 1 stallion who were considered competition fit by their trainers were subjected to 2 forms of exercise. The first phase of this study involved an incremental exercise test performed at the Byford City Council horse training facility. The horses were cantered/galloped around a 1300m all weather training track at velocities of 6, 8, 10, and 12 m/sec for a duration of 2 minutes for each workload. …

Phosphorylation Within The Amino-Terminal Acidic Domain I Of The Phosphoprotein Of Vesicular Stomatitis Virus Is Required For Transcription But Not For Replication, Asit K. Pattnaik, Leroy Hwang, Tong Le, Nathan Englund, Manjula Mathur, Tapas Das, Amiya Banerjee

School of Veterinary and Biomedical Sciences: Faculty Publications

Phosphorylation by casein kinase II at three specific residues (S-60, T-62, and S-64) within the acidic domain I of the P protein of Indiana serotype vesicular stomatitis virus has been shown to be critical for in vitro transcription activity of the viral RNA polymerase (P-L) complex. To examine the role of phosphorylation of P protein in transcription as well as replication in vivo, we used a panel of mutant P proteins in which the phosphate acceptor sites in domain I were substituted with alanines or other amino acids. Analyses of the alanine-substituted mutant P proteins for the ability to support …

The Drosophila Melanogaster Rad54 Homolog, Dmrad54, Is Involved In The Repair Of Radiation Damage And Recombination, Rolf Kooistra, José B. M. Zonneveld, Anja De Jong, Jan C. J. Eeken, Chris J. Osgood, Jean-Marie Buerstedde, Paul H. M. Lohman, Albert Pastink

Biological Sciences Faculty Publications

The RAD54 gene of Saccharomyces cerevisiae plays a crucial role in recombinational repair of double-strand breaks in DNA. Here the isolation and functional characterization of the RAD54 homolog of the fruit fly Drosophila melanogaster, DmRAD54, are described. The putative Dmrad54 protein displays 46 to 57% identity to its homologs from yeast and mammals. DmRAD54 RNA was detected at all stages of fly development, but an increased level was observed in early embryos and ovarian tissue. To determine the function of DmRAD54, a null mutant was isolated by random mutagenesis. DmRAD54-deficient flies develop normally, but the females …

The Rna-Binding Site Of Bacteriophage Qβ Coat Protein, Francis Lim, Marc Spingola, David Peabody

Biology Department Faculty Works

The coat proteins of the RNA bacteriophages Qβ and MS2 are specific RNA binding proteins. Although they possess common tertiary structures, they bind different RNA stem loops and thus provide useful models of specific protein-RNA recognition. Although the RNA-binding site of MS2 coat protein has been extensively characterized previously, little is known about Qβ. Here we describe the isolation of mutants that define the RNA-binding site of Qβ coat protein, showing that, as with MS2, it resides on the surface of a large β-sheet. Mutations are also described that convert Qβ coat protein to the RNA binding specificity of MS2. …

Distribution Of R-Banded Chromosome Variants In A Bottlenose Dolphin (Tursiops Truncatus) Population, Kristine Anne Gunter

Dissertations and Theses

When combined with observational and behavioral data, chromosome variant analysis provides a means of assessing population structure in bottlenose dolphins, Tursiops truncatus. In this cytogenetic analysis of thirty-six bottlenose dolphins, a distribution of fluorescent R-band variants was catalogued and used as a way to characterize the genetic structure of a Tursiops population inhabiting the Matagorda Bay, Texas, area. Three different population genetic analytical techniques were evaluated for their usefulness in analyzing cytogenetic data. The chromosome variant data obtained from this study was then compared with cytogenetic data gathered from a resident dolphin population residing near Sarasota, Florida.

The purpose …

Numa Assembles Into An Extensive Filamentous Structure When Expressed In The Cell Cytoplasm, Alejandro Saredi, Louisa Howard, Duane A. Compton

Dartmouth Scholarship

NuMA is a 236 kDa protein that participates in the organization of the mitotic spindle despite its strict localization in the nucleus during interphase. To test how cells progress through mitosis when NuMA is localized in the cytoplasm instead of the nucleus, we have deleted the nuclear localization sequence of NuMA using site-directed mutagenesis and transiently expressed this mutant protein (NuMA-DeltaNLS) in BHK-21 cells. During interphase, NuMA-DeltaNLS accumulates in the cytoplasm as a large mass approximately the same size as the cell nucleus. When cells enter mitosis, NuMA-DeltaNLS associates normally with the mitotic spindle without causing any apparent deleterious effects …

Protein-Protein And Protein-Dna Interactions At The Bacteriophage T4 Dna Replication Fork. Characterization Of A Fluorescently Labeled Dna Polymerase Sliding Clamp, Daniel J. Sexton, Theodore E. Carver, Anthony J. Berdis, Stephen J. Benkovic

Chemistry Faculty Publications

The T4 DNA polymerase holoenzyme is composed of the polymerase enzyme complexed to the sliding clamp (the 45 protein), which is loaded onto DNA by an ATP-dependent clamp loader (the 44/62 complex). This paper describes a new method to directly investigate the mechanism of holoenzyme assembly using a fluorescently labeled cysteine mutant of the 45 protein. This protein possessed unaltered function yet produced substantial changes in probe fluorescence intensity upon interacting with other components of the holoenzyme. These fluorescence changes provide insight into the role of ATP hydrolysis in holoenzyme assembly. Using either ATP or the non-hydrolyzable ATP analog, adenosine …

The Carboxyl Terminus Of The Bacteriophage T4 Dna Polymerase Is Required For Holoenzyme Complex Formation, Anthony J. Berdis, Patrice Soumillion, Stephen J. Benkovic

Chemistry Faculty Publications

To further elucidate the mechanism and dynamics of bacteriophage T4 holoenzyme formation, a mutant polymerase in which the last six carboxyl-terminal amino acids are deleted, was constructed, overexpressed, and purified to homogeneity. The mutant polymerase, designated ΔC6 exo−, is identical to wild-type exo− polymerase with respect to kcat, kpol, and dissociation constants for nucleotide and DNA substrate. However, unlike wild-type exo− polymerase, the ΔC6 exo− polymerase is unable to interact with the 45 protein to form the stable holoenzyme. A synthetic polypeptide corresponding to the carboxyl terminus of the wild-type exo− polymerase was tested as an in vitro inhibitor of …

Circadian Clock-Controlled Genes Isolated From Neurospora Crassa Are Late Night- To Early Morning-Specific, Deborah Bell-Pedersen, Mari L. Shinohara, Jennifer J. Loros, Jay C. Dunlap

Dartmouth Scholarship

An endogenous circadian biological clock controls the temporal aspects of life in most organisms, including rhythmic control of genes involved in clock output pathways. In the fungus Neurospora crassa, one pathway known to be under control of the clock is asexual spore (conidia) development. To understand more fully the processes that are regulated by the N. crassa circadian clock, systematic screens were carried out for genes that oscillate at the transcriptional level. Time-of-day-specific cDNA libraries were generated and used in differential screens to identify six new clock-controlled genes (ccgs). Transcripts specific for each of the ccgs …

Mechanisms Of Cytokine-Mediated Localized Immunoprotection., Nora Sarvetnick

Journal Articles: Regenerative Medicine

No abstract provided.

Cloning Of Human Acetyl-Coa Carboxylase-Beta And Its Unique Features., Joohun Ha, Jung-Kee Lee, Kyung-Sup Kim, Lee A. Witters, Ki-Han Kim

Dartmouth Scholarship

Acetyl-CoA carboxylase, which has a molecular mass of 265 kDa (ACC-alpha), catalyzes the rate-limiting step in the biosynthesis of long-chain fatty acids. In this study we report the complete amino acid sequence and unique features of an isoform of ACC with a molecular mass of 275 kDa (ACC-beta), which is primarily expressed in heart and skeletal muscles. In these tissues, ACC-beta may be involved in the regulation of fatty acid oxidation, rather than fatty acid biosynthesis. ACC-beta contains an amino acid sequence at the N terminus which is about 200 amino acids long and may be uniquely related to the …

Open-Path Fourier Transform Infrared Studies Of Large-Scale Laboratory Biomass Fires, Robert J. Yokelson, David W. T. Griffith, Darold E. Ward

Chemistry and Biochemistry Faculty Publications

A series of nine large-scale, open fires was conducted in the Intermountain Fire Sciences Laboratory (IFSL) controlled-environment combustion facility. The fuels were pure pine needles or sagebrush or mixed fuels simulating forest-floor, ground fires; crown fires; broadcast burns; and slash pile burns. Mid-infrared spectra of the smoke were recorded throughout each fire by open path Fourier transform infrared (FTIR) spectroscopy at 0.12 cm⁻¹ resolution over a 3 m cross-stack pathlength and analyzed to provide pseudocontinuous, simultaneous concentrations of up to 16 compounds. Simultaneous measurements were made of fuel mass loss, stack gas temperature, and total mass flow up the …

High-Power Broadly Tunable Picosecond Ir Laser System For Use In Nonlinear Spectroscopic Applications, D. E. Gragson, B. M. Mccarty, G. L. Richmond, D. S. Alavi

Chemistry and Biochemistry

We developed a high-power tunable picosecond IR laser system suitable for nonlinear spectroscopic investigations. We employ a Ti:sapphire regenerative amplifier that produces 1.9-ps, 17-cm⁻¹, 800-nm pulses at 1 kHz as a pump source. White-light generation in ethylene glycol and optical parametric amplification in potassium titanyl phosphate are used to produce the IR pulses. The tuning range extends from 2.4 to 3.8 μm in the idler and 1.0 to 1.2 μm in the signal. A total efficiency (signal plus idler) as high as 20% was achieved. The spatial, spectral, and temporal characteristics of the IR beam are presented along …

Pancreatic Expression Of Interleukin-4 Abrogates Insulitis And Autoimmune Diabetes In Nonobese Diabetic (Nod) Mice., Regula Mueller, Troy Krahl, Nora Sarvetnick

Journal Articles: Regenerative Medicine

Diabetes in nonobese diabetic (NOD) mice is a T cell-dependent autoimmune disease. The destructive activities of autoreactive T cells have been shown to be tightly regulated by effector molecules. In particular, T helper (Th) 1 cytokines have been linked to diabetes pathogenesis, whereas Th2 cytokines and the cells that release them have been postulated to be protective from disease. To test this hypothesis, we generated transgenic NOD mice that express interleukin (IL) 4 in their pancreatic beta cells under the control of the human insulin promoter. We found that transgenic NOD-IL-4 mice, both females and males, were completely protected from …

Interleukin-10 Inhibits Tumor Metastasis Through An Nk-Cell Dependent Mechanism, Li-Mou Zheng, David M. Ojcius, F. Garaud, C. Roth, E. Maxwell, Z. Li, H. Rong, J. Chen, X. Y. Wang, J. J. Catino, I. King

All Dugoni School of Dentistry Faculty Articles

Interleukin-10 (IL-10) is a recently described pleiotropic cytokine secreted mainly by type 2 helper T cells. Previous studies have shown that IL-10 suppresses cytokine expression by natural killer (NK) and type 1 T cells, thus down-regulating cell-mediated immunity and stimulating humoral responses. We here report that injected IL-10 protein is an efficient inhibitor of tumor metastasis in experimental (B16-F10) and spontaneous (M27 and Lox human melanoma) metastasis models in vivo at doses that do not have toxic effects on normal or cancer cells. Histological characterization after IL-10 treatment confirmed the absence of CD8+ and CD4+ T cells and macrophages at …

The Effects Of Zinc On Skeletal Alkaline Phosphatase Activity And Skeletal Tissues, Susan L. Hall

Loma Linda University Electronic Theses, Dissertations & Projects

Zinc is essential for normal growth, however, very little is known about the mechanisms by which zinc influences growth. ALP, an ectoenzyme expressed in high levels in osteoblasts, is a zinc-containing metalloenzyme and is important in bone formation. Preliminary studies showed that low levels of zinc could increase ALP specific activity in human osteosarcoma, SaOS-2 cells. Current studies were designed to characterize this zinc effect, test possible mechanisms, and determine whether the effect is physiologically relevant. Incubation with zinc resulted in a time and dose dependent increase in ALP specific activity in the extracts and conditioned media of SaOS-2 cells, …

Application Of Monoclonal Antibodies Against Creatine Kinase Mm And Bb Isoenzymes For Creatine Kinase Mb Isoenzyme Detection, Lei-Chieh Chuang

Chemistry & Biochemistry Theses & Dissertations

Nondiagnostic ECGs and unknown characteristic chest pains increase the diIIiculty of Acute Myocardial Infarction (AMI) diagnosis. Biochemical markers, such as creatine kinase MB isoenzymes (CK-MB) and myoglobin, are employed for the early detection of AMI. Compared to myoglobin, CK-MB reveals a higher specific indicator for AMI detection owing to its abundance in cardiac tissue. Hence, a successful CK-MB detection is expected to improve AMI diagnosis and therapy.

A CK-MB detecting ELISA system was set up based on a sandwich assay principle by using specific CK-B and CK-M monoclonal antibodies. The dimeric nature of the CK-MB isoenzyme provides two epitopes on …

A Truncated Form Of The Pho80 Cyclin Of Saccharomyces Cerevisiae Induces Expression Of A Small Cytosolic Factor Which Inhibits Vacuole Inheritance., Teresa Nicolson, Barbara Conradt, William Wickner

Dartmouth Scholarship

Vacuoles project streams of vesicles and membranous tubules into the yeast bud where they fuse, founding the daughter cell organelle, vac5-1, which encodes a truncated form of the Pho80 cyclin, inhibits normal vacuole inheritance. An in vitro inheritance assay which measures the fusion of vacuoles serves as a model for several steps of this process. We find that cytosol isolated from the vac5-1 mutant is unable to promote the fusion of wild-type vacuoles in the in vitro assay. Wild-type vacuoles are irreversibly inactivated in a time- and temperature-dependent manner if preincubated with vac5-1 cytosol and ATP, suggesting the presence of …

Anion Induced Blue To Purple Transition In Bacteriorhodopsin, Mrunalini Pattarkine, Anil K. Singh

Faculty Works

Purple membrane (PM, λ" role="presentation">λmax" role="presentation">max 570 nm) of H. halobium on treatment with sulphuric acid changes its colour to blue (λ" role="presentation">λmax" role="presentation">max 608 nm). The purple chromophore can be regenerated from the blue chromophore by exogeneous addition of anions such as CI−" role="presentation">− and HPO42−" role="presentation">2−4. Chloride ion is found to be more effective than the dibasic phosphate ion in regenerating the purple chromophore. Nevertheless, one thing common to the anion regeneration is that both CI−" role="presentation">− and HPO42−" role="presentation">2−4 show marked pH effect. At pH 1.0 the efficiency of …

Selenocysteine, Identified As The Penultimate C-Terminal Residue In Human T-Cell Thioredoxin Reductase, Corresponds To Tga In The Human Placental Gene, Vadim Gladyshev, Kuan-Teh Jeang, Thressa C. Stadtman

Vadim Gladyshev Publications

The possible relationship of selenium to immunological function which has been suggested for decades was investigated in studies on selenuim metabolism in human T cells. One of the major ⁷⁵Se-labeled selenoproteins detected was purified to homogeneity and shown to be a homodimer of 55-kDa subunits. Each subunit contained about 1 FAD and at least 0.74 Se. This protein proved to be thioredoxin reductase (TR) on the basis of its catalytic activities, cross-reactivity with anti-rat liver TR antibodies, and sequence identities of several tryptic peptides with the published deduced sequence of human placental TR. Physicochemical characteristics of T-cell TR were …

Il-10 Is Necessary And Sufficient For Autoimmune Diabetes In Conjunction With Nod Mhc Homozygosity., Myung-Shik Lee, Regula Mueller, Linda S. Wicker, Laurence B. Peterson, Nora Sarvetnick

Journal Articles: Regenerative Medicine

Contrary to expectations based on in vitro experiments, we previously found that pancreatic IL-10 did not inhibit autoimmune diabetes but accelerated it in an MHC-dependent manner. Therefore, the ability of IL-10 to overcome the absence of all non-MHC diabetes susceptibility (Idd) alleles was studied in transgenic mice expressing pancreatic IL-10 backcrossed to B10.H2g7 congenic mice, which have no Idd alleles other than NOD MHC (H2g7). IL-10 transgenic backcross 1 (BC1) mice with H2g7/g7 haplotype developed clear-cut insulitis and diabetes, but neither transgenic mice with the H2g/b haplotype nor nontransgenic BC1 mice did so. Further implicating IL-10 in autoimmune diabetes, anti-IL-10 …

Characterization Of Vasoactive Intestinal Peptide Receptors On Human Megakaryocytes And Platelets, Sang K. Park, Thomas A. Olson, Nuran Ercal, Monica Summers, M. Sue O'Dorisio

Chemistry Faculty Research & Creative Works

Vasoactive intestinal peptide receptor I (VIPRI) expression was examined in megakaryocytes using reverse transcriptase-polymerase chain reaction (RT- PCR). VIPRI protein was characterized in platelet membranes using covalent crosslinking techniques. Human megakaryocytes were isolated from suspension cultures of cord blood and adult bone marrow mononuclear cells using a murine monoclonal antibody to human platelet glycoprotein IIB/IIIA (CD41) and immunomagnetic beads. RT-PCR primers were constructed for the VIP, VIPRI, and VIPRII genes as well as for megakaryocyte specific genes, c-mpl and platelet factor 4 (PF-4). VIP, VIPRI, c-mpl, and PF-4 were coexpressed in megakaryocyte mRNA. Southern blot analysis confirmed the expression of …