Life Sciences Commons^™

Fidelity Of Dna Polymerase In Vitro, Yannacos C. Philopoulos

Dissertations

The fidelity of DNA Polymerase I, isolated from B. licheniformis and B. stearothermophilus, was tested in the presence of carcinogens and anticancer compounds. All of these compounds inhibited (62.4-99.2%) the synthesis of DNA using Poly(dA-dT) as a template. The compounds were inhibitory at all concentration levels tested, 0.01, 1.0 and 100 mM. The fidelity of the enzyme appeared to be enhanced by the presence of these compounds, as indicated by misincorporation (50-70%), that is, the incorporation of incorrect nucleotide. Misincorporation in the presence of either carcinogens or anticancer compounds was less than that of control. The results from both organisms …

Electroimmunoassay: A New Competitive Protein Binding Assay Using Antibody Selective Electrodes, George Raymond Connell

Chemistry & Biochemistry Theses & Dissertations

The purpose of this study was to utilize the concept of antibody selective electrodes to developed a new competitive protein binding assay. This is a new concept and a potentially important analytical technique because it combines the advantages of RIA (sensitivity and selectivity) with the advantages of electrical assays (speed and low cost). A conjugate of PGE₂ and dibenzo—18- crown-6 (a cation selective ionophore) was synthesized, and incorporated into a plastic membrane. The ionophore conjugate increased the selectivity of a polyvinyl chloride (PVC) membrane to monovalent cations. Transmembrane potential was altered by anti-PGE₂ antisera in a concentration dependent …

Isomers Of 5.8s Ribosomal Rna, David Smith

Chemistry & Biochemistry Theses & Dissertations

When whole cell RNA was separated by high resolution PAG (i.e., polyacrylamide gel) electrophoresis, 5.8S rRNA migrated as three higher mobility intense bands, and three faint bands of lower mobility. The three intense bands represent conformational isomers which migrated as a single band under denaturing conditions on PAG. The three faint bands represent conformational isomers of a minor form of 5.8S rRNA which is elongated at the 5' terminus of the molecule. This minor form migrates as a single band of lower mobility than the major form when electrophoresed on a denaturing PAG. The minor form was determined to have …

Development Of A Procedure For Analysis Of High Density Lipoprotein Subclasses, Chan Chin

Chemistry & Biochemistry Theses & Dissertations

Human serum high density lipoprotein subclasses, HDL₂ and HDL₃ , were isolated by preparative salt density gradient ultracentrifugation and further analyzed by electrophoresis on a 4 to 15% concentration gradient polyacrylamide gel.

The separation of the major classes of serum lipoproteins was achieved after a single ultracentrifugation for 272,000 g at 15°c in a swinging bucket rotor. High resolution concentration gradient gel electrophoresis was found to be particularly suitable for the separation of lipoproteins. The isolation of HDL by precipitation methods, heparin-manganese and dextran-magnesium have also been evaluated in this study, and were found not to completely separate …

Electroimmunoassay A New Competitive Protein Binding Assay Using Antibody Selective Electrodes, George Raymond Connell

Chemistry & Biochemistry Theses & Dissertations

The purpose of this study was to utilize concept of antibody selective electrodes to develop a new competitive protein binding assay. This is a new concept and potentially important analytical technique because it combines the advantage of RIA (sensitivity and selectivity) with advantages of electrical assays (speed and low cost).

Development Of An On-Line Computer System For Enzyme Kinetics Studies, Jacqueline R. Thweatt

Chemistry & Biochemistry Theses & Dissertations

The PDP-11/10 minicomputer was interfaced to the Beckman 25 ultraviolet/visible spectrophotometer and this on-line computer system was applied to enzyme kinetics studies. This interface involved hardware equipment to locate, amplify, and transmit the analog signal to an analog-to-digital converter, and software to acquire and store the digital data. Data processing programs were used to calculate kinetic parameters and statistical information from the acquired data.

The Varian Cary 219 ultraviolet/visible spectrophotometer can be readily interfaced to the Apple II Plus microcomputer by hardware equipment which can be purchased from the manufacturer. Various software may also be purchased for use in enzyme …

Assay, Purification, And Characterization Of A Pantetheine Hydrolyzing Enzyme From Pig Kidney, Carl Thomas Wittwer

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

A microsomal glycoprotein hydrolyzing pantetheine to pantothenate and cysteamine has been solubilized and purified to homogeneity as determined by sodium dodecylsulfate electrophoresis. Four rapid, independent assays of pantetheine hydrolysis are described and compared along with a method for localizing enzymatic activity on polyacrylamide gels. The enzyme is solubilized on exposure to butanol and purified by heat treatment, (NH4)₂SO₄ fractionation, hydrophobic chromatography, and hydroxyapatite chromatography. The glycoprotein, purified 5600-fold in 22% yield, has a specific activity of 14 μmoles pantothenate produced/min/mg of protein, 35 times that previously reported. The enzyme has a pH optimum of 9.0-9.5 and a …

The Isolation And Characterization Of A Novel Iron Protein From Desulfovibrio Gigas, Alan Kay Smith

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The isolation, purification, and partial characterization of a novel iron-containing protein from the sulfate-reducing anaerobic bacterium, Desulfovibrio gigas, is described. The highly insoluble protein was isolated from the cell debris following osmotic shock of the bacteria. The insoluble fraction consistently contained about 90% of the cell-associated iron. This protein was treated with acid, chelating agents, detergents and proteases in order to study their effect on the iron solubility. The results of elemental analysis of a crude protein preparation were 5.3% iron, 2.9% sulfur and 11 .9% nitrogen. An independent colorimetric iron analysis showed 6.4% iron. The iron could be …

High-Temperature Battery Calorimeter, L. D. Hansen, R. H. Hart, D. M. Chen, H. F. Gibbard

Faculty Publications

A battery calorimeter was built for the measurement of thermal energy generation of high-temperature lithium–aluminum/iron sulfide battery cells, which are under development for electric vehicle propulsion and other energy storage applications. The calorimeter was designed with a temperature range of 400˚–500˚C, a detection limit of 1 mW, and an upper limit of heat flow of 50 W. The results of measurements on 200-Ah LiAl/FeS cells were in excellent agreement with the predictions of thermodynamic calculations based on precise measurements of the total cell polarization and the temperature coefficient of the emf. Details of the construction and operation principles of this …

Ortho Cyclization Reactions Of Catecholamines, Robert Lee Hudkins

Chemistry & Biochemistry Theses & Dissertations

A study was undertaken to examine the potential for ortho cyclization associated with the synthesis of 1,2,3,4,-tetrahydroisoquinolines using the Pictet-Spengler reaction. Specif1cally, the condensation of the biogenic amines, dopamine, norepinephrine and epinephrine were carefully examined in light of current literature and the facility and extent of ortho cyclization in each case. The para isomers of these three biogenic amine condensation products were isolated and characterized, while only the ortho isomer derived from dopamine was found stable and could be isolated, and structurally identified. These studies demonstrate that a second cyclization product does, in fact, form in each instance and that …

The Voluntary Water Quality Monitoring Program : 1982 Project Report, Barbara Welch, Judy Potvin, Mary Ellen Dennis

Maine Collection

The Voluntary Water Quality Monitoring Program : 1982 Project Report

by Barbara Welch, Judy Potvin & Mary Ellen Dennis

Division of Environmental Evaluation and Lake Studies, Maine Department of Environmental Protection, Augusta, Maine 1982.

Contents: Understanding Maine's Lakes and Ponds / Acknowledgement / Temperature / Dissolved Oxygen / Other Measures of Lake Water Quality / Nutrients / Algae and Chlorophyll a / Transparency / Morphometry / Sampling / Protection / Classification / Answers to Most Frequently Asked Questions / Glossary of Terms / Appendix A : Maximum, Minimum, and Mean Values / Appendix B : Phosphate Content of Common Cleansers …

The Uptake And Release Of Label From Endocytosed 35so4-Labeled Proteins In Cultured Mouse Peritoneal Macrophages, Susan Patterson Buktenica

Dissertations

No abstract provided.

Effects Of Pulsed Electric Fields On Mammalian Cell Membranes, Guy K. Smith

Theses and Dissertations

The effect of pulsed electric fields on cell membranes were studied to investigate the effects of an electric field, per se, on biological systems, without the densometric and other technical problems associated with other forms of nonionizing radiation. Exposure of mouse splenocytes to a high voltage pulse resulted in an increase in membrane permeability to K⁺ that was dependent on both the electric field strength and the pulse duration. Exposure to a 2 μsec, 3.0 kV/cm pulse elicited a 50% loss of intracellular K⁺ indicating that the critical transmembrane potential (V_m) at breakdown was 1.26 volts …

Organic N-Chloramines: Chemistry And Toxicology, Frank E. Scully, Jr., Maxwell A. Bempong

Chemistry & Biochemistry Faculty Publications

The stability of aqueous solutions of organic N-chloramines, suspected of contaminating chlorinated water, has been studied. Two factors influence the decomposition of solutions of N-chloropiperidine and N-chlorodiethylamine: a spontaneous decomposition and photodecomposition. Since solutions of these compounds are relatively long-lived, a need for an analytical method for their identification is discussed. A new method is described which involves reaction of organic N-chloramines with arenesulfinic acid salts. The method gives high yields of stable arenesulfonamides. Several toxicological studies of N-chloropiperidine are described. The compound is mutagenic by Ames assay in Salmonella typhimurium strain TA 100 and does not require metabolic activation …

Nuclear Envelope Proteins: Chemical Characteristics And Molecular Interactions Of The Lamins, David Lee Cochran

Theses and Dissertations

The lamins are a group of proteins in a residual nuclear envelope fraction derived from the nuclear lamina. The absence of specific functional assays for the two predominant species, lamin A and lamin B, has complicated their biochemical characterization. Initial studies reported in this work involved the enzymic cleavage of isolated lamins A and B and another lamina protein with a molecular weight of 61,000-daltons. The results obtained with each of three enzymes indicated strong similarities between the three nuclear envelope proteins.

Further chemical characterization of the lamins has been achieved. Lamins A, B and C have been separated by …

Iodothyronine 5'-Deiodinase: Solubilization, Partial Purification, And Characterization Of A Thyroxine Deiodinase Extracted From Rat Liver Membranes, Samuel C. Powell

Dissertations

No abstract provided.

Electron Microscopic Demonstration Of Neural Connections Using Horseradish Peroxidase: A Comparison Of The Tetramethylbenzidine Procedure With Seven Other Histochemical Methods, Keith A. Carson, M-Marsel Mesulam

Biological Sciences Faculty Publications

Eight methods for the electron microscopic demonstration of horseradish peroxidase (HRP) labeling have been compared in adjacent series of vibratome sections of mouse lumbar spinal cord. The tracer, a HRP-wheat germ agglutinin (WGA) conjugate, was injected into the gastrocnemius muscle complex. Following retrograde axonal transport to the lumbar motor neurons and transganglionic anterograde transport of the tracer to the dorsal horn, the HRP activity was demonstrated in eight series of adjacent sections of lumbar spinal cord using eight methods. These included procedures using tetramethylbenzidine (TMB), benzidine dihydrochloride (BDHC), o-tolidine, paraphenylenediamine-pyrocatechol (PPD-PC), and 4 methods using 3,3'-diaminobenzidine (DAB). All eight methods …

The Effects Of Maternal Protein Deficiency On The Development Of Synaptic Plasma Membranes In Rat Offspring, Duane M. Smith

Dissertations

No abstract provided.

Elastin Content Of The Aorta From The Quantitation Of The Desmosines, H. Patrick Covault

Dissertations

No abstract provided.

A Microbial Reductive Conversion Of The Steffimycin Chromophore: Chemical And Mechanistic Studies On A Soluble, Nadhdependent, Nonflavin-Containing Reductase From Streptomyces Steffisburgensis, John Rollin Hofstetter

Dissertations

An understanding of a microbial reduction of the anthracycline antibiotic, steffimycin (1) was approached from two directions. First, the NADH-dependent enzyme responsible for the reduction was purified from the Streptomyces steffisburgensis cytosol and characterized. Second, information was obtained on the reduction and oxidation of 1.

The enzyme was a single polypeptide chain, acidic, and stable in solution. It had a molecular weight of 80,000 and no prosthetic groups.

The 5-step purification process resulted in greater than 1,000-fold purification and a 9% yield of the homogenous enzyme from extracts of S. steffisburgensis cells.

The enzyme reduced 1 by a two-electron mechanism …

Correlation Between Glutathione Peroxidase Activity And Selenium Concentrations In Both Plasma And Erythrocytes, Agatha Dudzai Munyanyi

Chemistry & Biochemistry Theses & Dissertations

Plasma and erythrocyte glutathione peroxidase activities and selenium concentrations were determined for 38 apparently healthy female subjects. In addition, erythrocyte lead and plasma lead and iron concentrations were determined in each subject.

Effects of age, smoking, vitamin and mineral supplementation, use of caffeinated beverages and oral contraceptives on glutathione peroxidase activity were determined and interelement relationships were investigated using the one tailed t-test and Spearman Rank Correlation Coefficients

In Vitro Enzymatic Assay Of Rna Methylation, Pamela Jean Eubanks Gallup

Chemistry & Biochemistry Theses & Dissertations

An assay procedure for in vitro enzymatic methylation of mannnalian ribosomal RNA has been developed in this study. The assay procedure, utilized for the comparison of normal and neoplastic methylase activities (using mouse liver and Ehrlich ascites cells as sources of enzyme), is a modification of previously published methods (52,53). A 100,000 x g supernatant (SlOO) enzyme preparation was incubated with 28S-5.8S rRNA and tritium-labeled S-adenosyl-L-methionine. The RNA was extracted, applied to DEAE cellulose paper, washed, and the radioactivity counted. The neoplastic cell methylase preparation was more active in methylating both exogenous neoplastic and normal 28S-5.8S rRNA than the normal …

The Mechanism Of Aerotaxis In Salmonella Typhimurium, Daniel John Laszlo

Loma Linda University Electronic Theses, Dissertations & Projects

Chemotaxis is the migrational response of bacteria to chemical gradients. Aerotaxis, the response to oxygen, was among the earliest observed tactic phenomena, but progress in the study of aerotaxis was previously limited by the available assays. These assays relied upon oxygen depletion by the bacteria to generate the oxygen gradient which elicits the behavioral response. Such assays are unsuitable for analyzing the role of the electron transport system in aerotaxis. In this study the biochemical mechanism of aerotaxis and its relationship to other chemotaxis pathways in Salmonella typhimurium was explored with the aid of two new assays which imposed an …

Chemical And Biological Structure Of Lake Mead Sediments, Richard T. Prentki, Larry J. Paulson, John R. Baker

Publications (WR)

Lake Mead has undergone a serious decline since Glen Canyon Dam was constructed 450 km upstream in 1963.

State fisheries management agencies are concerned that the decline was caused by water level fluctuations and more severe drawdowns during the bass spawning season, when the operation of Hoover Dam was altered during the post-Lake Powell period.

The construction of Glen Canyon Dam and formation of Lake Powell in 1963 drastically altered the natural discharge and temperature cycles and decreased suspended sediment and nutrient loading in the Colorado River inflow to Lake Mead. Recent studies indicate that these changes in nutrient loading …

An Enzyme-Linked Immunosorbent Assay (Elisa) For Pantothenate, Allen H. Smith

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

An enzyme-linked immunosorbent assay (ELISA) for pantothenate has been developed. Antibodies induced in rabbits against bovine serum albumin-pantothenate conjugate were specifically purified by affinity chromatography. This process served to reduce the amount of endogenous pantothenate attached to the antibody, as well as to purify the antibody. The purified antibodies were covalently linked to alkaline phosphatase (Sigma type VII) with glutaraldehyde (0.05% aqueous solution). An immobilized pantothenate substrate was first obtained by attaching human serum albumin-pantothenate conjugate to the surface of polystyrene culture tubes by passive adsorption. The binding of the enzyme labelled antibody (E-AB) to this substrate is proportionately inhibited …

The Transport And Intracellular Distribution Of Iron In The Basidiomycete, Ustilago Sphaerogena, Charles W. Passavant

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The transport and intracellular distribution of iron in Ustilago sphaerogena via the native siderochrome ferrichrome A has been examined. Evidence is presented which suggests the involvement of deferriferrichrome in iron transport when the metal is supplied to the cell as ferrichrome A. This process requires both oxygen and ATP synthesis and may require a transmembrane electrochemical gradient. Once inside the cell, iron derived from ferrichrome A is rapidly distributed, with a major fraction found in the mitochondria, presumably as heme.

The kinetics of ferrichrome and ferrichrome A uptake have also been investigated. The Km of transport of ferrichrome was …

High Performance Liquid Chromatographic Analysis Of Biogenic Amines, Thais Edwina Ahrendt Keegan

Chemistry & Biochemistry Theses & Dissertations

The objectives of this research were to develop a sensitive method for separation and quantitation of biogenic amines and to apply that procedure by monitoring neurochemical changes resulting from stimuli to the test system.

Primary aromatic amines were derivatized with o-phthalaldehyde-mercaptoethanol prior to chromatography, Separation of the fluorescent adduct was achieved using a μBondapak/ phenyl reversed-phase column and a two-step gradient of methanol:0.08 M acetic acid, pH J.O. Fluorescence was monitored using a J40 nm excitation energy and an emission filter which transmits radiation greater than 400 nm (50% T at 418 nanometers).

Following optimization of the chromatographic conditions, studies …

The Synthesis And Evaluation Of The Biological Activity Of Heterocyclic Analogs Of Phencyclidine, Julian Dew Mckenney Jr.

Chemistry & Biochemistry Theses & Dissertations

Using the classical synthetic approach described by Maddox and co-workers⁴³, the decyanation reaction of 1- piperidinocyclohexane carbonitrile (PCC), 5, was attempted using the heterocyclic anions of 2-methyl-5-nitropyridine, 22, 2-amino-5-nitropyridine, 23, and 2-hydroxy-5-nitropyridine, 24; however, the desired pyridine analogs of phencyclidine (PCP), 1, could not be prepared under these conditions. Attempts to prepare the same analogs via addition of these anions to the iminium tosylate and hydrobromide salts also proved unsuccessful.

The failure to observe the desired decyanation reaction has been attributed to the special sterochemical restrictions of the hindered nitrile of PCC in that facile …

Food Habits Of Largemouth Bass, Micropterus Salmoides (Lacepede) And Spotted Bass, Micropterus Punctulatus (Rafinesque), From Beech Fork Reservoir, A New Impoundment In Wayne And Cabell Counties, West Virginia, Michael A. Arcuri

Theses, Dissertations and Capstones

A study was undertaken to determine the food habits of two game fishes, the largemouth bass Micropterus salmoides (Lacepede) and the spotted bass M. punctulatus (Rafinesque) in Beech Fork Reservoir shortly after its impoundment. A total of 171 largemouth bass and 95 spotted bass were collected from July 1978, through June 1979, and their stomach contents analyzed.

Largemouth bass fed primarily on fishes, particularly young-of-the-year bluegill. Aquatic and terrestrial invertebrates were also important food items, especially during the spring and fall . The dominant aquatic forms were chironomid larvae and pupae while the dominant terrestrial forms were adult dipterans.

Spotted …

Peptidohydrolases Of Soybean Root Nodules, Nasir S. A. Malik, N. E. Pfeiffer, D. R. Williams, Fred W. Wagner

Department of Biochemistry: Faculty Publications

Nodule extracts prepared from Glycine max var Woodworth possessed endopeptidase, aminoptidase, and carboxypeptidase actiities. Three distinct endopeptidase activities could be resolved by disc-gel electrophoresis at pH 8.8. According to their order of increasing electrophoretic mobility, the first of these enzymes hydrolyzed azocasein and n-benzoyl-LLeu-β-naphthylamide, while the second hydrolyzed n-benzoyl-L-Arg-/8- naphthylamine (Bz-L-Arg-/βNA), n-benzoyl-L-Arg-β-nitroanilide (Bz-L-Arg-βNA), and azocasein. The third endopeptidase hydrolyzed Bz-L-Arg-βNA, Bz-L-Arg-βNA, and hemoglobin. Fractions of these enzymes extracted from electrophoresis gels were shown to have pH optima from 7.5 to 9.8. AU of the endopeptidases were completely inhibited by diisopropylphosphorofluoridate, demonstrating that they were serine proteases.