Biochemistry Commons^™

Heterologous Expression And Characterization Of Three Active Recombinant Fungal Peroxidases From Phanerochaete Chrysosporium Using Baculovirus, Todd M. Johnson

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

In a collaborative effort with Dr. Steven D. Aust's group at Utah State University, this research is the first to describe the heterologous expression of active peroxidases from the filamentous fungus Panerochaete chrysosporium. The cDNA clones λML-1, λML-6, λML-5 and λMP-1, which had been tentatively identified as fungal peroxidase clones, were subcloned into baculovirus transfer vectors and recombinant Autographa californica nuclear polyhedrosis viruses were constructed which contained copies of these potential peroxidase genes. The recombinant baculovirus clones were used to infect insect cells for the heterologous production of active recombinant enzymes. Three recombinant lignin peroxidases (H2, H6 and H8) …

Peptide Mapping Analysis Of Vp5 And Vp7 From Five Bluetongue Viruses Isolated In The United States, Yi-Yuan Yang

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

BTV-2, 10, 11, 13 and 17 were purified by three discontinuous sucrose gradients. Viral proteins were separated from the genomic double-stranded RNA using a newly developed KCI-SDS (sodium dodecyl sulfate) precipitation method. Cyanogen bromide (CNBr) peptide mapping of VP5 and VP7 from bluetongue virus (BTV) serotypes 2, 10, 11, 13 and 17 was then done using silver-stained SDS-polyacrylamide gels. The antigenic epitopes of VP5 and VP7 digested with CNBr were also investigated with polyclonal antibodies produced against KCl-SCS precipitated BTV-13 viral proteins. The peptide maps and the antigenic epitopes of VP7 were similar among the five American serotypes with very …

The Involvement And Relationship Of Ferritin And Ceruloplasmin In Oxygen Radical Toxicities, David Wilson Reif

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The intent of this dissertation is to demonstrate that ferritin iron may be involved in alloxan, diquat, and radiation-dependent damage and that ceruloplasmin may afford protection by limiting the availability of ferritin iron. Initial studies demonstrated that the alloxan radical formed by glutathione-dependent reduction of alloxan reductively mobilized ferritin iron and the released iron catalyzed lipid peroxidation. SOD and catalase were without effect, whereas ceruloplasmin effectively inhibited both iron release and lipid peroxidation. Using ¹³⁷Cs gamma-radiation it was demonstrated that radiolytically generated superoxide released iron from ferritin with an efficiency of 67%. Lipid peroxidation strictly required both superoxide and …

A Study Of The Action Of Bovine Cathepsin D On Intramuscular Connective Tissue, Elimosaria Elikalia Maeda

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Cathepsin D has been postulated to play a role in the tenderization of meat with ageing. Meat toughness has been related to connective tissue content and degree of myofibrillar shortening. This investigation was undertaken to determine the activity of cathepsin D on bovine intramuscular connective tissue.

Highly purified cathepsin D preparations from bovine spleen and bovine sternomandibularis muscle were obtained by ammonium sulfate fractionation followed by affinity chromatography on pepstatin coupled to controlled pore size alkyl amine glass beads. Maximum proteolytic activity using hemoglobin as substrate was obtained at pH 3.5 and 3.8 for bovine spleen and muscle cathepsin D, …

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 …

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 Regulatory Effects Of Certain S-Triazine Compounds On The Nutritive Composition Of Seeds And Leaves Of The Pea (Pisum Sativum L.) And Sweet Corn (Zea Mays L.) And Responses Of Metabolic Systems, Ming-Tsung Wu

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The study was conducted to investigate the influence of eight s-triazine compounds--simazine, atrazine, terbutryne, GS-14254, propazine, prometryne, prometone, and ametryne--on the composition of seeds of peas and sweet corn through soil and foliar application. In order to understand the mechanism of the effects of these a-triazines on protein accumulation, the responses of the metabolic systems of pea and sweet corn seedlings to s-triazine treatment was studied with emphasis on the stimulation of certain enzymes, and the changes in the rates of incorporation of ¹⁴C-L-leucine (U) into soluble protein. In addition, the effect of simazine on the ultrastructure …

The Effects Of Sources And Levels Of Vitamin A And D On Some Aspects Of Sulfur Metabolism, Akbar Malekpour

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Two trials were conducted using 48 infant pigs of both sexes. Pigs were taken from the sow when three to seven days old and placed in wire-bottomed metal cages by pairs. Animals in the first trial received synthetic diets containing 100 or 1000 international units of ergocalciferol and 1000 or 4000 international units of retinyl acetate or four or sixteen milligrams of beta carotene per kilogram of feed. Baby pigs in the second trial received the same synthetic diet plus 1000 international units of ergocalciferol (D₂) or cholecalciferol (D₃) and 2000 international units of vitamin A …

Gas Chromatographic Determination Of Changes In Concentration Of Pipecolic Acid And Other Free Amino Acids In Hypersensitive Tobacco Plants Induced By Tobacco Mosaic Virus And Temperature, David Francis Brown

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Changes in concentrations of pipecolic acid and other free amino acids in Nicotiana tabacum, L. cv. Xanthi induced by tobacco mosaic virus (TMV) incubated at 75 or 95 F after inoculation were investigated gas chromatographically. A preliminary extraction and gas chromatographic technique for separation and quantitative determination of the free amino acids, using hydroxyproline as an internal standard, was developed.

Amino acids were converted into their trifluoroacetyl n-butyl (TAB) derivatives and gas chromatographed on 0.325 percent ethylene glycol adipate (EGA) columns. Resolution was generally good, but glycine-isoleucine and phenylalanine-aspartic acids were not resolved on old columns. The over-all …

Quantitative Evaluation Of Saponin Content In Du Puits Alfalfa Foliage (Medicago Sativa L.) With Trichoderma, Wang Li-Chun

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Selections in Du Puits alfalfa (Medicago sativa L. ) were evaluated for saponin content. The data obtained from the Trichoderma bioassay were converted to saponin values by regressions and standard curves. It was necessary to adjust the concentration of alfalfa extract in the bioassay so that the saponin content in alfalfa was estimated in that portion of the curve showing a marked response to saponin. A different regression was established for each group of low and high saponin plants of Du Puits alfalfa. The standard curve prepared with extract of Du Puits alfalfa used on the basis of its …

Biosynthesis Of Piperidine-2-Carboxylic Acid In Fluoride-Necrotic Glycine Max, Susan S. Martin

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Synthesis of piperidine-2-carboxylic acid (pipecolic acid) in soybean was studied by infiltration of radioactive carbon labeled precursor and by experiments using enzyme extracts from control and fluoride-fumigated plants.

Pipecolic acid was synthesized from lysine in both control and treated plants. Normal plants, in which pipecolic acid had not been detected by conventional procedures, were capable of synthesizing pipecolic acid. Greater pipecolic acid synthesis was shown in treated plants than in controls, and was attributed to increased enzyme concentration induced by fluoride treatment.

The Biosynthesis Of Aminolevulinic Acid In Relation To Iron And Enzyme, Daniel Ming-Yi Teng

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The survey of plant species for aminolevalinic acid synthetase (ALAS) source was conducted. Under the conditions of this study, spinach leaves were found to be a good source of ALAS activity. The amount of amino levulinic acid (ALA) synthesized by the plant extracts was very small and was rapidly metabolized to porphyrins. ALA and porphyrin synthesis was found to be confined to the particulate fractions. Rupturing of the particles with ultrasonifier enhanced the penetration of succinyl-CoA and increased the activity. Direct study of the ALAS activity was enabled by the utilization of C¹⁴-compounds to the isolation by column …

Interaction Of Inorganic And Organic Fluoride Ions With Available Calcium On Root Growth In Triticum Vulgare, L., Subbanaidu Ramagopal

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

A method has been described to suspend seedling wheat roots in a saturated atmosphere. Wheat (Triticum vulgate, L.) seeds were germinated in aluminum trays and the seedlings transferred to seedling chambers. After a day's growth, the roots were treated with various concentrations of solutions containing CaSO₄, KCl, KF, NaCL, NaF, CH₃COONa, FCH₂COONa, K₂C₂O₄ and Na₂EDTA and observations were recorded 24 and 48 hours after treatment. The new root growth was described with respect to root hairs, root tissue collapse and root discoloration. Total length and …

Fluoroorganic Compounds In Relation To Hydrogen Fluoride Exposures In Leaves Of Glycine Max Merr., Julie Ye-O Cheng

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Soybean plants were fumigated in HF and the organic acid-fractions of the leaf extracts were compared to those of the leaf extracts from soybean plants grown in nutrient solution containing fluoroacetate. The extracts from both treatments contained fluoroorganic compounds that were chromatographically similar to fluoroacetate and fluorocitrate.

The fluorocitrate separated from HF fumigated leaves was found to inhibit aconitase prepared from pig-heart. This inhibition was similar to that caused by commercial fluorocitrate and the fluorocitrate synthesized using guinea-pig kidney. The aconitase from soybean leaves was found to be sensitive to fluorocitrate as the aconitase that isolated from pig-heart. Inorganic fluoride …

Alkali Liberated Phenolic Acids From Alfalfa, Leslie Huey-Seng Huang

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Fourteen compounds were isolated from an alkaline hydrolyzed extract of alfalfa leaves by using cellulose thin layer plates. The R_f values, fluorescent characteristics, and color reactions were given for all the compounds isolated.

Eight of these were identified as trans-ferulic acid, trans-p-coumaric acid, trans-caffeic acid, o-coumaric acid, tricin, syringic acid, cis-ferulic acid and cis-p-coumaric acid. The identity of the compounds was confirmed by comparison of the R_f values on thin layer chromatograms, fluorescent characteristics under ultraviolet light, and color reactions of spray reagents with those of authentic samples. Three of …

The Complex Formation Of Silver Ion With Ribonucleic Acid, Guanosine, Inosine And Related Compounds And Peroxidase-Like Activity Of A Haemundecapeptide Prepared From Horse Heart Cytochrome C, José Angel Reinosa

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The importance of nucleic acids in plant and animal cells as carriers of genetic information and as protein biosynthesis agents is well recognized. It is also known that nucleic acid is a component of all viruses.

Takahashi (45) and Fraenkel-Conrat (16) demonstrated that the protein component of tobacco mosaic virus is non-infectious to the host plant, although it is identical to the original virus morphologically. The virus ribonucleic acid (RNA) alone was infectious, however.

Deoxyribonucleic acid (DNA), which is present in chromosomes, displays a very specific function. The chromosome long has been accepted as the carrier of the hereditary unit, …

Saponin Content And Some Pod And Blossom Characteristics Of Alfalfa As Related To Seed Infestation By The Alfalfa Seed Chalcid, Ronald D. Morse

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The alfalfa seed chalcid, Bruchophagus ruddi Guss., is a jet-black hymenopteran wasp. The destructive nature of this pest has been recognized since the latter part of the nineteenth century. Every year thousands of acres of alfalfa seed are destroyed, with infestation reaching as high as 8 5 percent in some areas. In Utah the chalcid annually ruins from 5 to 25 percent of the alfalfa seed. Much of this damage goes unnoticed, as infested seed is commonly blown out in the trash during harvesting and cleaning operations . The extent of damage is not restricted to the United States. Wherever …

The Effect Of Fluoride On C14O2-Fixation And The Synthesis Of Organic Acids, Amino Acids, Protein And Carbohydrates, Chin-Fung Li

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Many studies have been conducted on the effect of fluoride on plants, but the exact mechanisms by which fluoride causes damage is little understood. The following physiological and metabolic processes are known to be affected by fluoride: a decrease in chlorophyll (McNulty and Newman, 1961), a diminished rate of photosynthesis (Thomas and Hendricks, 1956; Bishop, et al., 1958), decreased plant growth (Bonner and Thimann, 1950), inhibition of ascorbic acid synthesis (Smith, 1952), variation in starch and total polysaccharides (Donald and Emerson, 1961), inhibition of glucose transport (Jiri, et al., 1962), inhibition of metabolism (Ordin and Skoe, 1963), effects …

A Study Of 5-Fluorouracil: Its Effects And Mode Of Action In Drosophila Melanogaster, John B. Jenkins Jr.

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Investigations involving the fluorinated pyrimidine, 5-fluorouracil, and its effects on viruses, bacteria, plants and mammals, have brought to light the following facts: FU inhibits the growth of bacteria, plants and mammalian cells and it decreases the ability of certain viruses to produce progeny. In other words, FU is an antimetabolite that inhibits growth and/or reproduction. Two mechanisms have emerged from these studies to define the mode of action of FU. The first mechanism involves the incorporation of the compound into the RNA of the organism thus altering the base sequence and creating a non functional or damaging nucleic acid. The …

Effect Of Fluoride On The Respiration Of Leaves From Higher Plants, Ming-Ho Yu

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Studies concerning the effects of fluoride on the metabolism of both animals and plants have been of interest to many workers. In plants, it has been known that fluoride will inhibit the growth, that it will induce chlorosis as an evidence of visible injury, and that necrosis will eventually result. The economic importance associated with fluoride damage has been widely recognized, and, as a consequence, fluoride has become a major problem of air pollution.

Studies Of The Incorporation Of 5-Fluorouracil Into Tobacco Mosaic Viral Rna By Cell-Free Synthesing Systems Isolated From Infected Tobacco Plants, Helen Lu-Sheng Wang

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Research on biosynthesis of tobacco mosaic virus (TMV) ribonucleic acid in vitro has been reported by a number of laboratories (8, 16, 17). Bonner et al. postulated that ribonucleic acid could be synthesized from nucleotides by chromatin extracted from nuclei of pea seedlings (5, 14, 22). The evidence from in vitro synthesis of viral RNA suggested that an enzyme, RNA polymerase, links the four nucleotides (ATP, UTP, CTP, GTP) with viral RNA present as a primer and magnesium ions participating as a catalyst (7, 8).

The Respiratory Chain Involved In Oxidative Phosphorylation On Higher Plants In Relation To Carbon Dioxide-Bicarbonate Inhibition, Wan-Jean Young

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Increased concentrations of CO₂ with the accompanying high bicarbonate level in the culture medium have been reported to have a depressing effect in certain plant species on growth (34), respiration (23, 33), oxidative phosphorylation (9), mineral nutrient absorption (17), certain enzymatic reactions (4, 22, 27), accumulation of organic and amino acids (11), and protein synthesis (16). Since accumulation of organic acids occurs in lime-induced chlorotic tissue together with a depression in the respiration of plant tissue, an effect on the energy transfer mechanism of such plants is suspected. Inhibition of electron transfer from the substrate level could result in …

Biosynthesis Of Infectious Cucumber Mosaic Virus Ribonucleic Acid In A Cell-Free Medium, Men Hui Lee

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The mechanism of biosynthesis of the nucleic acids, RNA, remained obscure until 1955 when the announcement was made (14) that an enzyme isolated from bacterial sources catalyzes the synthesis of ribopolynucleotides from nucleoside-5'-diphosphates. It has been shown that the biosynthesis of polynucleotides occurs through the stepwise polymerization of nucleoside-5'-diphosphates or triphosphates. A great deal of research over the past few years has indicated that isolated nuclei from both animal (1) and plant (19) sources are able to synthesize ribonucleic acids in vitro (18).

Leaf Ribonuclease Activity As Related To Virus Infection, And Ribonucleic Acid Concentration, Andrew Chien-Yau Hsu

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Ribonucleic acid (RNA) appears to be the entity responsible for infectivity and genetic continuity in certain plant, animal and bacterial viruses. RNA also plays an important role in protein synthesis. The enzymes which are involved in RNA synthesis and degradation have received much attention. Among these enzymes ribonuclease (RNase) has been most thoroughly studied Rnase is widely distributed and has been isolated from many sources such as snake venom, bacteria, the pancreas, and many plant and animal tissues. Most studies with RNase have been concerned with its role in the hydrolysis of RNA in vitro. Therefore, it has generally been …

The Biochemical Differences Associated With Iron Chlorosis In Higher Plants, Daniel Ming-Yi Teng

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Chlorosis, which is associated with a reduction in the chlorophyll content of leaves, is a major problem in the growing of deciduous fruit trees and some field crops on calcareous soil. Under various soil conditions more than one factor may be involved in the development of chlorosis in plants. A deficiency of magnesium, manganese, zinc or iron, and virus infections can induce chlorosis. It is also induced by the presence of high concentrations of phosphate, bicarbonate, manganese, copper, zinc, or numerous other factors.

Biochemical Studies On The Effect Of Fluoride On Higher Plants, Shang-Fa Yang

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Fluoride occurs naturally in plants, but its presence has attracted attention primarily in certain industrial areas where concentrations are elevated above normal by accumulation from the atmosphere. Certain plant species have been observed to be injured as a result of accumulation of excessive fluoride from the atmosphere. The general symptoms of fluoride injury are necrotic lesions and burning which appear first in the leaf tips and margins.