Biochemistry, Biophysics, and Structural Biology Commons^™

Expression Of Holo And Apo Forms Of Spinach Acyl Carrier Protein-I In Leaves Of Transgenic Tobacco Plants, M. A. Post-Beittenmiller, Katherine M. Schmid, J. B. Ohlrogge

Scholarship and Professional Work - LAS

Acyl carrier protein (ACP) is a chloroplast-localized cofactor of fatty acid synthesis, desaturation, and acyl transfer. We have transformed tobacco with a chimeric gene consisting of the tobacco ribulose-1,5-bisphosphate carboxylase promoter and transit peptide and the sequence encoding the mature spinach ACP-I. Spinach ACP-I was expressed in the transformed plants at levels twofold to threefold higher than the endogenous tobacco ACPs as determined by protein immunoblots and assays of ACP in leaf extracts. In addition to these elevated levels of the holo form, there were high levels of apoACP-I, a form lacking the 4'-phosphopantetheine prosthetic group and not previously detected …

A Flow Cytometric Approach To Assessing The Environmental And Physiological Status Of Phytoplankton, Serge Demers, Kimberly Davis, Terry L. Cucci

CCPO Publications

No abstract provided.

Role Of Metabolites In The Reversible Light Activation Of Pyruvate,Orthophosphate Dikinase In Zea Mays Mesophyll Cells In Vivo, Chrissi A. Roeske, Raymond Chollet

Department of Biochemistry: Faculty Publications

Whole leaf and mesophyll cell concentrations of pyruvate, phosphoenolpyruvate (PEP), ATP, and ADP were determined in Zea mays during the reversible light activation of pyruvate, orthophosphate dikinase in vivo. Mesophyll cell levels of the four metabolites were estimated by extrapolation from values in freeze-quenched leaf samples that were fractionated by differential filtration through nylon mesh nets (adapted from M Stitt, HW Heldt [1985] Planta 164: 179-188). During the 3 minutes required for complete light activation of dikinase, pyruvate levels in the mesophyll cell decreased (from 166 ± 15 to 64 ± 10 nanomoles per milligram of chlorophyll [nmol/mg Chi]) …

Chloroplast Thylakoid Protein Phosphatase Is A Membrane Surface-Associated Activity, Gongqin Sun, Doreen Bailey, Michael W. Jones, John Markwell

Department of Biochemistry: Faculty Publications

Chloroplast thylakoid protein phosphatase activity was measured using ³²P-labeled histone as an exogenous substrate and an assay of the ³²P released involving formation of a phosphomolybdate complex and organic extraction. The activity was liberated from wheat (Triticum aestivum) thylakoids by washing the membranes in NaCI-containing solutions followed by centrifugation. The liberated phosphatase activity had a pH optimum of approximately 6.75, was inhibited by addition of 10 millimolar EDTA or EGTA, and was stimulated by addition of millimolar amounts of dithiothreitol, magnesium, manganese, or calcium ions. The rate of thylakoid protein dephosphorylation was decreased following liberation of a portion …

Multiple Isotope Effects On Enzyme-Catalyzed Reactions, Marion H. O'Leary

Department of Biochemistry: Faculty Publications

Isotope effects have long been popular as a method for studying mechanisms of chemical reactions (1, 2). Application of this method to enzymatic reac­tions has come slowly, in part because of the difficulty of making measurements of the necessary precision, and in part because of the difficulty of interpreting the variations in rate that occur in multistep reactions. The theory and practice of isotope effects has now reached the stage where a variety of interesting mechanistic studies are possible (3-9), including use of heavy­ atom isotope effects (5), application to multireactant enzymes (9a, 10), pH dependence of isotope effects (11, …

The Physiology And Biochemistry Of Isolated Skeletal Muscle Mitochondria : A Comparative Study, Mark Lowell Wagner

Dissertations and Theses

The physiological limit to maximum aerobic capacity (VO₂max) in vertebrates has been attributed to cardiovascular oxygen delivery, to the ability of the muscle cells to consume oxygen, or to a fine-tuned development of all components of the respiratory system such that no single component can be shown to limit VO₂max. The above hypotheses have each been developed using different experiments with different animals. The comparative studies uniting these animals and methods are limited. In order to further our knowledge of the cellular limit to VO₂max, skeletal muscle mitochondria were isolated from species representing four …

Biochemical Characterization Of The Axolemmal Mitogen For Cultured Schwann Cells, Mark A. Decoster

Theses and Dissertations

The molecule(s) involved at the axon plasma membrane (axolemma) which causes Schwann cells to proliferate has been investigated by three biochemical techniques: 1) Alkaline extraction of axolemma resulted in recovery of 95% of the mitogenic activity and 50% of the protein in the membrane-bound portion of the axolemma. 2) The axonal mitogen for Schwann cells may be associated with heparan sulfate proteoglycans at the axonal surface. Treatment of axolemma with heparitinase, which cleaves the glycosidic bonds of sulfated glycosaminoglycans, allowed mitogenic activity to be solubilized. 3) Treatment of axolemma with heparin, a highly sulfated glycosaminoglycan analogous to heparan sulfate, resulted …

Dietary And Hormonal Evaluation Of Men At Risk For Prostate Cancer, Donald J. Pusateri

Loma Linda University Electronic Theses, Dissertations & Projects

Relationships between dietary nutrients and plasma and fecal estrone, estradiol-17β, testosterone, and plasma prolactin levels were studied in young Seventh-day Adventist men: 18 nonvegetarians (NV), 20 lacto-ovovegetarians (LOV), and 15 vegans (V). Blood samples and 3 d dietary records were obtained. Contemporaneously collected diet composites and stool samples were analyzed for fiber. The vegetarians consumed significantly more fiber than the omnivores, while NVs and LOVs consumed more saturated fat than the Vs. Although plasma steroid hormone status did not differ, the Vs had significantly higher fecal estrogen levels than the NVs or LOVs. Plasma prolactin levels were significantly higher in …

A Method For The Large Scale Purification Of Hcg (Human Chorionic Gonadotropin) From The Urine Of Pregnant Women, Chun-Shyan Kuo

Chemistry & Biochemistry Theses & Dissertations

A large scale preparation of human chorionic gonadotropin (HCG) from the urine of pregnant women was accomplished by using kaolin adsorption, NH₄OH extraction and DEAE-cellulose ion exchange. A recovery about 81% of HCG and a specific activity in the 2,000-3,000 IU/mg range was achieved using this purification method.

Two HCG activity fractions, HCG I & HCG II, were isolated from DEAE-cellulose chromatography by a stepwise sodium chloride gradient elution. HCG I was eluted at 0.01-0.02 M NaCl while HCG II was eluted at 0.12-0.02 M NaCl.

The differences between HCG I and HCG II have been studied. The …

The Synthesis And Evaluation Of 5-Phenyloxazolidines As Potential Cardiovascular Drugs, Tammy C. Wang

Chemistry & Biochemistry Theses & Dissertations

Drugs such as adrenoceptor antagonists and certain centrally acting agents are known to lower blood pressure and thus serve as clinically useful antihypertensive agents. Norepinephrine 4, a natural ∝-adrenoceptor agonist, is the major agent associated with increased blood pressure in the periphery. Based on a careful structure-activity comparison, a series of substituted oxazolidines, 20, 21, 22 and 23, have been designed as possible antihypertensive candidates. Specifically, 22 and 23 have been synthesized to serve as potential norepinephrine antagonists.

Preliminary evaluation of these compounds indicated that they were not antihypertensive in nature. The more active compound, 23, …

An Analysis Of Protein-Ligand Interactions For Enzyme Ii-Mannitol From Escherichia Coli, Brent Lee Wood

Loma Linda University Electronic Theses, Dissertations & Projects

Enzyme II^mtl of Escherichia coli was purified from mtlA-overexpressing E. coli strain LJ1112 extraction of isolated membranes with deoxycholate, followed by hexyl agarose and ω-amino hexyl agarose chromatography in Lubrol PX. A 10-fold increase in total pure protein and 2.5-fold increase in specific activity over previously reported procedures was obtained. Tryptic fragments of Enzyme II^mtl were also purified and identified as the membrane and cytoplasmic domains by protein sequencing and SDS-PAGE. The fluorescence spectrum of Enzyme II^mtl suggested that two populations of tryptophan residues existed with emission maxima at 322 and 344 nm, corresponding to buried …

The Inhibition Of Bone Formation Occurring Under Weightlessness : The Effect Of Skeletal Unloading On Serum Levels Of A Bone-Derived Growth Factor, Jean D. Sibonga

Loma Linda University Electronic Theses, Dissertations & Projects

A deleterious effect of spaceflight on human health is the loss of bone associated with the prolonged lack of gravitational stress on the skeleton. A potential for bone loss is indicated by the hypercalciuria and decreased heel bone density determined in Skylab astronauts. Histological studies of juvenile laboratory rats in actual spaceflight and in simulation models document a suppression of bone formation with little alteration of bone resorption. The mechanism for net bone loss needs to be defined in order to develop measures for its prevention or correction.

Skeletal growth factors, which may mediate coupling and locally regulate bone volume …

The Characterization Of Ribosomal Rna Gene Chromatin From Physarum Polycephalum, Sally A. Amero, Vicky L. Montoya, Wendy L. Murdoch, Roy C. Ogle, John L. Keating, Robert M. Grainger

School of Medical Diagnostics & Translational Sciences Publications

We have isolated ribosomal RNA gene (rDNA) chromatin from Physarum polycephalum using a nucleolar isolation procedure that minimizes protein loss from chromatin and, subsequently, either agarose gel electrophoresis or metrizamide gradient centrifugation to purify this chromatin fraction (Amero, S. A., Ogle, R. C., Keating, J. L., Montoya, V. L., Murdoch, W. L., and Grainger, R. M. (1988) J. Biol. Chem. 263, 10725-10733). Metrizamide-purified rDNA chromatin obtained from nucleoli isolated according to the new procedure has a core histone/DNA ratio of 0.77:1. The major core histone classes comigrate electrophoretically with their nuclear counterparts on Triton-acid-urea/sodium dodecyl sulfate two-dimensional gels, although they …

A Light And Electron Microscopic Study Of The Rat Olfactory Tubercle: Normal Morphology And Acetylcholinesterase Localization, James Curtis Woodley

Biological Sciences Theses & Dissertations

A cytoarchitectural analysis of the rat olfactory tubercle using Nissl-stained coronal sections revealed that the dense cell layer (DCL) consisted of medium sized striatal cells in the cortical regions and small sized "granule" cells in the cap regions. Also delineated from this experiment was a rim of neuropil, nearly devoid of neurons as well as neuronal processes, outlining the islands of Calleja. Acetylcholinesterase (AChE) localization utilizing light microscopy revealed that the olfactory tubercles (OT) contained AChE-positive fibers that were orientated dorsoventrally in the molecular and multiform layers. The DCL consisted of only fibers en passant and putative terminals. Diisopropylfluorophosphate (DFP) …

Molecular Study Of The B19 (Human) Pathogenic Parvovirus, Jamshed Ayub

Theses and Dissertations in Biomedical Sciences

The B19 (human) parvovirus is a small single stranded DNA virus of 5.4 kilobases. B19 is specific for erythroid progenitor cells and has been propagated in vitro only with human erythroid bone marrow. Replication of viral DNA and the viral protein products of B19 appear similar to those of other animal parvoviruses. However, B19 differs from other parvoviruses in some important aspects, which include the initiation of all transcripts at a strong left side promoter (p6) and the absence of a functional internal promoter. B19 has an unusual transcription map which is described in this study.

The transcription map of …

Modulation Of Queuine Uptake In Cultured Human Fibroblasts By Phorbol Esters And Interferons, Debra L. Crane

Chemistry & Biochemistry Theses & Dissertations

Queuosine, a nucleoside found exclusively in the first position of the anticodon of transfer RNA (tRNA), is generated post-transcriptionally in an exchange of guanine for queuine by the modifying enzyme tRNA-guanine ribosyltransferase. Phorbol ester tumor promoters were shown to induce queuine hypomodified tRNA, and that phorbol ester action was due, in part, to inhibition of queuine transport across the cell membrane. An unidentified protein designated conditioned medium factor (CMF) that potentiated a phorbol-induced inhibition of queuine uptake was also documented. We suggest here that phorbol ester-induced inhibition of queuine uptake is not a significant factor in inducing queuine-deficient tRNA although …

Decreased Triiodothyronine Binding To The Hepatic Nuclear Thyroid Hormone Receptor In The Diabetic Mouse, Thomas J. Dewind

Loma Linda University Electronic Theses, Dissertations & Projects

The diabetic C57BL/KsJ-db m mouse has abnormal thyroid hormone levels and indications of thyroid hormone resistance. To investigate the basis of these abnormalities, the hepatic nuclear thyroid hormone receptor was extracted with 0.4 M KCl, 1.1 mM MgCl₂, 20 mM Tris/HCl, pH 7.9 from hepatocyte nuclei of normal C57BL/KsJ, heterozygous C57BL/KsJ-db m (db/m), and diabetic C57BL/KsJ-db m (db/db) mice. Normal and heterozygous mice were grouped together as the controls. Triiodothyronine (T₃) binding studies at 4°C using nitrocellulose filtration to separate free T₃ from receptor bound T₃ demonstrated an …

Characterization Of The Aquatic Environment In Lake Mead Near The Proposed Spring Canyon Pumped-Storage Project, And Assessment Of Potential Aquatic Impacts, Charles R. Liston, Stephen J. Grabowski, Bureau Of Reclamation

Publications (WR)

A pumped storage system consists of an upper reservoir and lower reservoir separated by an elevation difference. During low demand energy periods such as nights and weekends water is pumped from the lower to the upper reservoir using available energy from conventional steam electric power plants. During high energy demand periods, such as mornings and afternoons of weekdays, upper reservoir water is allowed to drop back down through the same system of water conduits and turbines, generating electricity to conveniently meet abrupt electrical energy requirements. The same water turbines thus act both as pumps and as conventional hydroelectric turbines. Because …

Kinetics Of Two-Substrate Reactions Using Integrated Rate Equations, Leonard D. Holmes

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The usual procedure employed in enzyme kinetic analysis is the method of initial rates. However, it has been appreciated for years that the analysis of enzyme-catalyzed reactions could, in principle, be more efficiently performed by examining the entire time course. There is much more information contained in a progress curve than in a simple initial rate. With the appearance of the computer, the formidable computations necessary for the use of integrated-rate equations are quite possible. The intention of this research was to develop the analytical and statistical methodology for applying an integrated-rate equation to a two-substrate reaction. I have analyzed …

The Bacterial Cell Membrane: Berlin Wall Of The Cell, Hans Kornberg

DSLS 1987-1988

(This information was taken from the Distinguished Scientist Lecture Series Program 1983-1984).

RECORDING UNAVAILABLE.

Born in Herford, Germany, Dr. Kornberg is Sir William Dunn Professor of Biochemistry, University of Cambridge, and Master of Christ's College, Cambridge. Dr. Kornberg received the B.Sc. and Ph.D. degrees from the University of Sheffield, the M.A. and D.Sc. degrees from the University of Oxford, and the Sc.D. degree from the University of Cambridge. He did postdoctoral research at Yale University, the University of California, Berkeley, and the Public Health Research Institute (New York City) before returning to England to join the Medical Research Council's Cell …

Use Of 52Cr For The Quantitative Determination Of Red Cell Volume, Claude Michael Hanbury

Chemistry & Biochemistry Theses & Dissertations

Chromium-52 has recently been suggested for use as a new agent to label red cells for the in vivo quantitation of red cell volume (1). In this paper, the development and validation of a routine ⁵²cr labeling procedure is described.

The accuracy, precision, and detection limits of chromium analysis by Zeeman effect atomic absorption spectroscopy was evaluated in the concentration range of 1 - 10 ug Cr/L.

Red cell chromium uptake was evaluated as a function of time, temperature, and concentration. Red cells labeled with a 2.5 mg/L chromium solution for 30 minutes at room temperature exhibited optimal label …

Reactions Of Aqueous Chlorine In The Gastric Fluid Of The Rat, Michael Gary Nickelsen

Chemistry & Biochemistry Theses & Dissertations

Gastric fluid was recovered from Sprague-Dawley rats that had been fasted for 24 hr or 48 hr and administered 4 mL deionized water. The protein, carbohydrate and chloride ion content of the gastric fluid was determined.

When gastric fluid is chlorinated with radiolabeled chlorine to concentrations between 323 and 375 mg/L, analyzed by high performance chromatography followed by liquid scintillation counting, as much as 59% of the radiolabeled chlorine was associated with organic components of the gastric fluid.

Model studies with the aromatic amino acids and the protein pepsin showed the formation of chlorinated analogs. Two chlorinated amino acid derivatives …

Laminin Receptors For Neurite Formation, H. K. Kleinman, Roy C. Ogle, F. B. Cannon, C. D. Little, T. M. Sweeney, L. Luckenbill-Edds

School of Medical Diagnostics & Translational Sciences Publications

Laminin, a basement membrane glycoprotein promotes both cell attachment and neurite outgrowth. Separate domains on laminin elicit these responses, suggesting that distinct receptors occur on the surface of cells. NG108-15 neuroblastoma-glioma cells rapidly extend long processes in the presence of laminin. We report here that 125I-labeled laminin specifically binds to these cells and to three membrane proteins of 67, 110, and 180 kDa. These proteins were isolated by affinity chromatography on laminin-Sepharose. The 67-kDa protein reacted with antibody to the previously characterized receptor for cell attachment to laminin. Antibodies to the 110-kDa and 180-kDa bands demonstrated that the 110-kDa protein …

Studies Of Transketolase Abnormality In Alzheimer's Disease / Kwan-Fu Rex Sheu, Phd; Donald D. Clarke, Phd; Young-Tai Kim, Phd; John P. Blass, Md, Phd; Bradford J. Harding; Joseph Decicco, Kwan-Fu Rex Sheu, Donald Dudley Clarke Phd, Young-Tai Kim, John P. Blass

Chemistry Faculty Publications

The partially purified transketolase from each of eight well-nourished patients with Alzheimer's disease contained significantly less heat-stable component with a significantly longer half-life of heat inactivation than that from eight controls. Immunochemical studies utilizing antibodies to the purified human liver transketolase did not distinguish between red blood cell transketolases of patients with Alzheimer's disease and those of controls. However, three brains from patients with Alzheimer's disease that were deficient in transketolase activity lacked a 69-kilodalton form on immunoblots. Subtle structural abnormalities of transketolase appear to occur in a high proportion of patients with Alzheimer's disease

The Purification Of Ribosomal Rna Gene Chromatin From Physarum Polycephalum, Sally A. Amero, Roy C. Ogle, John L. Keating, Vicky L. Montoya, Wendy L. Murdoch, Robert M. Grainger

School of Medical Diagnostics & Translational Sciences Publications

We have undertaken the purification of ribosomal RNA gene (rDNA) chromatin from the slime mold Physarum polycephalum, in order to study its chromatin structure. In this organism rDNA exists in nucleoli as highly repeated minichromosomes, and one can obtain crude chromatin fractions highly enriched in rDNA from isolated nucleoli. We first developed a nucleolar isolation method utilizing polyamines as stabilization agents that results in a chromatin fraction containing far more protein than is obtained by the more commonly used divalent cation isolation methods. The latter method appears to result in extensive histone loss during chromatin isolations. Two methods were then …

Improved Selective Medium For The Isolation Of Treponema Hyodysenteriae, Robert A. Kunkle, J. M. Kinyon

School of Veterinary and Biomedical Sciences: Faculty Publications

An agar medium with improved selection for Treponema hyodysenteriae was developed. Cultures of T. hyodysenteriae and T. innocens, feces from 11 clinically normal pigs, and colonic contents from 6 pigs with gross lesions consistent with swine dysentery were diluted in phosphate-buffered saline and plated on Trypticase soy agar (BBL Microbiology Systems, Cockeysville, Md.) with 5% citrated bovine blood (TSA), TSA with 400 ,ug of spectinomycin per ml (TSA-S400), TSA-S400 with 25 ,ug each of colistin and vancomycin per ml, and TSA with 5% pig feces extract and five antimicrobial agents (spiramycin, rifampin, vancomycin, colistin, and spectinomycin) (BJ). Viable numbers …

Comparison Of Cyclic Nucloetide Phosphodiesterase In Physarum Favicomum, C. Kay Holtman, Tammy K. Ebsen, Judith A. Bean, J. Scott Bryles, Thomas J. Lynch

Journal of the Arkansas Academy of Science

We have studied both cyclic AMP phosphodiesterase and cyclic GMP phosphodiesterase in the myxomycete Physarum flavicomum. The cyclic AMP phosphodiesterase preparations were isolated from both the diploid plasmodial stage of the lifecycle and the haploid myxamoebal stage. The plasmodial enzyme was prepared from spent medium (extracellular) and also from purified nuclei. The myxamoebal enzyme was prepared from purified nuclei. Cyclic GMP phosphodiesterase activity was studied in purified nuclei isolated from the plasmodium. One unusual feature of all the enzymes from the plasmodium is extreme heat stability; they remain catalytically active even after exposure to a boiling water bath for twenty …

Nucleotide Sequence Of The Fadr Gene, A Multifunctional Regulator Of Fatty Acid Metabolism In Escherichia Coli, Concetta C. Dirusso

Department of Biochemistry: Faculty Publications

The Escherichia coli fadR gene is a multifunctional regulator of fatty acid

and acetate metabolism. In the present work the nucleotide sequence of the

1.3 kb DNA fragment which encodes FadR has been determined. The coding

sequence of the fadR gene is 714 nucleotides long and is preceded by a typical

E. coli ribosome binding site and is followed by a sequence predicted to be

sufficient for factor-independent chain termination. Primer extension experiments

demonstrated that the transcription of the fadR gene initiates with an

adenine nucleotide 33 nucleotides upstream from the predicted start of

translation. The derived fadR peptide has …

Reduced Co2/O2 Specificity Of Ribulose-Bisphosphate Carboxylase/Oxygenase In A Temperature-Sensitive Chloroplast Mutant Of Chlamydomonas, Zhixiang Chen, Chris J. Chastain, Souhail R. Al-Abed, Raymond Chollet, Robert J. Spreitzer

Department of Biochemistry: Faculty Publications

The Chlamydomonas reinhardtii chloroplast mutant 68-4PP is phenotypically indistinguishable from wild type at 25°C but fails to grow photosynthetically at 35°C. It had about 30% of the wild-type level of ribulose-1,5-bisphosphate carboxylase/oxygenase (EC 4.1.1.39) holoenzyme and carboxylase activity when grown at 25°C, but less than 15% when grown at 35°C. Pulse-labeling with ³⁵S showed that the decrease in enzyme level at the restrictive temperature was not a result of reduced synthesis of enzyme subunits. The CO₂/O₂ specificity factor (V_cK_o/V_oK_c, where V_c and V_o are V_{max …}

Limnological Monitoring Data For Lake Mead During 1987: Technical Report No. 20, Larry J. Paulson

Publications (WR)

Limnological monitoring was conducted in Las Vegas Bay and Boulder Basin from April to December of 1987. The purpose of the monitoring was to (i) document possible changes in water quality resulting from decreased phosphorus loading in Las Vegas Wash, and (ii) establish a data base for evaluating the adequacy of water quality standards.