Biochemistry, Biophysics, and Structural Biology Commons^™

Extrathyroidal Human Biochemistry Of Monoiodotyrosine, Stanley Andrew Tan

Loma Linda University Electronic Theses, Dissertations & Projects

The existence of extrathyroidal monoiodotyrosine (MIT) has been debated because of uncertain results from measuring serum MIT. The reason is that the isotopic and chromatographic assays used have not been sensitive enough. This investigation undertook to establish the existence of MIT in serum utilizing a radioimmunoassay (RIA), to study the physiology of serum MIT, and to discover the pharmacologic effects of exogenous MIT.

MIT-binding antibody was raised in rabbits by immunization with an MIT-porcine thyroglobulin conjugate. The antiserum bound 40% of the labeled MIT at a final dilution of 1:1000. Incubation of 100 µ1 antiserum with 50 pg [^{125 …}

1-0-Alkyl-2-Acetyl-Sn-Glycero-3-Phosphorylcholine (Platelet Activating Factor): Interactions With Sheep Platelets, Duane O. Beaumont

Loma Linda University Electronic Theses, Dissertations & Projects

Platelet Activating Factor (PAF) seems to link platelet function with allergy and inflammation. Early events associated with the action of PAF on platelets are poorly understood, but PAF appears to interact with a platelet membrane receptor. Two photoaffinity-labelling analogs of PAF, diazoacetyl PAF and trifluorodiazoacetyl PAF, were designed for examining the interaction between PAF and its platelet receptor. These photoactive compounds were selected on the basis of the structure-activity relationship of PAF, and the general guidelines of the photoaffinity labelling technique. Some progress has been made in their synthesis and characterization.

The standard platelet aggregation assay was unreliable in testing …

The Effects Of Tri-O-Cresyl Phosphate On 2',3'-Cyclicnucleotide-3'-Phosphohydrolase Activity In Neural And Non-Neural Tissue Of The Rat, Mark K. Schlager

Chemistry & Biochemistry Theses & Dissertations

A total of 44 Long-Evans rats received 5.7 g/kg tri-O-cresyl phosphate (TOCP) orally every 3 days for 6 days. A second group of 13 rats received 4.6 g/kg orally every 3 days for 6 days. Animals were administered atropine sulfate (10 mg/kg) intraperitoneally every 6 hours or as needed to alleviate cholinergic symptoms. Ten control animals were given corn oil orally. Eight rats (18%) survived the 5.7 g/kg dose regimen and 5 rats (38%) survived the 4.6 g/kg dose regimen. Surviving rats were killed at 30-33 days post-dose. Signs of delayed neuropathy included ataxia and motor impairment.

The CNP activity …

Two-Dimensional Electrophoretic Studies Of Normal Spematozoal Proteins, Ying Lin

Chemistry & Biochemistry Theses & Dissertations

Protein profiles of human spermatozoa have been mapped by using high resolution two-dimensional electrophoresis which combines isoelectric focusing as the first dimension and polyacrylamide gradient gels electrophoresis in SDS as the second dimension. The profiles of normal spermatozoa! extraction, acrosomal extraction and seminal plasma showed high resolution in its separation capability. No qualitative or quantitative differences were observed by using Coomassie Brilliant Blue R-250 staining in a much higher concentration of proteins. However, coomassie blue lacks the sensitivity to detect many minor components. Therefore, a silver staining procedure was used, which resulted in greatly increased sensitivity.

Samples from normal fertile …

Time Course Of Enzyme Catalyzed Reactions: The Stoichiometry A --> P + Q, Tamra Tuttle

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The feasibility of using an integrated rate equation to analyze the kinetics of a second-order, enzyme-catalyzed reaction has been investigated. The inducible arginine decarboxylase from Escherichia coli B. was chosen for this study because it catalyzes an irreversible reaction with stoichiometry A --> P + Q, the simplest second order case. Values for five of the eight possible kinetic constants were determined from 21 time courses. Of the remaining three, the uncompetitive product inhibition constant for CO₂ was shown to be between 0 and 0.06, while the values of the competitive product inhibition for CO₂ and the uncompetitive …

Photoaffinity Labeling Of The Antimycin Binding Site In Rhodopseudomonas Sphaeroides, Emily Wilson

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The purpose of this study was to identify the site of interaction of antimycin with the ubiquinone-cytochrome b-c1 oxidoreductase in the photosynthetic bacteria, Rhodopseudomonas sphaeroides. To accomplish this goal, three areas of research were undertaken: the synthesis of a radiolabeled, photoaffinity analog of antimycin, identification of the inhibitory characteristics of this analog, and the photoaffinity labeling of the antimycin binding site. All three areas were accomplished.

The major finding of this study was the identification of an 11,000 dalton polypeptide as the predominantly labeled protein. Although this polypeptide was not exclusively labeled, it was consistently labeled and showed competition …

Investigation Of The Coenzyme Binding Site Of Lactate Dehydrogenase From Chicken Muscle, Jia-Hwei Lin

Chemistry & Biochemistry Theses & Dissertations

Investigation of the coenzyme binding site of lactate dehydrogenase was approached by using coenzyme analogs. Inhibition studies of this enzyme using n-alkylphosphates as coenzyme analogs were performed with a fluorometric method. The results showed that all of the n-alkylphosphates were competitive inhibitors with respect to NAD⁺.

The n-alkyl-ADPs were synthesized from AMP and dicycloheaylammonium salts of n-alkylphosphates. The synthetic compounds were subjected to NMR spectroscopic studies, thin layer chromatography and determination of the ratio of adenosine moiety to phosphate groups. The results indicated that the synthetic compounds were n-alkyl-ADPs.

These synthetic compounds were used as coenzyme analogs in …

A Method For The Comparison Of Hdl Containing Apoprotein E And Hdl Cholesterol, Steven A. Knizner

Chemistry & Biochemistry Theses & Dissertations

Human serum high density lipoproteins (HDL) were isolated by preparative salt density gradient ultracentrifugation and analyzed further by heparin-sepharose affinity chromatography. Separation of the major classes of serum lipoproteins was achieved after a single ultracentrifugation on a salt density gradient at 272,000 x g for 36 hours in a swinging bucket rotor. After isolation, the total HDL was subdivided into HDL containing apoprotein E and HDL without this apoprotein by heparin-sepharose affinity chromatography. Relative amounts of these subclasses were then calculated.

HDL-cholesterol was measured after precipitation of LDL and VLDL by phosphotungstic acid and magnesium. The statistical relationship between HDL …

Purification And Characterization Of A Camp- And Ca2+-Calmodulin-Independent Glycogen Synthase Kinase From Porcine Renal Cortex, Stephen J. Beebe, Erwin M. Reimann, Keith K. Schlender

Bioelectrics Publications

We recently reported the partial purification of a cAMP-independent and Ca²⁺-calmodulin-independent glycogen synthase kinase from porcine renal cortex (Schlender, K. K., Beebe, S. J., and Reimann, E. M. (1981) Cold Spring Harbor Conf. Cell Proliferation, 389-400). Subsequent purification indicated that the enzyme preparation consisted of at least three forms of glycogen synthase kinase which could be resolved by ATP gradient elution from aminoethylphosphate-agarose (AEP-agarose). The predominant form of glycogen synthase kinase, which eluted from AEP-agarose between 2 and 6 mM ATP, was purified approximately 800-fold and is designated GSK-A1. It had a molecular weight of 45,000-50,000 as determined …

Some Plant Leaves Have Orientation-Dependent Epr And Nmr Spectra, Douglas C. Mccain, Ted C. Selig, Govindjee, John L. Markley

Faculty Publications

Proton nuclear magnetic resonance (¹H NMR) spectra of leaves from 50 plant species were obtained at a spectrometer frequency of 470 MHz. Water present in leaf samples gives rise to characteristic spectral patterns. Most species show only one broad ¹H NMR peak; however, the leaves of some plants display complex, orientation-dependent spectra in which a common three-line pattern is discerned. The pattern varies with the angle between the leaf surface and the external magnetic field. Proton relaxation measurements show the presence of at least two water compartments in the leaves. The compartments are responsible for different components …

Potassium 1,3,3a,8a-Tetrahydro-3a,8a-Dihydroxy-2,8-Dioxoindeno[1,2-D]Imidazole-2-Ethanesulfonate, K+.C12h11n2o7s- / By D. D. Bray And D. D. Clarke Chemistry Department, Fordham University, Bronx, Nyew York 10458, Usa And N. Chatterjie And B. Sinha Institute For Basic Research In Development Disabilities, 1050 Forest Hill Road, Staten Island, New York 10314, Usa, Diana Bray, Donald Dudley Clarke Phd, Nithiananda Chatterjie, B. Sinha

Chemistry Faculty Publications

Mr = 366·40, monoclinic, P2/c, Z = 4, a=6·195 (1), b= 17·701 (4), c= 1.2·864 (3)A, P= 116·25(2)0 , V=l387·8A3, Dm=1·74, Dx= 1·754 Mg m-3, CuKa, l = 1·5418 A, ,u = 5·07 mm-1, T= 298 K, F(OOO) = 752, R = 0·074 for 2091 observed independent intensity data. Crystals were prepared by reacting the K salt of ureidotaurine with ninhydrin in acid solution. The product shows that the unsubstituted N -of the ureidotaurine reacts with the central C of ninhydrin

Cerebral Glutamine/Glutamate Interrelationships And Metabolic Compartmentation / S. Berl And D. D. Clarke, Soll Berl, Donald Dudley Clarke Phd

Chemistry Faculty Publications

The fact that in animals the concentration of glutamate and glutamine are higher in brain than in any other tissue initially attracted interest in attempting to understand the function or functions of these amino acids in the central nervous system. They were evidently required for protein synthesis and early studies established their close relationship to the Krebs tricarboxylic acid cycle and energy metabolism in brain. Glutamine formation in brain was also shown to be sensitive to elevations in plasma ammonia levels and to serve as the primary method for detoxifying this substance in brain in contrast to urea formation in …

Microheterogeneity Of Type Ii Camp-Dependent Protein Kinase In Various Mammalian Species And Tissues, Alison M. Robinson-Steiner, Stephen J. Beebe, Stephen R. Rannels, Jackie D. Corbin

Bioelectrics Publications

Excluding autophosphorylated species, at least six forms of the regulatory subunit of type II cAMP-dependent protein kinase (R(II)) from various mammalian tissues were identified by sodium dodecyl sulfate (SDS) gel electrophoresis of purified samples and of crude preparations photoaffinity labeled with 8-azido[32P] cAMP and by gel filtration. After autophosphorylation some heart R(II) forms termed type IIA (bovine, porcine, equine, and dog) shifted to a more slowly migrating band on SDS gels while others termed type IIB (rat, guinea pig, rabbit, and monkey) did not detectably shift. Both subclasses of R(II) exhibited variation in apparent M(r) on SDS gels. Bovine and …

Two Classes Of Camp Analogs Which Are Selective For The Two Different Camp-Binding Sites Of Type Ii Protein Kinase Demonstrate Synergism When Added Together To Intact Adipocytes, Stephen J. Beebe, Rob Holloway, Stephen R. Rannels, Jackie D. Corbin

Bioelectrics Publications

Twenty-five cyclic nucleotide analogs were tested individually to act as lipolytic agents and to activate adipocyte protein kinase. The lipolytic potency of individual analogs correlated better with their K(a) for protein kinase and their lipophilicity rather than with either parameters alone. Some of the most potent lipolytic analogs had high I50 values for the particulate low K(m) cAMP phosphodiesterase suggesting that their effect was not due to raising endogenous cAMP levels through inhibition of phosphodiesterase. The most potent lipolytic analogs contained a thio moiety at the C-8 or C-6 position. These analogs exhibited concave upward dose-response curves. At high concentrations …

The Marine Biogeochemistry Of Selenium: A Re-Evaluation, Gregory A. Cutter, Kenneth W. Bruland

OES Faculty Publications

Vertical and horizontal profiles from the North and South Pacific Oceans demonstrate the existence of three species of dissolved selenium: selenite, selenate, and organic selenide (operationally defined). In surface waters, organic selenide makes up about 80% of the total dissolved selenium, selenite concentrations are uniformly low, and selenate concentrations rise with increased vertical mixing. The organic selenide maximum (thought to consist of seleno-amino acids in peptides) coincides with the maxima of primary productivity, pigments, bioluminescence, and dissolved free amino acids. Deep ocean waters are enriched in selenite and selenate, while organic selenide is nondetectable. In suboxic waters of the tropical …

Comparison Of Surface Polysaccharides Of R. Leguminosarum Anu57 (Exo-1), R. Leg. Anu57(Pbr1an) (Nod+, Nif-), And R. Leg. Anu57(Pjb5j1) (Nod+, Nif-), Frank G. Hustmyer

Masters Theses

Extracellular polysaccharides (EPS) and lipopolysaccharides (LPS) are suspected to be involved in the legume/Rhizobium symbiotic process. Polysaccharide compositions from the EPS, LPS, capsular polysaccharide (CPS), and small polysaccharide (SmPS) were compared between Rhizobium leguminosarum ANU57 (parent, Exo^-1) and two mutants R. leg. ANU57(pBR1AN) (nod⁺, nif^-) and R. leg. ANU57(pJB5J1) (nod⁺, nif^-) both containing a Sym (symbiotic) plasmid insertion. R. leg. ANU57(pBR1AN) and R. leg. ANU57(pJB5J1) are nod⁺ on clover and peas, respectively. The nod⁺ mutants produce larger amounts of EPS and decreased amounts of LPS compared to …

Extracellular Phosphodiesterase From The Growth Medium Of The Myxomycete Physarum Flavicomum, Thomas J. Lynch, Mary E. Farrell

Journal of the Arkansas Academy of Science

The plasmodium of the myxomycete Physarum flavicomum secretes cyclic AMP phosphodiesterase into the medium. The extracellular enzyme had a pH optimum between 7 and 8 and a Km of about 500 μM cyclic AMP and was inhibited by theophylline, caffeine and 3-isobutyl-l-methyxanthine (MIX). A marked decrease of enzyme activity was noted in the presence of EDTA, suggesting the requirement of Mg+ + by the enzyme. Addition of Mg+ + and Ca + + stimulated the enzyme while Zn+ + , Co+ + , Pb+ + , Mn+ + , Fe + + + , Ni+ + , and Cu + …

Evaluation Of Peptide Assimilation In Ehrlich Ascites Tumor Cells, John M. Castellano

Biological Sciences Theses & Dissertations

The dipeptides glycyl-L-(³H)-leucine and L-phenylalanyl-L-(³H)-leucine were synthesized in radioactive form and utilized in the analysis of peptide transport and hydrolysis by Ehrlich ascites tumor cells. The tumor cells were cultured in the abdominal cavity of CF-1 albino mice and harvested into sterile isotonic buffer for all experimental analyses.

The results of this study reveal that the tumor cells rapidly accumulate radiolabel when incubated with the above mentioned peptides, but that the peptide is most probably hydrolyzed extracellularly and free L-(³)-leucine transported. This hypothesis is based on the observations of reduction in accumulation of radiolabel …

Reduction Of Ferric Leghemoglobin In Soybean Root Nodules, Keuk-Ki Lee, Robert V. Klucas

Department of Biochemistry: Faculty Publications

Reduction of ferric leghemoglobin to ferrous leghemoglobin in soybean nodules (Glycine max [L.] Meff. cv Woodworth) was studied using a spectrophotometer equipped with an in-cell space diffuse reflectance accessory. Nodule slices prepared and scanned under nitrogen gas showed a ferrous leghemoglobin absorption spectrum. Nodule slices equilibrated with 100% O₂ or air exhibited two absorption bands characteristic of oxygenated leghemoglobin. The addition of CO shifted those bands to CO leghemoglobin absorption bands. Potassium ferricyanide was not effective in oxidizing ferrous to feffic leghemoglobin in nodule slices. However, ferric leghemoglobin was formed by treating the nodule slices with hydroxylamine, and …

The Role Of Recognition In Plant Disease, J. M. Daly

Department of Biochemistry: Faculty Publications

As noted by Sequeira (110,111), the term recognition has become very popular in the literature of host-pathogen interactions over the last ten years. Despite recent facile usage, its meaning is not always clear to me. Recognition is defined by Clarke & Knox (16) as the "initial event in cell-cell communication that elicits a defined biochemical, physiological, or morphological response." This requires the not-so-easy task of defining "initial event" and "communica­ tion" with precise chemical or physical mechanisms. For pathologists, Se­ queira (110) has provided a more restricted meaning: "An early specific event that triggers a rapid, overt response by the …

Las Vegas Wash Advanced Water Quality Study: Final Report, Richard A. Roline, James J. Sartoris, U.S. Bureau Of Reclamation

Publications (WR)

The purpose of the Las Vegas Wash Advanced Water Quality Study is to determine the existence, extent, and mechanisms of nutrient and toxin stripping in Las Vegas Wash under present conditions and under future conditions, both with and without construction of the proposed salinity control unit as described by the Bureau of Reclamation (USBR, 1982b). This study was performed for the Lower Colorado Region Division of Planning by personnel of the Environmental Sciences Section of the Division of Research and Laboratory Services, E&R Center, Denver, Colorado. Work on the study began in February 1983. The general approach adopted for this …

The Synthesis Of Various Substituted 3-Amino-7-Hydroxy-2,2-Dimethyltetralins And Their Opioid-Related Activities, David Alan Lippman

University of the Pacific Theses and Dissertations

A series of aminotetralones and aminotetralins were synthesized from the common intermediate F, 3-amino-2,2-dimethyl-7-methoxy- 1-tetralone. The final compounds derived from F were simple substituted and/or reduced analogues. The products would allow a progressive structure activity relationship to be drawn based on pharmacological testing.

The common intermediate F was synthesized utilizing a six step procedure starting with p-methoxyphenylacetic acid. The overall yield from the precursor to the F:HC1 was 25%. Compound F was either O-demethylated to form 3-amino-2,2-dimethyl-7-hydroxy-l-tetralone (I) or was dimethylated on the amine and subsequently O-demethylated to yield the 3-dimethylamino-7-hydroxy-2,2-dimethyl-l-tetralone (J). The last major modification was the reduction of …

Isolation Of A Bacterium Capable Of Degrading Peanut Hull Lignin, Thomas J. Kerr, Robert D. Kerr, Ronald Benner

Faculty Publications

No abstract provided.

High Resolution Two Dimensional Electrophoresis Of Soluble Proteins In Human Spermatozoa, Hsing-Ying Liu

Chemistry & Biochemistry Theses & Dissertations

The evaluation of human semen is very important for diagnosis of male infertility. Combined with the analysis of seminal plasma, a comprehensive biochemical screening of human spermatozoan proteins may be of assistance in providing qualitative and quantitative information to the physician to assist with evaluation.

In this study, human spermatozoa were ruptured by sonication. The soluble spermatozoan proteins were separated from the cellular debris by centrifugation and then analyzed in dissociated forms by two dimensional electrophoresis. The separated proteins were visualized by a silver stain method.

It has been shown that these combined techniques can resolve- four to five hundred …

Determination Of N,N'-Diphenylguanidine In Biological Tissues And Fluids Using High Pressure Liquid Chromatography, Edward Sidney Hunter Iii

Chemistry & Biochemistry Theses & Dissertations

N,N'-Diphenylguanidine (DPG) is a potentially toxic compound used in the vulcanization of rubber products. Distribution of DPG in hybrid mice (C57BL/6J interbred with DAB₂) was determined following intraperitoneal injection. Urine, fecal material, whole blood and tissues were collected at various times after administration of DPG and the extracts analyzed for DPG by high pressure liquid chromatography. Recovery of DPG from spiked urine and whole blood exceed 90%, however, the recovery of DPG from spiked tissue samples was approximately 70%.

DPG concentration in whole blood reached a maximum within ten minutes and rapidly decreased through 180 minutes after administration …

Identification Of Four Complementary Rna Species In Akabane Virus-Infected Cells, Asit K. Pattnaik, G. Abraham

School of Veterinary and Biomedical Sciences: Faculty Publications

The analysis of RNA extracted from purified Akabane virus demonstrated the presence of three size classes of single-stranded RNAs with sedimentation coefficients of 31S (large, L), 26S (medium, M), and 13S (small, S). Molecular weights of these RNA species were estimated to be 2.15 x 10⁶,1.5 X 10⁶, and 0.48 x 10⁶ for the L, M, and S RNAs, respectively. Hybridization analysis involving viral genomic RNA and RNA from virus-infected cells resulted in the identification of four virus-specific cRNA species in infected cells. These cRNAs were found to be nonpolyadenylated by their inability to bind …

A Survey Of Glycosphingolipids In The Human Aorta: Their Relationship To Glycosphingolipids Of Blood And To Atherosclerosis, John L. Nappier

Dissertations

The four major glycosphingolipids, glucosyl ceramide, lactosyl ceramide, galactosyl lactosyl ceramide and globoside were isolated from both diseased and normal tissue from human aorta by solvent extraction followed by solvent partitioning and column chromatography. The levels of each were then determind quantitatively by high pressure liquid chromatography (HPLC) after derivatization. The individual glycosphingolipids were isolated by semi-preparative HPLC and the fatty acid composition of each was determined, after methanolysis, by gas chromatography.

In the diseased tissue, the amount of glycosphingolipid as well as the amount of crude lipid increased relative to the normal tissue. The relative amounts of cerebrosides increased …

Determination Of Erythrocyte Zinc Concentrations Using Zeeman Effect Atomic Absorption Spectroscopy, Tanya Maranda Lewis

Chemistry & Biochemistry Theses & Dissertations

The purpose of this research project was to develop assay procedures for determination of zinc in erythrocytes. Three methodologies were developed. The methods were evaluated on the basis of within-run and between-run precision. The most reproducible method, the 213.856 nm flame methodology, was used to determine red blood cell zinc concentrations in more than 200 samples obtained from the Neonatal Intensive care Unit at the Children's Hospital of the King's Daughters.

This study demonstrated that erythrocyte zinc levels in premature infants were similar to those of healthy adults. A linear relation between plasma and red blood cell zinc concentrations was …

Hemoglobins Of The Cutthroat Trout Salmo Clarki, Jonathan N. Southard

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Nine hemoglobins have been isolated from the blood of cutthroat trout. All nine hemoglobins bind oxygen cooperatively and appear to be tetramers with molecular weights of -64,000. The oxygen equilibria and subunit structures of the purified hemoglobins were studied. In addition, the red blood cells of cutthroat trout were examined for the presence of ATP and GTP, which are known to be physiological modulators of hemoglobins in fishes.

Five hemoglobins with isoelectric points from 9.1 to 7.0 are classified as cathodal hemoglobins. These five hemoglobins have identical oxygen binding properties by the criteria tested. All have oxygen equilibria which are …

Subunit Interactions In The Inducible Arginine Decarboxylase From Escherichia Coli B, Catalina N. Depusoy

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

The nature of the subunit interactions in the inducible arginine decarboxylase from Escherichia coli B is of considerable interest because of the observed differences in the catalytic activities of the dimer and the decamer; the decamer is active and the dimer is inactive. To study these interactions, inactive dimers were prepared by sodium borohydride reduction of the E-amino--pyridoxal-P Schiff base. Hybrid decamers were then prepared from varying molar ratios of native and reduced dimers. The hybrid decamers were indistinguishable from native decamers as observed in the analytical ultracentrifuge and on acrylamide gel electrophoresis. Kinetic studies indicated that true hybrids were …