Biochemistry Commons^™

The Cytotoxic Effect Of Tropical Plants On Human And Rat Hepatoma Cells, Mark Tafazoli

Honors Capstone Projects and Theses

No abstract provided.

Gal4 Disrupts A Repressing Nucleosome During Activation Of Gal1 Transcription In Vivo, Jeffrey D. Axelrod, Michael S. Reagan, John Majors

Biology Faculty Publications

Photofootprinting in vivo of GALl reveals an activation- dependent pattern between the UASG and the TATA box, in a sequence not required for transcriptional activation by GAL4. The pattern results from a nucleosome whose position depends on sequences within the UASG. In the wild-type gene, activation by GAL4 and derivatives disrupts this nucleosome. This activity is independent of interactions with DNA-bound core transcription factors and is proportional to the strength of the activator. Presence of the nucleosome correlates with low basal transcription levels under various conditions, suggesting a role in limiting basal expression. We propose a role for the GAL4 …

Human Lung Mast Cell Tryptase Isozymes: Separation And Examination Of Structural And Functional Differences, Susan S. Little

Electronic Theses and Dissertations

Tryptases are trypsin-like enzymes found in mast cell granules. Although in vivo substrates have not been positively identified, tryptases cleave a limited number of potential physiological substrates in vitro, including high molecular weight kininogen (HMWK) and vasoactive intestinal peptide (VIP). Purified human lung mast cell tryptase (HLT) apparently exists as a tetramer with an M$\sb{\rm r}$ of 135-144 kDa by gel filtration, whereas SDS-PAGE yielded two bands of M$\sb{\rm r}$ 29 Kda and 33 Kda. Tryptases are resistant to inhibition by most natural trypsin inhibitors and display some affinity for heparin. The existence of tryptase isozymes has been implied from …

Probing Protein-Protein Interactions Among Proteins Of A Nonaggregated Fatty Acid Synthetase From Euglena Gracilis Variety Bacillaris, Sande G. Williams

Electronic Theses and Dissertations

Enoyl-acyl carrier protein (ACP) reductase from chloroplast nonaggregated fatty acid synthetase (FAS) of Euglena gracilis variety bacillaris was purified to a single band on a denaturing polyacrylamide gel. The enzyme was partially characterized with respect to substrate specificity, reduced nucleotide requirement, and the effect of ACP and Ca$\sp{++}$ on enzyme activity. Antibodies against the purified protein were raised in hens and isolated from eggs. ACP was purified from Euglena in yields of about 1mg/100g (wet weight) of cells. Antibodies were raised against the purified protein. ACP antibodies inhibited the Euglena chloroplast FAS using Euglena or E. coli ACP as a …

Longitudinal Study Of Plasma And Erythrocyte Selenium Levels In Type I Diabetic Children During Development And Treatment Of Ketoacidosis, Pi-Hsia Fan

Chemistry & Biochemistry Theses & Dissertations

Most studies on the biological activity of selenium have focused on its role at the active site in the ubiquitous enzyme, glutathione peroxidase (GSH-Px) which catalyzes the removal of hydrogen peroxide and organic hydroperoxides (1). A connection between selenium deficiency and a wide range of diseases in animals and humans has been reviewed (2). Development of various forms of human cancer (3), failure of the immune system (4), as well as some cardiovascular diseases (5) have been reported or suspected to correlate with selenium deficiency. However, the physiological function of selenium in these different diseases is not fully understood, and …

Analysis Of The Proteoglycan Content In Fresh And Cryopreserved Porcine Cardiovascular Tissues, Yun Hee Shon

Theses and Dissertations in Biomedical Sciences

The purpose of this research was to study the effects of cryopreservation on the proteoglycan (PG) and mineral content of aorta conduit tissue. Proteoglycans from fresh and cryopreserved porcine aorta conduit tissues were isolated by extraction with 4 M guanidine (Gdn) - hydrochloride (HCl) for 48 hours. Quantitative analysis of extracted proteoglycans revealed that the content of proteoglycan material from cryopreserved tissue, measured as uronate and protein per unit weight of wet tissue, was similar to that from fresh tissue. Gel permeation column chromatography studies suggested that proteoglycans present in three CsCl fractions from cryopreserved tissues had molecular weights similar …

Separation And Quantification Of Queuine-Modified And Unmodified Transfer Rna Isoacceptors, Jorge Miguel Pereira De Oliveira Da Silva Santos

Chemistry & Biochemistry Theses & Dissertations

The discovery of a base-exchange modification in the first position of the anticodon of four transfer RNA (tRNA) isoacceptor families has been shown to be correlated with a decrease in the rate of cell division in several tumorigenic tissues. This fact led to the hypothesis that wobbling in the anticodon might be a control point wich mediates the rate of translation. A suitable method for separation of different tRNA isoacceptor families had to be created in order to generate a means for quantitatively measuring degrees of this base modification in tRNA. Radio labelled amino acids were used to charge tRNA …

Hepatic Lipase Treatment Of Chylomicron Remnants Increases Exposure Of Apolipoprotein E, Dawn L. Brasaemle, Kathleen Cornely-Moss, André Bensadoun

Chemistry & Biochemistry Faculty Publications

The consequences of hepatic lipase treatment of chylomicron remnants were studied. Rats were fed corn oil to induce production and secretion of chylomicrons and were then injected with polyclonal antiserum raised against hepatic lipase to specifically and quantitatively inhibit hepatic lipase activity in vivo. A fraction enriched in chylomicron remnants was isolated from rat plasma by a brief centrifugation step that preferentially isolates triglyceride-rich apolipoprotein (apo) B-48-containing lipoproteins. The chylomicron remnants were then treated with hepatic lipase in vitro, or incubated under identical conditions in the absence of enzyme (control incubations). Hepatic lipase-treated and control chylomicron remnants were isolated by …

Mouse Heparin Binding Protein-44 (Hbp-44) Associates With Brushin, A High-Molecular-Weight Glycoprotein Antigen Common To The Kidney And Teratocarcinomas, Masaru Nakamoto, Masayuki Ozawa, Sonia Jacinto, Tatsuhiko Furukawa, Yasuhiro Natori, Hiroshi Shirahama, Suguru Yonezawa, Tatsuo Nakayama, Takashi Muramatsu

Biology Faculty Publications

Heparin binding protein-44 (HBP-44) is a heparin binding protein of 44 kDa, found by cDNA cloning using antibodies against teratocarcinoma glycoproteins [Furukawa, T. et al. (1990) J. Biochem. 108, 297-302]. The N-terminal sequence analysis reported in this publication establishes the structure of its mature form. Immunohistochemical staining revealed that HBP-44 was located in the tubular brush border of the kidney. HBP-44 formed a complex with brushin, a high molecular weight (460 kDa) glycoprotein antigen common to the kidney and teratocarcinoma, but not with OR8 antigen, another antigen (350 kDa) of the same category. Brushin was shown to be the mouse …

Investigation Of The Structural Heterogeneity Of Lipooligosaccharides From Pathogenic Haemophilus And Neisseria Species And Of R-Type Lipopolysaccharides From Salmonella Typhimurium By Electrospray Mass Spectrometry, B W. Gibson, William Melaugh, Nancy J. Phillips, M A. Apicella, A A. Campagnari, J M. Griffiss

Chemistry Faculty Publications

Heterogeneity in the lipooligosaccharides (LOS) of pathogenic Haemophilus and Neisseria species is evident from the multiplicity of components observed with electrophoretic analyses. Knowledge of the precise structures that make up these diverse LOS molecules is clearly the key to reaching an understanding of pathogenic processes such as phase variation and molecular mimicry. Except for a few cases, little is known about the specific structural features of LOS that underlie phase variation and molecular mimicry, partly because of the inherent difficulties in the structural elucidation of these complex glycolipids. In the lipopolysaccharides (LPS) from Salmonella typhimurium and Escherichia coli, rough, or …

Reexamination Of The Thermolytic Rearrangement Of 4-Halophenyl Azides To 2-Aminophenols And Other Products / H. Gershon, D. D. Clarke, And M. Gershon, Department Of Chemistry, Fordham University, Bronx, Ny 10458, Usa, New York Botanical Garden, Bronx, Ny 10458, Usa, Herman Gershon, Donald Dudley Clarke Phd, Muriel Gershon

Chemistry Faculty Publications

The halogenation of derivatives of 2-aminophenol with N-chloro- and N-bromosuccinimides at ambient temperatures in acetic acid was studied. With the necessary compounds available, a reexamination of the thermolytic rearrangement of 2-halophenyl azides to 2-aminophenols and other products was undertaken. It is certain that the rearrangement of 4-halophenyl azides to 2-aminophenols occurs but the products identified in this study differ significantly from those reported previously by Suschitzky et al. (1963, 1966)

A Comparative 2-Dimensional Gel Protein Database Of The Intact And Regenerating Newt Limbs, Panagiotis A. Tsonis

Biology Faculty Publications

In this paper we describe a two-dimensional gel database of the regenerating newt limb. Protein synthesis was compared in the intact limb, in the 1-week regenerating limb, representing the dedifferentiation stage, and in the 2-week regenerating limb, representing the formation of the blastema. This comparative database provided data on differential expression of about 800 proteins during the process of limb regeneration. In addition, a map has been generated for these proteins for future guidance in characterizing further new, unknown proteins. The overall expression patterns of the proteins indicated that the dedifferentiation stage was marked by down-regulation of most proteins, while …

Exopeptidase Catalyzed Site-Specific Bonding Of Supports, Labels And Bioactive Agents To Proteins, Fred W. Wagner, Thomas R. Coolidge, Sheldon M. Schuster, Jay Stout, Dwane E. Wylie, Klaus Breddam, William Lewis

Department of Biochemistry: Faculty Publications

An auxiliary substance such as a label, support, or bioactive agent is attached to a protein at a site that is remote from the active site of the protein by the use of exopeptidase and a nucleophile which is an amino acid, amino acid derivative, amine or alcohol. In one embodiment, the nucleophile is attached to the carboxy terminus of a protein by catalysis with exopeptidase to form an adduct and then the adduct or its combination with a linker arm is bound to the auxiliary substance. In another embodiment, the auxiliary substance or its combination with a linker arm …

Purification And Ligand Binding Of A Soluble Class I Mhc Molecule Consisting Of The First Three Domains Of H-2kd Fused To B2-Microglobulin Expressed In The Baculovirus/Insect Cell System, Francois Godeau, Immanuel F. Luescher, David M. Ojcius, Cecile Saucier, Estelle Mottez, Lucien Cabanie, Philippe Kourilsky

All Dugoni School of Dentistry Faculty Articles

A recombinant baculovirus encoding a single-chain murine major histocompatibility complex class I molecule in which the first three domains of H-2Kd are fused to beta 2-microglobulin (beta 2-m) via a 15-amino acid linker has been isolated and used to infect lepidopteran cells. A soluble, 391-amino acid single-chain H-2Kd (SC-Kd) molecule of 48 kDa was synthesized and glycosylated in insect cells and could be purified in the absence of detergents by affinity chromatography using the anti-H-2Kd monoclonal antibody SF1.1.1.1. We tested the ability of SC-Kd to bind antigenic peptides using a direct binding assay based on photoaffinity labeling. The photoreactive derivative …

Identification Of The Nd Δ And Σ States And The 1,3Φ←←X  3Σ− G Transition Of O2 By Resonant Multiphoton Ionization, Robert J. Yokelson, R. J. Lipert, W. A. Chupka

Chemistry and Biochemistry Faculty Publications

Spectra of the 3dRydberg state region of O₂ have been obtained by two‐photon resonant ionization of the ground electronic state. By varying the rotational distribution and radiation polarization, all observed bands were identified and attributed to excitation of Σ, Δ, and Φ states. Earlier assignments were corrected. The Δ and Φ assignments are complete while the Σ assignments are so far incomplete.

Identification Of The Ns And Nd Rydberg States Of O2 For N=3–5, Robert J. Yokelson, R. J. Lipert, W. A. Chupka

Chemistry and Biochemistry Faculty Publications

The 4s‐3d and 5s‐4dRydberg complexes of diatomic oxygen have been studied by (2+1) resonance‐enhanced multiphoton ionization of the X  ³∑ _g ⁻ ground state of O₂. We have located and identified at least two vibrational levels of each of the following states: Three of four expected 4sσ Π states; all four expected 5sσ Π states; 18 of 22 expected 3d states (with only the states of the 3dσ orbital remaining unobserved); and 5 of the 10 predicted 4dπ states. State assignments …

Circular Dichroism And Molecular Modeling Yield A Structure For The Complex Of Human Immunodeficiency Virus Type 1 Trans-Activation Response Rna And The Binding Region Of Tat, The Trans-Acting Transcriptional Activator, Erwann P. Loret, Philippe T. Georgel, W. Curtis Johnson Jr., Pui Shing Ho

Biological Sciences Faculty Research

Transcription in the human immunodeficiency virus type 1 (HIV-1) retrovirus is regulated by binding the viral Tat protein (trans-acting transcriptional activator) to the trans-activation response (TAR) RNA sequence. Here, vacuum UV circular dichroism (VUV-CD) is used to study the structure of TAR and its complex with two peptide fragments that are important for Tat binding to TAR. The VUV-CD spectrum of TAR is typical of A-form RNA and is minimally perturbed when bound to either the short or the long Tat peptide. The CD spectra ofthe complexes indicate an extended structure in the argnine-rich region of Tat from amino acid …

Structural Characterization Of Fucoidan And Its Role In The Hemizona Assay, Manish S. Patankar

Chemistry & Biochemistry Theses & Dissertations

Fucoidan is a polysaccharide commonly found in brown algae. It is a sulfated polymer of the monosaccharide L-fucose. Fucoidan is thought to function in water retention by algae. Commercially available fucoidan is extracted from a brown algae called Eurus vesiculosus.

This polysaccharide has immense potential as a biologically active agent: (i) it exhibits anticoagulant action, (ii) it has also been reported to block retroviral replication and (iii) it is capable of blocking sperm-egg binding in sea urchin, mouse and rabbit. Recently, fucoidan has also been shown to block the interaction of human sperm and egg in the hemizona assay …

The Structure Of A Complex Of Bovine Ɑ-Thrombin And Recombinant Hirudin At 2.8-Å Resolution, Jacqueline Vitali, Philip D. Martin, Michael G. Malkowski, William D. Robertson, Jerome B. Lazar, Richard C. Winant, Paul H. Johnson, Brian Fp Edwards

Biochemistry and Molecular Biology Faculty Publications

Crystals of the complex of bovine alpha-thrombin with recombinant hirudin variant 1 have space group C222(1) with cell constants a = 59.11, b = 102.62, and c = 143.26 A. The orientation and position of the thrombin component was determined by molecular replacement and the hirudin molecule was fit in 2 magnitude of Fo - magnitude of Fc electron density maps. The structure was refined by restrained least squares and simulated annealing to R = 0.161 at 2.8-A resolution. The binding of hirudin to thrombin is generally similar to that observed in the crystals of human thrombin-hirudin. Several differences in …

Saccharomyces Cerevisiae Sec59 Cells Are Deficient In Dolichol Kinase Activity, Loree Heller, Peter Orlean, W. Lee Adair Jr.

Bioelectrics Publications

The temperature-sensitive Saccharomyces cerevisiae mutant sec59 accumulates inactive and incompletely glycosylated protein precursors in its endoplasmic reticulum at the restrictive temperature. O-mannosylation and glycosyl phosphatidylinositol membrane anchoring of protein are also abolished, consistent with a deficiency in dolichyl phosphate mannose. Membranes prepared from sec59 cells that had been shifted to the restrictive temperature, however, made normal amounts of dolichyl phosphate mannose when exogenous dolichyl phosphate was supplied, but dolichyl phosphate mannose synthesis was severely depressed in the absence of exogenous dolichyl phosphate. Quantitative measurements of dolichyl phosphate in sec59 cells showed that the levels were decreased to 48% of wild …

Isolation And Characterization Of The Triiodothyronine Regulatory Region Of The Fatty Acid Synthase Gene In Avian Species, Delfina Rivera Valencia

Masters Theses

Fatty acids are utilized for energy storage in the form of triglycerides or neutral fats. One way to regulate fatty acid biosynthesis is to regulate the activity of fatty acid synthase (FAS), the enzyme that catalyzes the reaction. Previous studies have shown that regulation of FAS activity and synthesis is regulated by the hormone triiodothyronine, T₃, at the level of transcription. Thus, isolation and characterization of the promoter region o f the FAS gene will enable the identification of the thyroid hormone response elements (TREs). This study could shed light on the mechanisms on how this gene can …

Appearance Of A Methotrexate Binding Plasma Protein During High Dose Methotrexate Therapy, M P. Iqbal, I Malik, N Mehboobali

Department of Biological & Biomedical Sciences

No abstract provided.

Biochemical Investigation Of Gout And Its Familial Incidences, Chin-Ling Hsu

Chemistry & Biochemistry Theses & Dissertations

Gout is a chronic metabolic disorder caused by deposition of uric acid on the joint. It is categorized into two types: primary and secondary. Primary gout is uric acid overproduction, caused by excessive synthesis of the purine precursors. Secondary gout occurs also as the result of overproduction or decreased renal excretion of uric acid resulting from other disease processes or medication.

The two purine bases, hypoxanthine and xanthine, liberated from ribosides by the action of nucleoside phosphorylase, are degraded to uric acid as the final excretory product in the purine degradation pathway. Hypoxanthine and xanthine are the physiological substrates of …

Kinetic Characterization Of A Recombinant C-Terminal Mutant Of Reverse Transcriptase From The Human Immunodeficiency Virus, Thomas S. Heard

Chemistry & Biochemistry Theses & Dissertations

The human immunodeficiency virus (HIV) reverse transcriptase (RT) (EC 2.7.7.49) is the central replication enzyme for HIV. In general, the kinetic mechanism for this and all other polymerases involves the ordered binding of two substrates: a primer-template (PT) followed by a deoxyribonucleoside triphosphate (dNTP). Previous investigations prompted this research when it was discovered that the substrate dNTP, in absence of PT, could protect a recombinant c-terminal mutant HIV-1 RT from inhibition by pyridoxal-5'-monophosphate (PLP), an active-site dNTP inhibitor. In contrast, the non-mutant recombinant HIV-1 RT required both substrates for protection from PLP inhibition. This investigation sought to determine if this …

The Structure Of Residues 7-16 Of The Aɑ-Chain Of Human Fibrinogen Bound To Bovine Thrombin At 2.3 Å Resolution, Philip D. Martin, William Robertson, Dusan Turk, Robert Huber, Wolfram Bode, Brian Fp Edwards

Biochemistry and Molecular Biology Faculty Publications

The tetradecapeptide Ac-D-F-L-A-E-G-G-G-V-R-G-P-R-V-OMe, which mimics residues 7f-20f of the A alpha-chain of human fibrinogen, has been co-crystallized with bovine thrombin from ammonium sulfate solutions in space group P2(1) with unit cell dimensions of a = 83.0 A, b = 89.4 A, c = 99.3 A, and beta = 106.6 degrees. Three crystallographically independent complexes were located in the asymmetric unit by molecular replacement using the native bovine thrombin structure as a model. The standard crystallographic R-factor is 0.167 at 2.3-A resolution. Excellent electron density could be traced for the decapeptide, beginning with Asp-7f and ending with Arg-16f in the active …

Methotrexate As An Alternative To Surgery In Ectopic Pregnancy: A New Role For An Old Drug, M P. Iqbal

Department of Biological & Biomedical Sciences

Abstract is not provided by the author/publisher

Biochemical Analysis Of Human Seminal Plasma, Jamshed Ayub

Chemistry & Biochemistry Theses & Dissertations

Presently there is no single biochemical criterion for differentiating between a potentially fertile male and an infertile male. However, for evaluating male fertility the biochemical analysis of seminal plasma is a very convenient and quantitative technique if it is combined with careful clinical examination.

In seminal plasma analysis, fructose and proteins are chemical indicators of seminal vesicle activity, citric acid and acid phosphatase indicate prostate function while glycerolphosphorylcholine indicates epididymis activity. In this research these five indicators are studied and an attempt is made to establish their normal ranges.

This study shows that the data obtained from the seminal plasma …

Estimation And Separation Of N-Acetyl-Beta-D-Glucosaminidase Isoenzymes In Urine, M H. Javed, A N. Hussain, M A. Waqar

Department of Biological & Biomedical Sciences

N-acetyl-beta-D-glucosaminidase is a lysosomal enzyme made up of two isoenzymes (A and B). It has been used extensively as a marker for kidney damage. However, its estimation in urine has not been standardized. We have established a method for the estimation and separation of NAG isoenzymes by ion-exchange chromatography. In 19 experiments done so far, this method has given reproducible results. The significance of this method is that with a single experiment, one can estimate total as well as individual isoenzyme activity. Furthermore, urine constituents do not appear to interfere in this assay.

Structural Changes That Accompany The Reduced Catalytic Efficiency Of Two Semisynthetic Ribonuclease Analogs, V. Srini J. De Mel, Philip D. Martin, Marilynn S. Doscher, Brian Fp Edwards

Biochemistry and Molecular Biology Faculty Publications

The structures of two catalytically defective semi-synthetic RNases obtained by replacing aspartic acid 121 with asparagine or alanine have been determined and refined at a resolution of 2.0 A (R = 0.186 and 0.172, respectively). When these structures are compared with the refined 1.8-A structure (R = 0.204) of the fully active aspartic acid-containing enzyme (Martin, P.D., Doscher, M.S., and Edwards, B. F. P. (1987) J. Biol. Chem. 262, 15930-15938), numerous and widespread changes, much greater in number and magnitude than the small structural variations noted previously between the semisynthetic complex and RNase A, are found to have occurred. These …

Studies On Photoaffinity Labeling And Mass Spectrometry Of Bacteriorhodopsin, Charles John Beischel

MUSC Theses and Dissertations

The objective of these studies was to further define the amino acids of bacteriorhodopsin in close proximity to the ring end of retinal by labeling them using a retinal analog modified to contain a photoactivatable group. The sites of labeling were to be identified using mass spectrometry and tandem mass spectrometry. Eight potentially photoactivatable retinal analogs were characterized and thoroughly examined as bacteriorhodopsin analog pigments thus identifying those analogs that were most similar to the native retinal. A versatile flash photolysis apparatus was compared to a continuous wave light source on the basis of total photon output and on efficiency …