Molecular Biology Commons^™

How Oxygen Availability Affects The Antimicrobial Efficacy Of Host Defense Peptides: Lessons Learned From Studying The Copper-Binding Peptides Piscidins 1 And 3, Adenrele Oludiran, David S. Courson, Malia D. Stuart, Anwar R. Radwan, John C. Putsma, Myriam L. Cotten, Erin B. Purcell

Chemistry & Biochemistry Faculty Publications

The development of new therapeutic options against Clostridioides difficile (C. difficile) infection is a critical public health concern, as the causative bacterium is highly resistant to multiple classes of antibiotics. Antimicrobial host-defense peptides (HDPs) are highly effective at simultaneously modulating the immune system function and directly killing bacteria through membrane disruption and oxidative damage. The copper-binding HDPs piscidin 1 and piscidin 3 have previously shown potent antimicrobial activity against a number of Gram-negative and Gram-positive bacterial species but have never been investigated in an anaerobic environment. Synergy between piscidins and metal ions increases bacterial killing aerobically. Here, we …

Leptin Regulates Cd16 Expression On Human Monocytes In A Sex-Specific Manner, Joseph G. Cannon, Gyanendra Sharma, Gloria Sloan, Christiana Dimitropoulou, R. Randall Baker, Andrew Mazzoli, Barbara Kraj, Anthony Mulloy, Miriam Cortez-Cooper

Medical Diagnostics & Translational Sciences Faculty Publications

Fat mass is linked mechanistically to the cardiovascular system through leptin, a 16 kDa protein produced primarily by adipocytes. In addition to increasing blood pressure via hypothalamic-sympathetic pathways, leptin stimulates monocyte migration, cytokine secretion, and other functions that contribute to atherosclerotic plaque development. These functions are also characteristics of CD16-positive monocytes that have been implicated in the clinical progression of atherosclerosis. This investigation sought to determine if leptin promoted the development of such CD16-positive monocytes. Cells from 45 healthy men and women with age ranging from 20 to 59 years were analyzed. Circulating numbers of CD14++16++ monocytes, which are primary …

Identification Of Persistent Long Range Interactions In GA95 And GB95 Through Thermal Unfolding Simulations, Milen Redai Tesfamariam

Chemistry & Biochemistry Theses & Dissertations

For over five decades, different experiments have been performed to research how proteins attain their native three dimensional structures. However, the folding problem continues to be a puzzle in modern science. The design of two proteins that have maximal sequence identity but different folds and functions is one method that is being used to study the relationship between protein structure and amino acid sequence. In particular, mutant proteins of Streptococcus protein G, G_A and G_B, have 95% sequence identity and a 3a helix fold and β4/a fold, respectively. Molecular dynamics simulations of G_A95 …

The Effects Of Cocaine And Ecstasy On Cardiac Myocytes And The Intact Myocardium, David A. Tiangco

Theses and Dissertations in Biomedical Sciences

Cocaine and ecstasy are widely used illicit drugs. Both drugs have undergone intense scrutiny as information regarding their side-effects has become available. One important yet incomplete area of investigation pertains to their effects on the heart. The purpose of the current studies was to test the hypothesis that exposure to cocaine or ecstasy will adversely affect cellular homeostasis and normal heart function. Cultured cardiac myocytes (H9c2) and New Zealand White rabbits (Oryctolagus cuniculus) were used to measure the responses to various concentrations of cocaine or ecstasy at both the cellular and intact organ system levels. We observed that cocaine and …

Positive Regulation Of Pka On Human Gonadotropin-Releasing Hormone (Hgnrh) Gene Expression In Human Placental Jeg-3 Cells, Zhaoyang Wen

Chemistry & Biochemistry Theses & Dissertations

Using the human placental choriocarcinoma JEG-3 cell line as an in vitro human placental model, we studied the mechanisms of the PKA positive regulation of the hGnRH gene expression in the human placenta. Studies in JEG3 cells showed that through the PKA catalytic subunit a, human GnRH upstream promoter activity was stimulated by PKA signaling pathway in a cAMP dependent mechanism. The sequence between —202 (Afl II) and —554 (BamH I) base pair in the human GnRH upstream promoter region appeared to be responsible for the PKA positive regulation of the gene expression. Furthermore, Western blot analysis demonstrated the involvement …

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 …

Laminin Potentiates Differentiation Of Pcc4uva Embryonal Carcinoma Into Neurons, T. M. Sweeney, Roy C. Ogle, C. D. Little

Medical Diagnostics & Translational Sciences Faculty Publications

The embryonal carcinoma PCC4uva differentiates into neurons in response to treatment with retinoic acid and dbcAMP. We used this in vitro model system to study the effects of laminin on early neural differentiation. Laminin substrata markedly potentiate neural differentiation of retinoic acid and dbcAMP-treated cultures. Only laminin induced more rapid neural cell body clustering, neurite growth and neurite fasciculation as compared to type IV collagen, type I collagen, and fibronectin substrata. Exogenous laminin substrata promoted greater cell attachment, cellular spreading and growth to confluence than type IV collagen, type I collagen, fibronectin and glass substrata. Laminin-induced effects were inhibited by …

Regulation Of Queuine Insertion Into Transfer Rna: Effects Of Tumor Promoters, Bonnie Jean Brooks

Chemistry & Biochemistry Theses & Dissertations

The purpose of this study was to treat normal human fibroblasts with known tumor promoters and observe queuine modification of tRNA. The queuine insertion enzyme, tRNAguanine ribosyltransferase, was studied in vivo and in vitro. Tumor promoter-treated human fibroblast cultures exhibited variable queuine-insertion rates with a transient inhibition that correlated with variable levels of queuine modified tRNA over time in culture from passages 3 — 8. In contrast the in vitro studies showed that phorbol cetera and saccharin actually increased insertion with strong evidence indicating phosphorylation as a positive modulating force of the enzyme activity. It is proposed that chronic stimulation …

Microwell Based Competitive Enzyme Immunoassay For Quantifying Serum Triiodothyronine And Thyroxine Levels, Yiwen Yang

Chemistry & Biochemistry Theses & Dissertations

A microwell based competitive enzyme immunoassay for determination of total thyroxine and triiodothyronine in human serum was developed with use of horseradish peroxidase conjugate. The assay, which can be performed in only 1.5 hours at room temperature, involves simultaneous incubation of T₃- or T₄- HRPO conjugate and serum sample in anti-T₃ or anti-T₄ gamma globulin fraction coated polystyrene microwells.

Isolation of anti-T₃ and anti-T₄ gamma globulin fractions were accomplished from whole antiserum by using DEAE-cellulose chromatograph. A carbodiimide coupling method was described to prepare the T₄- and T₃-HRPO …

Collagen Binding Proteins Derived From The Embryonic Fibroblast Cell Surface Recognize Arginine-Glycine-Aspartic Acid, Roy C. Ogle, Charles D. Little

Medical Diagnostics & Translational Sciences Faculty Publications

Several cell surface proteins (Mr = 120,000, 90,000, 63,000 and 47,000) apparently integral to embryonic fibroblast plasma membranes were extracted with detergent and isolated by collagen affinity chromatography. Certain of these proteins (Mr = 120,000, 90,000, and 47,000) were specifically eluted from collagen affinity columns by synthetic peptides containing the amino acid sequence arginyl-glycyl-aspartic acid (RGD). These data show that a number of collagen binding proteins exist on the embryonic fibroblast cell surface. Some of the proteins may be collagen receptors binding to RGD sequences in the collagen molecule while at least one of the proteins (Mr = 63,000) recognizes …

Hypoxanthine-Induced Differentiation Of Cultured Human Leukemia Cells, Gayle Jennette Singleton

Chemistry & Biochemistry Theses & Dissertations

Human cultured leukemia cells appear to have a decreased amount of inosine in their tRNA. When cells with inosine deficient tRNA are placed in a hypoxanthine fortified media, they incorporate hypoxanthine into their tRNA by the action of the enzyme tRNA-hypoxanthine ribosyl transferase. This generates the nucleoside inosine in the tRNA. The cultured human leukemia cell lines, CCRF-CEM, HL-60, and HGPRT(-) HL- 60, incorporate hypoxanthine into their tRNA, as determined by tRNA isolation, hydrolysis, and HPLC analysis. Hypoxanthine treatment dramatically inhibited cell growth in conjunction with partial induction of differentiation in the CCRF-CEM, HL-60, and HGPRT ( - ) HL-60 …

Isolation And Partial Characterization Of Basic, Sperm-Specific, Nuclear Proteins In The Dog And Hampster, Richard William Hall

Biological Sciences Theses & Dissertations

The purpose was to determine whether dogs and hamsters contain the same sperm-specific protamine and testis specific proteins (TP) as other eutherian mammals previously studied. The proteins (protamine and TPs) were isolated from spermatids and mature spermatozoa, and were subjected to electrophoresis in an acid-urea, polyacrylamide gel system according to the procedure of Panyim and Chalkley (1962). The results indicated that, regardless of the anatomical location of sperm acquisition, acid precipitation technique, or denaturing agents used, epididymal and testicular samples from both the dog and hamster contained protamine in elongated spermatids and mature spermatozoa. The protamines from dogs and hamsters …

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 …

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

Medical Diagnostics & Translational Sciences Faculty 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 …

High Resolution Two-Dimensional Electrophoretic Study Of Human Seminal Plasma Proteins, Edward E. Gaunt

Chemistry & Biochemistry Theses & Dissertations

A comprehensive biochemical screening procedure for the evaluation of seminal plasma constituents is needed which, when used in conjunction with semen analysis, can provide the physician with both qualitative and quantitative information regarding the fertility status of an individual.

High resolution two-dimensional electrophoresis is a technique whereby proteins in a complex mixture such as a biological fluid are separated in one dimension by molecular charge and in the second dimension by molecular mass. It has been the purpose of this investigation to adapt and evaluate this technique for the analysis of human seminal plasma.

Our efforts have shown that this …