Biochemistry Commons^™

Synthesis, Structural Characterization, And In Vitro Biological Assessment Of Trans- Platinum (Ii) Thionate Complexes As Potent Anticancer Agents, Mia Alshami

Chemistry & Biochemistry Undergraduate Honors Theses

The content of this thesis has been originally reported in our published paper, “trans-Platinum (II) Thionate Complexes: Synthesis, Structural Characterization, and in vitro Biological Assessment as Potent Anticancer Agents” ChemPlusChem 2019 84, 1525-1535, DOI: 10.1002/cplu.201900394, in which I served as coauthor. Cancer caused 9.6 million deaths in 2018 worldwide, with 18.1 million new diagnoses during that same year.The most widely used metal in anticancer drugs is platinum (Pt), and these drugs are used to treat almost 50% of cancer patients. To optimize drug effectiveness, trans-configured Pt(II) complexes have been introduced as a strategy to potentially overcome the …

Hyper Stable Variants Of Fgf-1-Fgf-2 Dimer, Madison Shields Mcclanahan

Chemistry & Biochemistry Undergraduate Honors Theses

Fibroblast Growth Factors (FGFs), including FGF-1 and FGF-2, are proteins that play a crucial role in cell proliferation, cell differentiation, cell migration, and tissue repair. FGF-1 and FGF-2 are useful in accelerating the healing process in the human body; however, these proteins are naturally thermally unstable, resulting in a relatively low half-life in vivo. 1,8 In efforts to improve the stability of this protein, FGF-1 and FGF-2 proteins are engineered by combining the amino acid sequences of the two proteins to form a heterodimer and obtain novel properties. These two FGF variants are chosen for their specific wound healing capabilities. …

Effects Of Localized Oxygen Production By Electrolysis On The First-Generation Glucose Sensor Response, Nandita Halder

Graduate Theses and Dissertations

Glucose sensors are very important for detecting blood glucose both in vitro and in vivo. First-generation glucose biosensors were based on the glucose oxidase (GOx) enzyme using molecular oxygen as the electron acceptor and therefore oxygen dependent. Unfortunately for in-vivo work, oxygen in the body is variable and limited. Alternative approaches to overcome the oxygen dependency came with their own limitations. The widely used and commercially available ex-vivo glucose test strip uses a mediator in place of oxygen to free it from oxygen dependency. The mediator-based technology, in most cases cannot be transferred to in vivo applications due to the …

Biochemical Characterization Of Small Molecule Inhibitor Binding On A Ras Related Gtpase And Its Effector Interactions, Djamali Muhoza

Graduate Theses and Dissertations

The Ras superfamily of GTPases has 167 proteins that are involved in various cellular processes such as proliferation, transformation, migration, and inhibition of cell death. Mutations, abnormal expression, and function of these proteins are observed in many diseases, including several forms of cancer. Even though these GTPases were among the first discovered oncogenes, no successful Ras drug candidate has successfully passed clinical trials. Drugs targeting these proteins have failed mainly because of the complexity of their regulation, their high affinity to GTP, and their structure’s dynamic nature. Recently, novel promising targeting approaches have renewed interest in the Ras drug discovery …

Design Of Hfgf1 Variant(S) With Increased Stability And Enhanced Bioactivity, Shilpi P. Agrawal

Graduate Theses and Dissertations

Fibroblast growth factors (FGFs) are involved in various cellular processes such as cell growth,proliferation, differentiation, migration, angiogenesis, wound healing and embryonic development. Human acidic fibroblast growth factor (hFGF1) binds non-selectively to all the four FGF-receptors and is therefore considered as a powerful mitogen with broadest specificity. However, pharmacological applications of hFGF1 are restricted due to the low thermal stability of the growth factor. hFGF1 has low thermodynamic stability under physiological temperatures which leads to impairment of cellular signaling process thereby preventing its potential mitogenic properties. hFGF1 has a heparin binding pocket at the C-terminus which comprises of positively charges residues. …

Investigation Of The Role Of Heparin-Binding Pocket In Amyloid Fibrils Formation Of Fgf-1, I Gusti Ayu Agung Septiari

Graduate Theses and Dissertations

Human acidic fibroblast growth factor (aFGF/hFGF-1) is one of the promising molecules to be investigated to generate an in-depth understanding of the pathological mechanism of Alzheimer's disease (AD) neurodegenerative disorder characterized by the presence of amyloid fibrils. Some in vivo and human brain tissue studies proved the correlation of high-level expression of FGF-1-induced neuroinflammation and the occurrence of AD. The presence of amyloid fibrils as a hallmark of AD can be related to the generic property of the proteins to form amyloid fibrils; High level of FGF-1, in this case, may contribute to the formation of amyloid fibrils. As a …

Probing Of Carbohydrate-Protein Interactions Using Galactonoamidine Inhibitors, Jessica B. Pickens

Graduate Theses and Dissertations

Glycoside hydrolases are ubiquitous and one of the most catalytically proficient enzymes known, and thus understanding their mechanisms are crucial. Most research has focused on the interaction of the glycon of substrates and their inhibitors within the active site of glycoside hydrolases. The inhibitors employed to probe these interactions generally had small aglycons (i.e. a hydrogen atom, amidines, small aliphatic groups, or benzyl groups). Here, the interactions of the aglycon with glycoside hydrolases are examined by probing the active sites with a library of 25 galactonoamidines. The studies described in this dissertation aim to increase the understanding of stabilization of …

Toward Understanding The Mechanism Of Protein Targeting In The Chloroplast Signal Recognition Particle Pathway, Mercede Furr

Graduate Theses and Dissertations

Protein targeting is a vital cellular function. The signal recognition particle (SRP) pathway is a universally conserved targeting system present in the cytosol and used to co-translationally target many proteins to the inner membrane of prokaryotes and the endoplasmic reticulum of eukaryotes. The chloroplast has a homologous SRP system which post-translationally targets light harvesting chlorophyll binding proteins (LHCPs) to the thylakoid membrane for integration. The chloroplast SRP (cpSRP) is a heterodimer with a 54 kDa subunit equivalent to SRP54 in the canonical pathway. In addition, cpSRP contains a novel 43 kDa subunit which is a unique and irreplaceable component. cpSRP43 …

Production And Purification Of Basic Fibroblast Growth Factor Fused To Two Collagen Binding Domains Expressed In E. Coli Bl21 Using Flask And Fed-Batch, Hazim Aljewari

Graduate Theses and Dissertations

Delivering effective and non-toxic doses of bioactive materials that can aid in activating tissue regeneration to wounded tissue has proven to be an enormous challenge. This study was designed to produce a potential therapeutic recombinant protein by fusing two collagen binding domains to basic fibroblast growth factors (bFGF) through a collagenase cleavage site linker, so it can release the bFGF in a wound site by the action of this enzyme. The novel fusion protein was expressed in Escherichia coli BL-21 (E. coli) using traditional flask shaker and fed-batch cultivation. Cell lysate was purified by FPLC using Immobilized metal affinity chromatography …

Influence Of Single And Multiple Histidine Residues And Their Ionization Properties On Transmembrane Helix Dynamics, Orientations And Fraying, Fahmida Afrose

Graduate Theses and Dissertations

Since aromatic and charged residues are often present in various locations of transmembrane helices of integral membrane proteins, their impacts on the molecular properties of transmembrane proteins and their interactions with lipids are of particular interest in many studies. In this work, I used solid-state deuterium NMR spectroscopy in designed model peptide GWALP23 [GGALW(LA)6LWLAGA] with selective deuterium labels to addresses the pH dependence and influence of single and multiple “guest” histidine residues in the orientation and dynamic behaviors of transmembrane proteins. The mutations include Gly to His (G2/22 to H2/22), Trp to His (W5/19 to H5/19) and Leu to His …

Single Molecule Fluorescence Studies Of Protein Structure And Dynamics Underlying The Chloroplast Signal Recognition Particle Targeting Pathway, Dustin R. Baucom

Graduate Theses and Dissertations

The work presented in this dissertation explores the structural dynamics in the chloroplast signal recognition particle pathway. Findings include cpSRP shows scanning functionality similar to that in the cytosolic SRP with the ribosome. The intrinsically disordered C-terminal tail of the Albino3 protein has some transient secondary structure. Upon binding to cpSRP43 in solution, separate secondary structure formation was identified in the C-terminal tail of Albino3. Finally, to increase efficiency of analyzing fluorescence time traces for this work, a modular software was produced.

Targeting Sec61Α By Ipomoeassin F Leads To Highly Cytotoxic Effect, Zhijian Hu

Graduate Theses and Dissertations

Ipomoeassin F is a flagship congener of a resin glycoside family that inhibits growth of many tumor cell lines with only single-digital nanomolar IC50 values. However, biological and pharmacological mechanisms of ipomoeassin F have been undefined. To facilitate exploration of the biological and pharmacological properties, we performed sophisticate SAR (Structure–activity relationship) studies of ipomoeassin F to understand its pharmacophore and structure properties so that we can design favorable probes for further biological investigation. By applying appropriate deviates that possess fluorescent groups and similar bio-activity, the target protein was found to be localized in endoplasmic reticulum (ER). Through biotin affinity pull …

The Effects Of Eicosapentaenoic Acid (Epa) And Docosahexaenoic Acid (Dha) On Brown Adipogenesis In Stem Cell Culture, Darynne Dahlem

Animal Science Undergraduate Honors Theses

Polyunsaturated fatty acids (PUFAs), especially eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), are major maternal dietary supplements due to their positive benefits on neurological tissue growth during the first 12 weeks of gestation. Previous studies show that EPA and DHA inhibit muscle formation but promote adipogenesis. However, no research has addressed the question whether high intake of EPA and DHA affects brown fat development during gestation. The objective of this study was to measure the effect of EPA and DHA supplement on brown adipogenesis and potential pathways related to mitochondrial biosynthesis using fibroblasts as in vitro model. Using Oil-Red-O staining …

The First In Vivo Human Methionine Sulfide Proteome And The Impact Of Smoking, Abdullah Qassab

Graduate Theses and Dissertations

Reactive oxygen species are naturally generated within the human body and they are known to modulate signaling pathway and mediate other physiological activities. However, excessive generation of ROS and the inability of body defense system in detoxifying them results in the so called “oxidative stress”. Methionine has powerful antioxidant properties due to the presence of electronegative sulfur in its structure. Therefore, Met is readily oxidized, and methionine sulfoxide has been linked to several pathological conditions.

The urinary proteome is an attractive candidate for the discovery of biomarkers to diagnose and classify health conditions because of the non-invasive collection procedure. However, …

Engineering A Mutation In The Heparin Binding Pocket Of The Human Fibroblast Growth Factor, Roshni Patel

Chemistry & Biochemistry Undergraduate Honors Theses

Fibroblast growth factors (FGFs) are family of proteins that belong to a group of growth factors that are found in mammals and play an important role in angiogenesis, differentiation, organogenesis, and tissue repair. In summary, their main functionality is involved in cell division and proliferation. Because FGFs plays such a vital role in cell proliferation, they are mainly involved in the process of wound healing and injuries. FGF binds to its ligand, heparin—a heavily sulfated glycosaminoglycan. The binding of heparin to FGF occurs through electrostatic interactions, specifically between the negatively charged sulfate groups on heparin and positively charged residues such …

Deciphering The Role Of Glycine134 In The Human Acidic Growth Factor-1’S Binding To Heparin, Adam W. Burroughs

Chemistry & Biochemistry Undergraduate Honors Theses

Human acidic fibroblast growth factor 1 (FGF-1) is a potent modulator of cell survival and exhibits a universal role in various physiological processes. Though potent, FGF-1 unbound to heparin is known to show a poor thermal stability and a relatively short in vivo half-life. Much is known about the structure and relation of FGF-1 with heparin yet there is still unknown information regarding the exact role of heparin in stabilizing FGF-1. Thus, the aim of this study is to mutate glycine at position 134 to glutamic acid in wild type FGF1. G134 is located in the heparin binding pocket, thus …

Local Modulation And Measurement Of Macrophage-Derived Bioactive Proteins From Implanted Biomaterials In Rat, Geetika Bajpai

Graduate Theses and Dissertations

Fibrosis around the implanted medical devices is a severe problem that can plague long-term device reliability. Activation of macrophage phenotype (macrophage polarization) has emerged as a new and possible means for reducing fibrosis in the fields of biomaterials and regenerative medicine. Macrophages are phagocytic cells that respond to microenvironmental cues that direct their phenotype. Macrophage activation has been widely studied in mouse and human in the context of tumor biology, yet little information is available regarding how macrophage activation could be used in a biomaterials context. Further, rats rather than mice are the common subjects in biomaterials experiments. A significant …

Design, Development, And Characterization Of Novel Antimicrobial Peptides For Pharmaceutical Applications, Yazan H. Akkam

Graduate Theses and Dissertations

Candida species are the fourth leading cause of nosocomial infection. The increased incidence of drug-resistant Candida species has emphasized the need for new antifungal drugs. Histatin 5 is a naturally occurring human salivary antifungal peptide and the first line of defense against infections of the oral cavity. This research has focused on understanding the activity of histatin 5, and subsequently designing novel peptides that may serve as models for the further development of therapeutics to treat fungal infection.

This objective has been achieved in three steps: studying the structural requirement of histatin 5 involved in antifungal activity, the identification of …

Oxidation Of Thrombomodulin Methionine 388 In Cigarette Smokers, Samrat B. Thapa

Graduate Theses and Dissertations

This work tested the hypothesis that oxidation of methionine 388 in thrombomodulin is higher in cigarette smokers, and thus a likely contributor towards the hypercoagulable state in smokers. Thrombomodulin, a protein cofactor found on endothelial cell surfaces, regulates the activity of thrombin. Thrombin bound to thrombomodulin no longer converts fibrinogen to fibrin, but instead activates Protein C which, in turn, stops the coagulation cascade by inactivation of clotting factors. The oxidation of methionine 388 of thrombomodulin has been shown in vitro to dramatically decrease the anticoagulant cofactor activity of thrombomodulin. The blood of cigarette smokers is more prone to clot …