Biochemistry Commons^™

Progress Toward The Synthesis Of Aminomethyl-Substituted Qpt-1 Bacterial Topoisomerase Inhibitors, Riley Scoville, Michael Barbachyn

Summer Research

Bacterial resistance to antibiotics is a growing public health threat. Each year, ~2 million infections and 23,000 deaths are caused by antibiotic resistance in the US.1 Developing antibiotics for treating Gram-negative bacteria is especially challenging because they utilize two membranes, an outer membrane that is hydrophilic, and an inner membrane that is lipophilic, features that make cellular entry a challenge. These bacteria also have antibiotic de-activating enzymes and RND efflux pumps that work to remove the antibiotics from the cell. These features make targeting of Gram-negative organisms very difficult. Agents targeting the essential bacterial type II topoisomerases DNA gyrase and …

Investigation Of The Saccharomyces Cerevisiae Gpi Transamidase: Insights Into Its Activity And Subunit-Subunit Interactions, Travis Ness

Wayne State University Dissertations

Glycosylphosphatidylinositol (GPI) anchoring of proteins is a eukaryotic, posttranslational

modification catalyzed by GPI transamidase (GPI-T). The Saccharomyces

cerevisiae GPI-T is composed of five membrane-bound subunits: Gaa1, Gpi8, Gpi16,

Gpi17, and Gab1. Structural and functional studies have been hindered by the

complexity of this enzyme. Conditions to purify the Gpi8:Gaa1:Gpi16 GPI-T heterotrimer

from yeast have been reported, but an understanding of the subunit functions,

interactions, and stoichiometry remain unclear. Furthermore, a reliable, quantitative, in

vitro assay for this important post-translational modification has remained elusive for

nearly three decades.

Our laboratory has developed an in vitro peptide cleavage assay that correlates

changes …

Biochemical And Cellular Studies Of Apobec3 Family Dna-Cytosine Deaminases, Sachini Umedi Siriwardena

Wayne State University Dissertations

The AID/APOBEC family of enzymes deaminate cytosines in single-stranded DNA to uracils leading to base substitutions and strand breaks. Members of APOBEC3 family in humans are induced by cytokines produced during the body's inflammatory response to infections and provide innate immunity against viruses. However, there is emerging consensus that these enzymes can cause mutations in the cellular genome depending on the physiological state of the cell and the phase of the cell cycle they are expressed. Since aberrant expression of APOBEC3B was recently identified as a possible source of cancer, we initiated a study to determine the maximally active catalytic …

Design, Synthesis, And Reactivity Of Homo- And Heterobimetallic Complexes Bridged By A Xanthene Linker, Thilini Samangi Hollingsworth

Wayne State University Dissertations

Cooperative reactivity of bimettalics can be is observed in many different areas of chemistry and have been increasingly investigated because of the advantageous reactivity when compared to the corresponding mononuclear systems. The focus of my dissertation is on (1) investigation of the homobimetallic cooperativity in lactide polymerization catalysis; (2) investigation of the heterobimetallic cooperativity in the biomimetic studies of Mo-Cu carbon monoxide dehydrogenase (CODH) enzyme in order to make a functional model of its active site.

Three new main group bis(alkoxide) complexes Mg(OR)2(THF)2, Zn(Cl)(μ2-OR)2Li(THF) and In(OR)2(μ2-Cl)2Li(THF)2 featuring bulky alkoxide [OCtBu2Ph] were synthesized serve as metal alkoxide precursors for bimetallic lactide …

Effects Of Lipid Activating Chemical Compounds On The Growth And Production Of Fatty Acids And Metabolites In Green Algae, David Nguyen, Nishikant Wase, Concetta Dirusso

UCARE: Research Products

This study examines the effects of small molecule lipid activating chemical compound, discovered from the high throughput screening (HTS) (2) methods of previous FATTT Lab studies on the growth and production of lipids and metabolites in microalgae. For this study, the effects of two lipid inducing chemical compounds from the HTS method were implemented with the microalgae strain Chlamydomonas reinhardtii CC-125. This experiment focused on scaling up to large amounts of culture, approximately 1L. These cultures were grown in specially designed large bioreactors that were able to accommodate for such large volume of algal culture. Algal cells were treated with …

Characterization Of Metabolic Network In T-Helper 2 Cells, Bailee Lichter

UCARE: Research Products

T-helper 2 (Th2) cells function in the adaptive immune system to coordinate responses to large extracellular pathogens (Luckheerham, 2012). Th2 cells are one of the differentiated subtypes cells of CD4+ T cells. CD4+ T cells recognize foreign pathogens and differenitated in specialized subtypes to affectively destroy the pathogen (Luckheerham, 2012). Upon CD4+ T cell activation, the cells undergo metabolic reprogaming, resulting in a shift in metabolism (Almedia, 2016).

We have successfully created a Th2 metabolic network model. The Th2 metabolic network model contains 1151 genes, 2601 metabolites, and 4654 reactions.

Investigation Of The In Vivo Activity Of Ribosome-Targeting Peptides And Aminoglycosides In Escherichia Coli, Nisansala Sarangi Thilakarathne Muthunayake

Wayne State University Dissertations

The development of short peptides that specifically bind to higher-order structures of ribosomal RNA is one promising way to address the problem of antibiotic resistance. However, the poor correlation between in vitro and in vivo activities of these peptides is one of the major questions in antibiotic peptide research. Therefore, one of the main objectives of this dissertation work was to utilize a plasmid-based system to in vivo express ribosome-targeting peptides and study their direct inhibitory effects on bacteria. A specific plasmid system was optimized to in vivo express oncocin, a prolin-rich antimicrobial peptide and its variants in bacteria. Our …

The Effects Of Site-Directed Mutagenesis On Hemerythrin-Like Protein Rv2633c, Kelly M. Rosch

Honors Undergraduate Theses

Mycobacterium tuberculosis (Mtb) is the causative agent of tuberculosis, one of the top ten causes of death worldwide. One of the genes upregulated in Mtb during macrophage infection is rv2633c, but the structure and function of its gene product remain unknown. Preliminary research has indicated that Rv2633c is a hemerythrin-like protein that exhibits catalase activity and binds two iron atoms using an HHE domain. Additionally, Rv2633c appears to exist as a dimer. The purpose of this project is to identify specific residues outside of the HHE domain that contribute to the protein's iron-binding ability and/or catalase activity, …

Investigation Of Aminoglycoside Induced Nanoparticle Self-Assemblies, Michael Leong

Honors Undergraduate Theses

Aminoglycosides are a group of broad-spectrum antibiotics that, under neutral pH conditions, carry a positive charge. The net cationic charge arises from the high number of amino groups in the core structure of aminoglycosides. Previous studies performed have shown that negatively charged citrate ligand-capped gold nanoparticles (AuNPs) can interact with various biomolecules such as aminoglycosides. AuNPs bound to biomolecules have been used in conjugation with various assaying techniques to detect and study compounds in vitro and in vivo. AuNPs also have strong light scattering properties that can be used with a wide variety of imaging and assaying techniques. Our laboratory …

Functional Study Of Smyd2 Glutathionylation In Cardiomyocytes, Dhanushka Nalin Perera Munkanatta Godage

Wayne State University Dissertations

Reactive oxygen species (ROS) are important signaling molecules that contribute to the etiology of multiple muscle-related diseases, including cardiomyopathy and heart failure. There is emerging evidence that cellular stress can lead to destabilization of sarcomeres, the contractile unit of muscle. However, it is not completely understood how cellular stress or ROS induce structural destabilization of sarcomeres or myofibrils. Protein glutathionylation is one of the major protein cysteine oxidative modifications that play an important role in redox signaling and oxidative stress. In this report, we used a clickable glutathione approach in a cardiomyocyte cell line, and found that SET and MYND …

Biosynthetic Mechanism Of The Antibiotic Capuramycin, Erfu Yan

Theses and Dissertations--Pharmacy

A-102395 is a member of the capuramycin family of antibiotics which was isolated from the culture broth of Amycolatopsis sp. SANK 60206. A-102339 is structurally classified as a nucleoside antibiotic, which like all members of the capuramycin family, inhibits bacterial MraY (translocase I) with IC₅₀ of 11 nM which is the lowest among the capuramycin family. A semisynthetic derivative of capuramycin is currently in clinical trials as an antituberculosis antibiotic, suggesting high potential for using A-102395 as a starting point for new antibiotic discovery. In contrast to other capuramycins, A-102395 has a unique arylamine-containing polyamide side chain. The biosynthetic …

Quantifying The Spectrum Of Depression, Octavious Bishop

Walden Dissertations and Doctoral Studies

Walden University

College of Social and Behavioral Sciences

This is to certify that the doctoral dissertation by

Octavious Bishop

has been found to be complete and satisfactory in all respects,

and that any and all revisions required by

the review committee have been made.

Review Committee

Dr. Thomas Edman, Committee Chairperson, Psychology Faculty

Dr. Ricardo Thomas, Committee Member, Psychology Faculty

Dr. Joanna Bissell-Havran, University Reviewer, Psychology Faculty

Chief Academic Officer

Eric Riedel, Ph.D.

Quantifying the Spectrum of Depression

by

Octavious Bishop

MA, University of Texas at Austin, 2008

BS, University of Texas at Austin, 2001

Dissertation Submitted in Partial Fulfillment …

Role Of Glycerol-3-Phosphate Permeases In Plant Defense, Juliana Moreira Soares

Theses and Dissertations--Plant Pathology

Systemic acquired resistance (SAR) is a type of plant defense mechanism that is induced after a localized infection and confers broad-spectrum immunity against related or unrelated pathogens. During SAR, a number of chemical signals and proteins generated at the site of primary infection travel to the uninfected tissues and are thought to alert the distal sites against secondary infections. Glycerol-3-phosphate (G3P) is one of the chemical signals that play an important role in SAR. G3P is synthesized in the cytosol and chloroplasts via the enzymatic activities of G3P Dehydrogenase (G3Pdh) or Glycerol Kinase (GK). Interestingly, a mutation in three of …

Carbohydrate-Based Inducers Of Cellular Stress For Targeting Cancer Cell Metabolism, Fidelis Ndombera

Wayne State University Dissertations

ABSTRACT

CARBOHYDRATE-BASED INDUCERS OF CELLULAR STRESS FOR TARGETING CANCER CELL METABOLISM

by

FIDELIS TOLOYI NDOMBERA

May 2018

Advisor: Dr. Young-Hoon Ahn

Major: Chemistry (Biochemistry)

Degree: Doctor of Philosophy

Metabolic reprogramming and redox control of cancer cells is vital for their proliferation, but also provides selective strategies for treating cancer. Increased generation of reactive oxygen species (ROS) and an intricate control of redox status in cancer cells relative to normal cells provide a basis for designing ROS-inducing anticancer agents. In my work, I designed, synthesized and evaluated carbohydrate-based small molecules for ROS-generation, cytotoxicity and redox signaling and stress response. Our data …

Asymmetric Patterns Of Small Molecule Transport After Nanosecond And Microsecond Electropermeabilization, Esin B. Sözer, C. Florencia Pocetti, P. Thomas Vernier

Bioelectrics Publications

Imaging of fluorescent small molecule transport into electropermeabilized cells reveals polarized patterns of entry, which must reflect in some way the mechanisms of the migration of these molecules across the compromised membrane barrier. In some reports, transport occurs primarily across the areas of the membrane nearest the positive electrode (anode), but in others cathode-facing entry dominates. Here we compare YO-PRO-1, propidium, and calcein uptake into U-937 cells after nanosecond (6 ns) and microsecond (220 µs) electric pulse exposures. Each of the three dyes exhibits a different pattern. Calcein shows no preference for anode- or cathode-facing entry that is detectable with …

Short-Chained Oligo(Ethylene Oxide)-Functionalized Gold Nanoparticles: Realization Of Significant Protein Resistance, Kathryn R. Riley, C. M. Sims, I. T. Wood, D. J. Vanderah, M. L. Walker

Chemistry & Biochemistry Faculty Works

Protein corona formed on nanomaterial surfaces play an important role in the bioavailability and cellular uptake of nanomaterials. Modification of surfaces with oligoethylene glycols (OEG) are a common way to improve the resistivity of nanomaterials to protein adsorption. Short-chain ethylene oxide (EO) oligomers have been shown to improve the protein resistance of planar Au surfaces. We describe the application of these EO oligomers for improved protein resistance of 30 nm spherical gold nanoparticles (AuNPs). Functionalized AuNPs were characterized using UV-Vis spectroscopy, dynamic light scattering (DLS), and zeta potential measurements. Capillary electrophoresis (CE) was used for separation and quantitation of AuNPs …

Functional Characterization Of Scaffold Protein Shoc2, Hyein Jang

Theses and Dissertations--Molecular and Cellular Biochemistry

Signaling scaffolds are critical for the correct spatial organization of enzymes within the ERK1/2 signaling pathway and proper transmission of intracellular information. However, mechanisms that control molecular dynamics within scaffolding complexes, as well as biological activities regulated by the specific assemblies, remain unclear.

The scaffold protein Shoc2 is critical for transmission of the ERK1/2 pathway signals. Shoc2 accelerates ERK1/2 signaling by integrating Ras and RAF-1 enzymes into a multi-protein complex. Germ-line mutations in shoc2 cause Noonan-like RASopathy, a disorder with a wide spectrum of developmental deficiencies. However, the physiological role of Shoc2, the nature of ERK1/2 signals transduced through this …

Detection Of Local Steroidogenic Enzyme Gene Expression In Brown Anoles, Ada Spahija

Honors Program Theses

The endocrine system in vertebrates responds to stress by releasing steroid hormones, mainly glucocorticoids (GC), which increase blood glucose levels to supply key organs and muscles with energy needed for survival. Steroid hormones are synthesized via an enzymatic pathway that converts cholesterol into either GCs, androgens, or estrogens in a step-wise manner. The adrenal cortex is known to produce GCs, but evidence suggests that individual organs can also produce steroid hormones de novo in response to stress. This study aims to quantify gene expression of four steroidogenic enzymes, encoded by CYP19A1, CYP17A1, StAR, and HSD17ß3 genes, via qRT-PCR to determine …

The Contributions Of Fc Gamma Receptors And Macropinocytosis To The Internalization, Sorting And Clearance Of Antibody Coated Nanovesicles In Macrophages, George Opoku-Kusi Jr.

Electronic Theses and Dissertations

Macrophages are tissue-resident phagocytes that play critical roles in immune response and tissue homeostasis. They have a tremendous capacity to internalize objects of various sizes ranging from nanoscale viral particles to micron sized bacteria and tumor cells. This phenomenon, termed phagocytosis for the uptake of large particles, or endocytosis for the uptake of small particles, is integral to the immune response in multicellular organisms. Macrophages express FcγRs, which are tyrosine kinase receptors that bind IgG on an opsonized target. Binding of IgG Fc domain to the extracellular domain of an FcγR triggers signaling cascades that coordinate internalization of the opsonized …

Study Of Anti-Proliferative Activity Of Cucurbitacins Inspired Estrone Analogs On Hepatocellular Carcinoma, Sara Elgazwi

Electronic Theses and Dissertations

Hepatocellular carcinoma (HCC) is considered the third leading cause of death from cancer. Overall survival rate is significantly low, due to the emerging resistance to chemotherapeutic agents and lack of selectivity. Recent studies have demonstrated that epidermal growth factor receptor (EGFR) is a promising molecular target for cancer therapy, especially HCC. Current studies showed that cucurbitacins are potent anticancer compounds which target EGFR. This prompted us to investigate the antiproliferative activity of novel cucurbitacins inspired estrone analogs (CIEA) against sensitive and resistant HepG2 cell lines. Anti-proliferation activity of 20 CIEA analogs were examined against HepG2 using MTT assay and showed …

A Comprehensive Analysis Of Aromatic-Proton Mediated Hydrogen Bonds, Mona S. Alshamrani

Electronic Theses and Dissertations

Hydrogen bonds play critical role in folding, structure and recognition of biological macromolecules (e.g., proteins, RNA, DNA). In addition to classical hydrogen bonds (e.g., ─OH---O=, ─OH---O─, ─NH---O─ etc.), structural analysis of protein and nucleic acids, almost a decade ago, showed that hydrogen bonds (e.g., CH---O) with hydrogen atoms on aliphatic carbon atoms (hereafter, aliphatic-protons) also play very important role in the structure and function of biomolecules. Even though, protons of aromatic ring systems (hereafter, aromatic-protons) are more polar than the aliphatic-protons, systematic analysis of hydrogen bonds of aromatic-protons have not been carried out. Therefore, I carried out a systematic analysis …

Interleukin-6 And Exercise; Early Evidence Of A Novel Myokine, Brendan Hogg Mr.

Graduate Student Theses, Dissertations, & Professional Papers

Interleukin-6 (IL-6) was first identified as a pleiotropic cytokine, with a host of body-wide functions. Cytokine functions are characterized by chronically elevated levels within various inflammatory states. In this regard, IL-6 is largely associated with the acute phase response to many stimuli and performs specific actions when produced from certain cell types. Accrued evidence indicates IL-6 release from skeletal muscle often includes myokine functions. Novel myokine functions are adaptive in nature, and as compared to inflammatory/cytokine roles, exhibit a transient time course. Following exercise plasma IL-6 peaks and returns to resting levels within 1-2 hours. In contrast, IL-6 is observed …

Dielectric Properties Of Isolated Adrenal Chromaffin Cells Determined By Microfluidic Impedance Spectroscopy, A. C. Sabuncu, M. Stacey, G. L. Graviso, N. Semenova, P. T. Vernier, N. Leblanc, I. Chatterjee, J. Zaklit

Bioelectrics Publications

Knowledge of the dielectric properties of biological cells plays an important role in numerical models aimed at understanding how high intensity ultrashort nanosecond electric pulses affect the plasma membrane and the membranes of intracellular organelles. To this end, using electrical impedance spectroscopy, the dielectric properties of isolated, neuroendocrine adrenal chromaffin cells were obtained. Measured impedance data of the cell suspension, acquired between 1 kHz and 20 MHz, were fit into a combination of constant phase element and Cole-Cole models from which the effect of electrode polarization was extracted. The dielectric spectrum of each cell suspension was fit into a Maxwell-Wagner …

Photosystem Ii Subunit S Overexpression Increases The Efficiency Of Water Use In A Field-Grown Crop, Katarzyna Glowacka, Johannes Kromdijk, Katherine Kucera, Jiayang Xie, Amanda P. Cavanagh, Lauriebeth Leonelli, Andrew D. B. Leakey, Donald R. Ort, Krishna K. Niyogi, Stephen P. Long

Department of Biochemistry: Faculty Publications

Insufficient water availability for crop production is a mounting barrier to achieving the 70% increase in food production that will be needed by 2050. One solution is to develop crops that require less water per unit mass of production. Water vapor transpires from leaves through stomata, which also facilitate the influx of CO2 during photosynthetic assimilation. Here, we hypothesize that Photosystem II Subunit S (PsbS) expression affects a chloroplastderived signal for stomatal opening in response to light, which can be used to improve wateruse efficiency. Transgenic tobacco plants with a range of PsbS expression, from undetectable to 3.7 …

Simulation Of Stimulation: Cytokine Dosage And Cell Cycle Crosstalk Driving Timing-Dependent T Cell Differentiation, Matteo Barberis, Tomáš Helikar, Paul Verbruggen

Department of Biochemistry: Faculty Publications

Triggering an appropriate protective response against invading agents is crucial to the effectiveness of human innate and adaptive immunity. Pathogen recognition and elimination requires integration of a myriad of signals from many different immune cells. For example, T cell functioning is not qualitatively, but quantitatively determined by cellular and humoral signals. Tipping the balance of signals, such that one of these is favored or gains advantage on another one, may impact the plasticity of T cells. This may lead to switching their phenotypes and, ultimately, modulating the balance between proliferating and memory T cells to sustain an appropriate immune response. …

Computational Systems Biology Approach For The Study Of Rheumatoid Arthritis: From A Molecular Map To A Dynamical Model, Vidisha Singh, Marek Ostaszewski, George D. Kalliolias, Gilles Chiocchia, Robert Olaso, Elisabeth Petit-Teixeira, Tomáš Helikar, Anna Niarakis

Department of Biochemistry: Faculty Publications

In this work we present a systematic effort to summarize current biological pathway knowledge concerning Rheumatoid Arthritis (RA). We are constructing a detailed molecular map based on exhaustive literature scanning, strict curation criteria, re-evaluation of previously published attempts and most importantly experts’ advice. The RA map will be web-published in the coming months in the form of an interactive map, using the MINERVA platform, allowing for easy access, navigation and search of all molecular pathways implicated in RA, serving thus, as an on line knowledgebase for the disease. Moreover the map could be used as a template for Omics data …

The Colomoto Interactive Notebook: Accessible And Reproducible Computational Analyses For Qualitative Biological Networks, Aurelien Naldi, Celine Hernandez, Nicolas Levy, Gautier Stoll, Pedro T. Monteiro, Claudine Chaouiya, Tomáš Helikar, Andrei Zinovyev, Laurence Calzone, Sarah Cohen-Boulakia, Denis Thieffry, Loic Pauleve

Department of Biochemistry: Faculty Publications

Analysing models of biological networks typically relies on workflows in which different software tools with sensitive parameters are chained together,many times with additional manual steps. The accessibility and reproducibility of such workflows is challenging, as publications often overlook analysis details, and because some of these tools may be difficult to install, and/or have a steep learning curve. The CoLoMoTo Interactive Notebook provides a unified environment to edit, execute, share, and reproduce analyses of qualitative models of biological networks. This framework combines the power of different technologies to ensure repeatability and to reduce users’ learning curve of these technologies. The framework …

A Mechanistic Computational Model Reveals That Plasticity Of Cd4+ T Cell Differentiation Is A Function Of Cytokine Composition And Dosage, Bhanwar Lal Puniya, Robert G. Todd, Akram Mohammed, Deborah M. Brown, Matteo Barberis, Tomáš Helikar

Department of Biochemistry: Faculty Publications

CD4+ T cells provide cell-mediated immunity in response to various antigens. During an immune response, naïve CD4+ T cells differentiate into specialized effector T helper (Th1, Th2, and Th17) cells and induced regulatory (iTreg) cells based on a cytokine milieu. In recent studies, complex phenotypes resembling more than one classical T cell lineage have been experimentally observed. Herein, we sought to characterize the capacity of T cell differentiation in response to the complex extracellular environment. We constructed a comprehensive mechanistic (logical) computational model of the signal transduction that regulates T cell differentiation. The model’s dynamics were characterized and analyzed under …

Reactive Oxygen Species In Chronic Obstructive Pulmonary Disease, Samia Boukhenouna, Mark A. Wilson, Karim Bahmed, Beata Kosmider

Department of Biochemistry: Faculty Publications

Chronic obstructive pulmonary disease (COPD) includes chronic bronchitis and emphysema. Environmental exposure, primarily cigarette smoking, can cause high oxidative stress and is the main factor of COPD development. Cigarette smoke also contributes to the imbalance of oxidant/antioxidant due to exogenous reactive oxygen species (ROS). Moreover, endogenously released ROS during the inflammatory process and mitochondrial dysfunction may contribute to this disease progression. ROS and reactive nitrogen species (RNS) can oxidize different biomolecules such as DNA, proteins, and lipids leading to epithelial cell injury and death. Various detoxifying enzymes and antioxidant defense systems can be involved in ROS removal. In this review, …

Investigating Phosphorylation Patterns And Their Effect On The Activity Of Transcription Factor Taabf1 In Imbibing Cereal Grains, Grace Uwase

Honors Theses

The wheat transcription factor TaABF1 plays an important role in hormone-mediated regulation of seed dormancy and germination of cereal grains. Evidence shows that TaABF1 activity is regulated by phosphorylation, and previous work in our lab showed that when serine residues in its conserved regions; S36, S37, S113, S115 were altered to phosphomimetic aspartate, the 4xD TaABF1 mutant had increased activity as a transcription factor. However, when only S113 was altered, TaABF1’s activity was greatly reduced. The work presented here explored whether the S36D/S37D/S115D mutant would have stronger activity than the 4xD mutant. Using the particle bombardment technique to introduce the …