Biochemistry Commons^™

A Synthetic Biology Approach To Engineering New Anticancer Agents, Shane Robert Wesener

Theses and Dissertations

Histone deacetylase (HDAC) inhibitors are becoming increasingly valuable therapeutic agents in treatment of several types of malignancies. FK228 is a depsipeptde anticancer compound produced by Chromobacterium violaceum no. 968 through a nonribosomal peptide synthetase (NRPS)-polyketide synthase (PKS) hybrid assembly line. In the present study, reconstitution of the biosynthetic pathway responsible for the production of FK228 revealed cross-talk between modular PKS and fatty acid synthase. This pathway contains two PKS modules on the DepBC enzymes that lack a functional acyltransferase (AT) domain, and no apparent AT-encoding gene exists within the gene cluster or its vicinity. We reported through heterologous expression of …

Iron-Regulated Cyanobacterial Predominance And Siderophore Production In Oligotrophic Freshwater Lakes, Ryan J. Sorichetti

Electronic Thesis and Dissertation Repository

The frequency and intensity of cyanobacterial blooms (cyanoblooms) is increasing globally. Contrary to existing phosphorus (P) and nitrogen (N) paradigms describing cyanobloom proliferation in eutrophic (nutrient-rich) freshwater lakes, many of the recent cyanobloom reports pertain to oligotrophic (nutrient-poor) freshwater lakes with no prior history of cyanobloom occurrence. There exists a critical research need to re-visit existing conceptual models, identify regulating factors currently unaccounted for and improve our ability to effectively detect and measure cyanobacterial toxins (cyanotoxins) in lakes. Iron (Fe) is required in nearly all pathways of cyanobacterial macronutrient use, though its direct role in regulating cyanobacterial biomass is not …

Mscs-Like Mechanosensitive Channels In Plants And Microbes, Margaret E. Wilson, Grigory Maksaev, Elizabeth S. Haswell

Biology Faculty Publications & Presentations

The challenge of osmotic stress is something all living organisms must face as a result of environmental dynamics. Over the past three decades, innovative research and cooperation across disciplines have irrefutably established that cells utilize mechanically gated ion channels to release osmolytes and prevent cell lysis during hypoosmotic stress. Early electrophysiological analysis of the inner membrane of Escherichia coli identified the presence of three distinct mechanosensitive activities. The subsequent discoveries of the genes responsible for two of these activities, the mechanosensitive channels of large (MscL) and small (MscS) conductance, led to the identification of two diverse families of mechanosensitive channels. …

A Supramolecular Strategy To Assemble Multifunctional Viral Nanoparticles, Limin Chen, Xia Zhao, Yuan Lin, Yubin Huang, Qian Wang

Faculty Publications

Using a one-pot approach driven by the supramolecular interaction between β-cyclodextrin and adamantyl moieties, multifunctional viral nanoparticles can be facilely formulated for biomedical applications.

Cellular Metabolism And Its Effect On The Type Iii Secretion System Of Dickeya Dadantii 3937, William Cortrell Hutchins

Theses and Dissertations

Nutrition, in both eukaryotes and prokaryotes, is vital to the life and well-being of the species. In organisms such as Escherichia coli, metabolism and its regulation have been well established, whereas in Dickeya dadantii 3937, the metabolic pathways and their effects on other processes have not been elucidated. Little is known is how carbon metabolism is able to regulate virulence and pathogenicity in this organism. In this work, we have investigated how the metabolic network contributes to positively and negatively regulating the pathogenicity of Dickeya dadantii 3937.

Chapter 1 provides an overview of the history and virulence processes in …

Function And X-Ray Crystal Structure Of Escherichia Coli Yfde, Elwood A. Mullins, Kelly L. Sullivan, T. Joseph Kappock

T. Joseph Kappock

Many food plants accumulate oxalate, which humans absorb but do not metabolize, leading to the formation of urinary stones. The commensal bacterium Oxalobacter formigenes consumes oxalate by converting it to oxalyl-CoA, which is decarboxylated by oxalyl-CoA decarboxylase (OXC). OXC and the class III CoA-transferase formyl-CoA:oxalate CoA-transferase (FCOCT) are widespread among bacteria, including many that have no apparent ability to degrade or to resist external oxalate. The EvgA acid response regulator activates transcription of the Escherichia coli yfdXWUVE operon encoding YfdW (FCOCT), YfdU (OXC), and YfdE, a class III CoA-transferase that is ~30% identical to YfdW. YfdW and YfdU are necessary …

Cooperative Effects Of Drug-Resistance Mutations In The Flap Region Of Hiv-1 Protease, Jennifer Foulkes-Murzycki, Christina Rosi, Nese Yilmaz, Robert Shafer, Celia Schiffer

Celia A. Schiffer

Understanding the interdependence of multiple mutations in conferring drug resistance is crucial to the development of novel and robust inhibitors. As HIV-1 protease continues to adapt and evade inhibitors while still maintaining the ability to specifically recognize and efficiently cleave its substrates, the problem of drug resistance has become more complicated. Under the selective pressure of therapy, correlated mutations accumulate throughout the enzyme to compromise inhibitor binding, but characterizing their energetic interdependency is not straightforward. A particular drug resistant variant (L10I/G48V/I54V/V82A) displays extreme entropy-enthalpy compensation relative to wild-type enzyme but a similar variant (L10I/G48V/I54A/V82A) does not. Individual mutations of sites …

Human Adenovirus E1a Binds And Retasks Cellular Hbre1, Blocking Interferon Signalling And Activating Virus Early Gene Transcription, Gregory J. Fonseca

Electronic Thesis and Dissertation Repository

Upon infection, human adenovirus (HAdV) must block interferon signaling and activate the expression of its early genes to reprogram the cellular environment to support virus replication. During the initial phase of infection, these processes are orchestrated by the first HAdV gene expressed during infection, early region 1A (E1A). E1A binds and appropriates components of the cellular transcriptional machinery to modulate cellular gene transcription and activate viral early genes transcription. We have identified hBre1/RNF20 as a novel target of E1A. hBre1 is an E3 ubiquitin ligase which acts with the Ube2b E2 conjugase and accessory factors RNF40 and WAC1 to monoubiquitinate …

A High Omega-3 Fatty Acid Diet Has Different Effects On Early And Late Stage Myeloid Progenitors, Melinda Varney, James Buchanan, Yulia Dementieva, W. Hardman, Vincent Sollars

Yulia Dementieva

The effects of the polyunsaturated omega-3 (n-3) and omega-6 (n-6) fatty acids (FA) on hematopoiesis are complex in that both FA forms are processed into leukotrienes, eicosanoids, and prostaglandins, which can have independent effects. These FA have antagonistic effects in that n-6 FA prostaglandins tend to be pro-proliferative and pro-inflammatory, while the effects of n-3 FA prostaglandins are the opposite. We have previously shown that diets high in n-3 FA reduce the size of the middle to later stage myeloid progenitor compartment in FVB X sv129 F1hybrid mice. To assay the effects of high n-3 FA diets on earlier stages …

Global Analysis Of Gene Expression Changes During Retinoic Acid-Induced Growth Arrest And Differentiation Of Melanoma: Comparison To Differentially Expressed Genes In Melanocytes Vs Melanoma, Mary H. Estler, Goran Boskovic, James Denvir, Sarah Miles, Donald A. Primerano, Richard M. Niles

Goran Boskovic

BACKGROUND: The incidence of malignant melanoma has significantly increased over the last decade. Some of these malignancies are susceptible to the growth inhibitory and pro-differentiating effects of all-trans-retinoic acid (RA). The molecular changes responsible for the biological activity of RA in melanoma are not well understood. RESULTS: In an analysis of sequential global gene expression changes during a 4-48 h RA treatment of B16 mouse melanoma cells, we found that RA increased the expression of 757 genes and decreased the expression of 737 genes. We also compared the gene expression profile (no RA treatment) between non-malignant melan-a mouse melanocytes and …

Quorum Sensing And Metabolism In Marine Environments, Amanda May

Doctoral Dissertations

Quorum sensing (QS) is a phenomenon that allows bacteria to communicate with each other. Small molecules known as autoinducers are synthesized and released by bacteria, and once enough members of the community are around to ensure survival, i.e. quorum, a phenotype, e.g. bioluminescence, is expressed. There are two types of QS molecules, intra- and inter-species.

S-4,5-Dihydroxy-2,3-pentanedione (DPD) is a byproduct of the activated methyl cycle which recycles methionine. This has led to the discussion as to whether DPD is a metabolic byproduct or is the interspecies signal as proposed previously. The detection and quantitation of DPD however, has not …

Interaction Of Bacillus Anthracis Exosporium Protein Bcla With Complement Factor H And Spore Persistence In The Lung, Sarah A. Jenkins

Dissertations & Theses (Open Access)

Anthrax outbreaks in the United States and Europe and its potential use as a bioweapon have made Bacillus anthracis an interest of study. Anthrax infections are caused by the entry of B. anthracis spores into the host via the respiratory system, the gastrointestinal tract, cuts or wounds in the skin, and injection. Among these four forms, inhalational anthrax has the highest lethality rate and persistence of spores in the lungs of animals following pulmonary exposure has been noted for decades. However, details or mechanisms of spore persistence were not known. In this study, we investigated spore persistence in a mouse …

Aerobic Degradation Of Α-, Β-, Γ-Hexachlorocyclohexane By Narragansett Bay Bacterioplankton, Ian M. Rambo

Senior Honors Projects

Hexachlorocyclohexanes (HCHs) are a family of chlorinated organic compounds that were previously used as agricultural insecticides. HCHs are recognized as persistent organic pollutants due to their toxicity, recalcitrant properties, and tendency to bioaccumulate in food webs. Although HCH was first synthesized in 1825, its use was not widespread until the discovery of the insecticidal activity of the γ-HCH isomer in 1942. γ-HCH and its toxic waste isomers α-HCH and β-HCH were banned from production and use by the United Nations in 2009, yet these chemicals still present environmental problems due to their persistence in soils and surface waters. HCHs continue …

Evolutionary And Molecular Analysis Of Conserved Vertebrate Immunity To Fungi, Erin Carter

Honors College

The innate immune system is highly conserved amongst all multicellular organisms. Yet a constant battle exists between host cells and pathogens due to the rapid evolution of immune system components. Functional genomics and in silico methods can be employed to elucidate the evolutionary patterns of vertebrate immunity to pathogenic fungi such as Candida albicans, an opportunistic fungal pathogen that can cause lethal candidiasis in the immunocompromised. Mammals such as humans and mice possess conserved C-type lectin receptors that recognize the C. albicans cell wall. However, these receptors have not been identified in fish. Here I describe how we identified potential …

Activities Of Methionine-Γ-Lyase In The Acidophilic Archaeon “Ferroplasma Acidarmanus” Strain Fer1, M. A. Khan, Madeline M. Lopez-Munoz, Charles W. Kaspar, Kai F. Hung

Kai F. Hung

Biogeochemical processes on exposed pyrite ores result in extremely high levels of sulfuric acid at these locations. Acidophiles that thrive in these conditions must overcome significant challenges, including an environment with proton concentrations at pH 3 or below. The role of sulfur metabolism in the archaeon “Ferroplasma acidarmanus” strain fer1’s ability to thrive in this environment was investigated due to its growth-dependent production of methanethiol, a volatile organic sulfur compound. Two putative sequences for methionine- γ-lyase (EC 4.4.1.11), an enzyme known to carry out α,γ-elimination on L-methionine to produce methanethiol, were identified in fer1. Bioinformatic analyses identified a conserved pyridoxal-5′-phosphate …

Activities Of Methionine-Γ-Lyase In The Acidophilic Archaeon “Ferroplasma Acidarmanus” Strain Fer1, M. Khan, Madeline Lopez-Munoz, Charles Kaspar, Kai Hung

Faculty Research & Creative Activity

Biogeochemical processes on exposed pyrite ores result in extremely high levels of sulfuric acid at these locations. Acidophiles that thrive in these conditions must overcome significant challenges, including an environment with proton concentrations at pH 3 or below. The role of sulfur metabolism in the archaeon “Ferroplasma acidarmanus” strain fer1’s ability to thrive in this environment was investigated due to its growth-dependent production of methanethiol, a volatile organic sulfur compound. Two putative sequences for methionine- γ-lyase (EC 4.4.1.11), an enzyme known to carry out α,γ-elimination on L-methionine to produce methanethiol, were identified in fer1. Bioinformatic analyses identified a conserved pyridoxal-5′-phosphate …

Activities Of Methionine-Γ-Lyase In The Acidophilic Archaeon “Ferroplasma Acidarmanus” Strain Fer1, M. A. Khan, Madeline M. Lopez-Munoz, Charles W. Kaspar, Kai F. Hung

Faculty Research & Creative Activity

Biogeochemical processes on exposed pyrite ores result in extremely high levels of sulfuric acid at these locations. Acidophiles that thrive in these conditions must overcome significant challenges, including an environment with proton concentrations at pH 3 or below. The role of sulfur metabolism in the archaeon “Ferroplasma acidarmanus” strain fer1’s ability to thrive in this environment was investigated due to its growth-dependent production of methanethiol, a volatile organic sulfur compound. Two putative sequences for methionine- γ-lyase (EC 4.4.1.11), an enzyme known to carry out α,γ-elimination on L-methionine to produce methanethiol, were identified in fer1. Bioinformatic analyses identified a conserved pyridoxal-5′-phosphate …

Novobiocin As An Allosteric Modulator Of Ste2p, Jeffrey K. Rymer, Melinda Hauser, Jeffrey M. Becker

EURēCA: Exhibition of Undergraduate Research and Creative Achievement

G protein-coupled receptors (GPCRs) are the target of 30-50% of all prescribed drugs for human medicine and are therefore the subject of intense study by the scientific community. It has been recognized recently that compounds called allosteric modulators can regulate GPCR activity by binding a GPCR at sites not occupied by the normal receptor-activating molecule. Such allosteric compounds are desirable drug candidates as they may produce fewer toxic side-effects than standard drugs that target GPCRs. The purpose of this study was to determine the interaction of different allosteric modulators with Ste2p, a model GPCR expressed in the yeast Saccharomyces cerevisiae. …

Lysine Biosynthesis In Bacteria: A Metallodesuccinylase As A Potential Antimicrobial Target, Danuta M. Gillner, Daniel P. Becker Ph.D., Richard C. Holz

Chemistry: Faculty Publications and Other Works

In this review, we summarize the recent literature on dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) enzymes, with an emphasis on structure–function studies that provide insight into the catalytic mechanism. Crystallographic data have also provided insight into residues that might be involved in substrate and hence inhibitor recognition and binding. These data have led to the design and synthesis of several new DapE inhibitors, which are described along with what is known about how inhibitors interact with the active site of DapE enzymes, including the efficacy of a moderately strong DapE inhibitor.

Phi X 174, David J. Hall

Infectious Pathogens

Phi X 174 database number # 1CD3. Phi X 174 is a virus that infects bacteria and was the first DNA-based genome to be sequenced in 1977. This bacteriophage has a [+] circular single-stranded DNA genome. The assembly of the capsid has been successfully done in vitro and is the focus of many studies on self-assembly, drug delivery and nanotechnology.

Human Papilloma Virus 16, David J. Hall

Infectious Pathogens

Human Papilloma virus 16, database# 1DZL Infection by most papillomaviruses is either asymptomatic or causes small benign tumors, known as papillomas or warts (e.g. human papillomavirus HPV6 or HPV11). Papillomas caused by some types, however, such as human papillomaviruses 16 and 18, carry a risk of becoming cancerous. Human papilloma virus is a non-enveloped, double stranded, circular DNA virus.

Dengue Virus, David J. Hall

Infectious Pathogens

Dengue virus, database # 1THD Dengue fever, is an infectious tropical disease caused by the dengue virus and transmitted by mosquito. Symptoms include fever, headache, muscle and joint pains, and a characteristic skin rash that is similar to measles. In a small proportion of cases the disease develops into the life-threatening dengue hemorrhagic fever. Dengue is a positive strand RNA virus that is part of the flavivirus family. Dengue is an enveloped virus meaning it has a nucleocapsid protecting the RNA and the nucleocapsid is surrounded by lipids (hence the name enveloped).

Hepatitis B, David J. Hall

Infectious Pathogens

Hepatitis B, database # 2g33. Hepatitis B is an infectious inflammatory illness of the liver caused by the hepatitis B virus (HBV). Hepatitis B virus (HBV) is a member of the Hepadnavirus family.The virus particle, (virion) consists of an outer lipid envelope and an icosahedral nucleocapsid core composed of protein. These virions are 42 nM in diameter. The nucleocapsid encloses the viral DNA and a DNA polymerase that has reverse transcriptase activity.

Inhibition Of Bacillus Cereus Growth By Bacteriocin Producing Bacillus Subtilis Isolated From Fermented Baobab Seeds (Maari) Is Substrate Dependent, Donatien Kaboré, Dennis S. Nielsen, Hagrétoui Sawadogo-Lingan, Bréhima Diawara, Mamoudou H. Dicko Prof., Mogens Jakobsen, Line Thorsen

Pr. Mamoudou H. DICKO, PhD

Optimizing Electrode Design For Microbial Fuel Cells Used For Wastewater Treatment, Lindsay Nichols, John A. Hogan

STAR Program Research Presentations

Microbial fuel cells (MFC) utilize bacteria to generate an electrical current that can be used in the decomposition of sludge and human urine. In a MFC there is an anode (for oxidation of organic compounds), cathode (reduction of oxygen or carbon dioxide), and a proton exchange membrane (PEM, allows protons to migrate); reduction-oxidation reactions between the anode and cathode produce a measurable current. Bacteria that are found in sludge can be used to produce electrons in a voltaic cell, but optimizing conditions for harnessing the energy is crucial to making a MFC efficient. Research has shown that the ratios of …

Expression And Characterization Of The Major Capsid Protein (Mcp) Of A Giant Marine Virus: Cafeteria Roenbergensis Virus, Sayan Chakraborty

Open Access Theses & Dissertations

The oceans play significant roles in maintaining the climate on Earth and providing nutrients to the whole biosphere. Oceanic microbes produce approximately half of the Earth's oxygen. It has been shown that the populations of microbial communities in oceans are largely regulated by viral infection and thus affect the marine ecosystem. Cafeteria roenbergensis Virus (CroV) is a giant marine virus with a linear, double stranded, and AT-rich DNA genome. The genome of CroV has a size of 744kb and encodes for 544 predicted genes. CroV derived its name from its host, a unicellular marine zooplankton Cafeteria roenbergensis (Cro) that is …

Malnutrition In Sickle Cell Anemia: Implications For Infection, Growth And Maturation, Hyacinth I. Hyacinth, Oluwatoyosi A. Adekeye, Christopher S. Yilgwan

Journal of Social, Behavioral, and Health Sciences

Sickle cell anemia (SCA) is a genetic disease that affects mostly individuals of African and/or Hispanic descent, with the majority of cases in sub-Saharan Africa. Individuals with this disease show slowed growth, delayed sexual maturity, and poor immunologic function. These complications could partly be explained by the state of undernutrition associated with the disease. Proposed mechanism of undernutrition include protein hypermetabolism, decreased dietary intake possibly from interleukin-6-related appetite suppression, increased cardiac energy demand/expenditure, and increased red cell turnover. All the above mechanisms manifest as increased resting energy expenditure. Nutritional intervention utilizing single or multiple nutrient supplementation has led to improved …

Bacteriostatic Effects Of Sucralose On Environmental Bacteria, Arthur Phillip Omran Jr.

UNF Graduate Theses and Dissertations

Sucralose is a zero calorie sweetener developed and manufactured by Tate and Lyle Sweetener Company in the 1980’s. They sell the sweetener compounded with maltodextrin and dextrose under the brand name Splenda®. Sucralose was developed as a low cost artificial sweetener that is non-metabolizable in humans and can withstand changes in pH and temperature. It is not degraded by the waste water treatment process. Since the molecule can withstand heat, acidification and microbial degradation it is accumulating in the environment, and has been found in waste water, estuaries, rivers and the Gulf Stream. The highest concentration of environmental sucralose detected …

Critical Events In Human Metapneumovirus Infection: From Entry To Egress, Brent A. Hackett

Theses and Dissertations--Molecular and Cellular Biochemistry

Human metapneumovirus (HMPV) is a respiratory pathogen in Paramyxovirus family that demonstrates extremely high morbidity in the population, with most individuals having been infected by the age of five. Despite the prevalence of this negative-sense RNA virus in the population for decades, it was only identified in 2001. As such, there is currently no specific treatment for HMPV and the potentially severe consequences of infection for elderly and immunocompromised individuals and particularly infants make development of antivirals targeting HMPV of high significance. HMPV constitutes a quarter of all respiratory hospitalizations among infants, placing it second only to RSV, in addition …

Exploitation Of The Bacteriophage-Derived Peptidase Chapk, Pierre-Mehdi Hadbi

Theses

This project investigated an anti-staphylococcal bacteriophage-derived peptidoglycan hydrolase enzyme, namely a cysteine-histidine amydohydrolase/peptidase (CHAP_k). The study focused firstly on optimizing the production of recombinant CHAP_k, which was previously cloned in an E. coli expression system, and sets out the resulting optimal Standard Operating Procedures (SOPs) for production, purification and determination of concentration and activity of the resulting protein stock. A typical yield resulting from the investigation was 300pg of CHAP_k from one litre of E.coli culture. The activity of the purified enzyme was typically in the region of 190U nmol ’. Secondly, the project investigated …