Life Sciences Commons^™

Characterization Of The Reconstituted And Native Pseudomonas Aeruginosa Type Iii Secretion System Translocon, Kathryn R. Monopoli

Masters Theses

The Type III Secretion (T3S) system is a system utilized by many pathogenic bacteria to inject proteins into host cells during an infection. Effector proteins enter the host cell by passing through the proteinaceous T3S translocon, which forms a pore on the host cell membrane. Pseudomonas aeruginosa is an opportunistic pathogen that utilizes the T3S system, and very little is known about how the P. aeruginosa translocon forms.

The proteins PopB and PopD are believed to assemble into the P. aeruginosa translocon. A pore-forming heterocomplex of PopB and PopD has been reconstituted in model membranes, however this heterocomplex has not …

Promoting Extracellular Matrix Crosslinking In Synthetic Hydrogels, Marcos M. Manganare

Masters Theses

The extracellular matrix (ECM) provides mechanical and biochemical support to tissues and cells. It is crucial for cell attachment, differentiation, and migration, as well as for ailment-associated processes such as angiogenesis, metastases and cancer development. An approach to study these phenomena is through emulation of the ECM by synthetic gels constructed of natural polymers, such as collagen and fibronectin, or simple but tunable materials such as poly(ethylene glycol) (PEG) crosslinked with short peptide sequences susceptible to digestion by metalloproteases and cell-binding domains. Our lab uses PEG gels to study cell behavior in three dimensions (3D). Although this system fosters cell …

Rnai Validation Of Resistance Genes And Their Interactions In The Highly Ddt-Resistant 91-R Strain Of Drosophila Melanogaster, Kyle Gellatly

Masters Theses

4,4’-dichlorodiphenyltrichloroethane (DDT) has been re-recommended by the World Health Organization for malaria mosquito control. Previous DDT use has resulted in resistance, and with continued use resistance will increase in terms of level and extent. Drosophila melanogaster is a model dipteran that has many available genetic tools, numerous studies done on insecticide resistance mechanisms, and is related to malaria mosquitoes allowing for extrapolation. The 91-R strain of D. melanogaster is highly resistant to DDT (>1500-fold), however, there is no mechanistic scheme that accounts for this level of resistance. Recently, reduced penetration, increased detoxification, and direct excretion have been identified as …

The Evolution Of Thermotolerance A Characterization Of A Directionally Evolved Cyanobacterium, Nathen Emil Bopp

Masters Theses

Chaperone proteins are essential components in the maintenance and turnover of the proteome. Many chaperones play integral functions in the folding and unfolding of cellular substrates under many conditions, including heat stress. Most chaperones can be characterized into two categories; the typical ATP dependent chaperones and the ATP independent chaperones. One ATP independent chaperone class it the Small Heat Shock Proteins (sHSPs), which as molecular life vests and are thought to protect misfolding proteins from irreversible aggregation. One such organism, the cyanobacterium Synechocystis sp. PCC 6803, is an excellent model for the study and understanding of these proteins and their …

Utilizing In Silico And/Or Native Esi Approaches To Provide New Insights On Haptoglobin/Globin And Haptoglobin/Receptor Interactions, Ololade Fatunmbi

Doctoral Dissertations

Haptoglobin (Hp), an acute phase protein, binds free hemoglobin (Hb) dimers in one of the strongest non-covalent interactions known in biology. This interaction protects Hb from causing potentially severe oxidative damage and limiting nitric oxide bioavailability. Once Hb/Hp complexes are formed, they proceed to bind CD163, a cell surface receptor on macrophages leading to complex internalization and catabolism. Myoglobin, (Mb) a monomeric protein, that is normally found in the muscle but can be released into the blood in high concentrations during myocardial injury, is homologous to Hb and shares many conserved Hb/Hp interface residues. Both monomeric Hb and Mb species …

Biophysical Characterization Of Katanin’S Regulation Of Microtubules, Megan E. Bailey

Doctoral Dissertations

Microtubules, as an essential part of the cytoskeleton, require proper function as well as correct spatial and temporal localization. In order to achieve correct organization, microtubule-associated proteins (MAPs) regulate microtubule dynamics. Katanin, a known microtubule-severing enzyme from the AAA family of proteins, plays a role in regulating microtubules, but the mechanisms of microtubule control and the mechanism of severing activity remain to be elucidated. In the following studies I examine mechanisms of katanin-based regulation of microtubule dynamics using a single molecule biophysics approach. I use this simplified in vitro approach to change specific parameters to investigate how katanin targets microtubules …

New Insights Into The Role Of The Udp-Glucose: Glycoprotein Glucosyltransferase 1 In The Endoplasmic Reticulum Quality Control, Abla Tannous

Doctoral Dissertations

The UDP-glucose:glycoprotein glucosyltransferase 1 (UGT1) is a central quality control factor in the Endoplasmic Reticulum (ER). It surveys the folding status of proteins in the ER and redirects them, via its reglucosylation activity, to bind to the ER carbohydrate binding (lectin) chaperones calreticulin (CRT) and calnexin (CNX). However, the cellular mechanism of UGT1 is not completely understood. Using a cell based reglucosylation assay, we found that UGT1 reglucosylated proteins that eventually fold. This modification was transient and resulted in delay of protein trafficking in the secretory pathway and prolonged binding to lectin chaperones in the ER. In addition, terminally misfolded …

Responsive Supramolecular Assemblies Based On Amphiphilic Polymers And Hybrid Materials, Longyu Li

Doctoral Dissertations

The design and synthesis of responsive supramolecular assemblies are of great interest due to their applications in a variety of areas such as drug delivery and sensing. We have developed a facile method to prepare self-crosslinking disulfide-based nanogels derived from an amphiphilic random copolymer containing a hydrophilic oligo-(ethylene glycol)-based side-chain functionality and a hydrophobic pyridyl disulfide functional group. This thesis first provides a concept of studying the influence of Hofmeister ions on the size and guest encapsulation stability of a polymeric nanogel. The size and core density of nanogel can be fine-tuned through the addition of both chaotropes and kosmotropes …

Chemical Biology-Based Probes For The Labeling Of Targets On Live Cells, Amanda M. Hussey

Doctoral Dissertations

Proper detection is the key to studying any processes on the cellular scale. Nowhere is this more evident than in the tight space which confines the synaptic cleft. Being able to ascertain the location of receptors on live neurons is fundamental to our understanding of not only how these receptors interact and move inside the cell but also how neurons function. Most detection methods rely on significantly altering the receptor; both tagging with a fluorescent protein or targeting the receptor by a fluorescent reporter in the form of a small molecule causes significant difficulties. These localization techniques often result in …

Molecular Mechanisms Underlying The Contralateral Repeated Bout Effect (Crbe) In Human Skeletal Muscle, Ling Xin

Doctoral Dissertations

Eccentric (muscle lengthening) exercise induces temporary muscle damage that can lead to long-term muscle adaptation, a process known as the repeated bout effect where subsequent exercise results in less damage. The existence of a contralateral repeated bout effect (CRBE) has been controversial. The primary goals of this study were to: 1) validate the existence of the CRBE; and 2) define the underlying molecular mechanisms. Thirty-six young men performed 100 maximal eccentric actions of the knee extensors using one leg (bout 1) and repeated the exercise with the contralateral leg five weeks later (bout 2). Vastus lateralis muscle biopsies were …

Design And Synthesis Of Polymeric Nanoparticles For Drug And Protein Delivery, Judy A. Ventura

Doctoral Dissertations

Nanoparticles are emerging as carriers in biological applications due to advances in their preparation, size control, surface modification and encapsulation capabilities. In addition, nanomaterials improve bioavailability by enhancing aqueous solubility of the guest molecule and increasing resistance time in the body. However, the delivery of guest molecules is still challenging due to the intrinsic characteristics of the guest molecule including large size and propensity to denature or degradation in the case of biomolecules and the encapsulation stability of the small guest molecules. Our group recently reported the preparation of self-cross-linked polymeric nanogels possessing surface functionalization capabilities. In this dissertation we …

Inhibition And Cofactor Targeting Of Hypoxia-Sensing Proteins, Cornelius Y. Taabazuing

Doctoral Dissertations

Hypoxia Inducible Factor (HIF) is a transcription activator considered to be the main regulator of O₂ homeostasis in humans. The transcriptional ability of HIF is regulated by the Fe²⁺/αKG-dependent enzyme, Factor Inhibiting HIF (FIH). FIH uses molecular oxygen to catalyze hydroxylation of an asparagine residue (Asn⁸⁰³) in the C-terminal transactivation domain (CTAD) of the HIFα subunit, abrogating HIF target gene expression. The mechanism of FIH and other αKG-dependent oxygenases involves the ordered sequential binding of αKG, substrate, and O₂, which becomes activated to form a reactive ferryl intermediate that hydroxylates the substrate. The …

Rnai Mediated Silencing Of Cell Wall Invertase Inhibitors To Increase Sucrose Allocation To Sink Tissues In Transgenic Camelina Sativa Engineered With A Carbon Concentrating Mechanism, Joshua Zuber

Masters Theses

Plant invertases are a class of proteins that have enzymatic function in cleaving sucrose to fructose and glucose. Cell wall invertase, located on the exterior of the cell wall of plant cells, plays a key role in the unloading of sucrose from the apoplast to the sink tissues. Cell wall invertase interacts with an inhibitor, cell wall invertase inhibitor, post-transcriptionally to regulate its activity. The inhibitor is constitutively expressed in pollen development, early developing seeds, and senescing leaves: indicative of sucrose allocation being a limiting factor at these stages of development. We introduced algal bicarbonate transporters LCIA/CCP1 to Camelina sativa …

Characterization Of Protein-Protein Interactions For Therapeutic Drug Design Utilizing Mass Spectrometry, Alex J. Johnson

Masters Theses

The number of transferrin based therapeutics progressing to clinical trials remains disappointingly small despite promising capabilities of transporting therapeutic payloads to cancer cells and across the blood brain barrier. This meager success record is largely due to the complexity and heterogeneity of all protein conjugation products that generates difficulties for their analytical characterization. Discussed in this work, transferrin is conjugated to lysozyme as a model therapeutic to deliver this bacteriostatic protein to target central nervous system infections. In this work ESI- and MALDI-MS were used to characterize the modification sites at lysine residues in hopes of characterizing heterogeneity within the …

Thermocycle-Regulated Wall Regulator Interacting Bhlh Encodes A Protein That Interacts With Secondary-Cell-Wall-Associated Transcription Factors, Ian P. Whitney

Masters Theses

Lignocellulosic biomass is one of the most abundant raw materials on earth that can be utilized to created carbon-neutral biofuels as a replacement for conventional fossil fuels. In order to create ideal energy crops, the regulation and deposition of cell wall polysaccharides must first be fully understood. Improved understanding of cell wall regulation will enable selection of traits that can optimize biofuel feedstocks. Herein, I utilize the grass model system Brachypodium distachyon in order to understand the transcriptional regulation of secondary cell wall deposition. Gene expression profiling was used to elucidate transcription factors that regulate secondary cell wall biosynthesis. Through …

Gelatin Diffusion Experiment, Jennifer Welborn

Nanotechnology Teacher Summer Institutes

In this activity, nanotech participants will:

- See how food dyes and gelatin are used to model the delivery of nanoscale medicines to cells in the human body - Measure diffusion distances of 3 different colors of food dye by: Eye, photo image on a computer, ADI software (Analyzing Digital Images) Some useful websites:

• http://www.echalk.co.uk/Science/biologyContent.htm Life Processes and Cells
• http://workbench.concord.org/database/activities/321.html Molecular Workbench Database. Diffusion, osmosis, ...