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Concurrent Identification, Characterization, And Reconstruction Of Protein Structure And Mixed-Mode Dynamics From Rdc Data Using Redcraft, Hanin Rafiq Omar

Theses and Dissertations

A complete understanding of the structure-function relationship of proteins requires an analysis of their dynamic behaviors and the static structure. However, all current approaches to studying dynamics in proteins have their shortcomings. A conceptually attractive and alternative approach simultaneously characterizes a protein's structure and its intrinsic dynamics⁠. Ideally, such an approach could solely rely on RDC data-carrying both structural and dynamical information. The major bottleneck in utilizing RDC data in recent years has been attributed to a lack of RDC analysis tools capable of extracting the pertinent information embedded within this complex data source.

Here we present a comprehensive strategy …

Macronutrient Intake: A Multi-Sport Study Of Female Division I Collegiate Athletes, Ffion Price

Theses and Dissertations

Background: Macronutrients play a critical role within collegiate athletes’ performance and health, with carbohydrates providing most of the energy needs for most athletes. There is little research examining the macronutrient intake of healthy collegiate female athletes across sports. The aim of the present study was to compare macronutrient intakes of female collegiate athletes within different sports and compare their intakes to recommendations. Methods: An observational study was conducted to determine whether a sample of female collegiate athletes (n=26) consumed the IOC nutritional recommendations. Sports included within the study were soccer, basketball, volleyball, and cross-country. Athletes were asked to complete a …

Molecular Dynamic Simulation Of Protein Devices And The Parameterization Of Azides And Alkynes For Use In Unnatural Amino Acid Models, Addison Kyle Smith

Theses and Dissertations

Proteins that have been modified by attaching them to a surface or to a polyethylene glycol (PEG) molecule can see many uses in therapeutics and diagnostics -- these unique proteins are called protein devices. Current techniques can perform these functionalizations at a specific residue on the protein, but what remains is identifying what happens to protein structure when mutated, and where to perform the attachment. Both of these issues can be examined using molecular dynamic (MD) simulations. Currently, simulations of the unnatural amino acid (uAA) mutations necessary for protein device functionalization cannot be executed, and full-protein screens of all possible …

Molecular Dynamic Simulation Of Protein Devices And The Parameterization Of Azides And Alkynes For Use In Unnatural Amino Acid Models, Addison Kyle Smith

Theses and Dissertations

Proteins that have been modified by attaching them to a surface or to a polyethylene glycol (PEG) molecule can see many uses in therapeutics and diagnostics -- these unique proteins are called protein devices. Current techniques can perform these functionalizations at a specific residue on the protein, but what remains is identifying what happens to protein structure when mutated, and where to perform the attachment. Both of these issues can be examined using molecular dynamic (MD) simulations. Currently, simulations of the unnatural amino acid (uAA) mutations necessary for protein device functionalization cannot be executed, and full-protein screens of all possible …

Biophysical Characterization Of Protein-Nanoparticle Interactions, Yasiru Randika Perera

Theses and Dissertations

Nanoparticles (NPs) have become a key tool in medicine and biotechnology; as drug delivery systems, biosensors, and diagnostic devices. However, the mechanism of biocorona formation on nanoparticle surfaces and their impact on drug delivery remains speculative. Nevertheless, functionalized nanoparticles have demonstrated major success in medical applications; having been shown to effectively treat disease. The mechanistic details of protein behavior on nanoparticle surfaces remain poorly understood to date; due to difficulty in determining the orientation and structure of protein on NPs. Furthermore, surface crowding, orientation, and degree of disorder have been shown to perturb the efficacy of protein on NPs; dramatically …

Social Media Posts' Influence On College Athletes' Protein Knowledge, Kriston T. Love

Theses and Dissertations

This research aimed to examine how Instagram can be used as a means of education for collegiate athletes and their knowledge of protein. More specifically this research aimed to compare the prior knowledge about protein that collegiate athletes have to the knowledge they may have retained after being exposed to posts on Instagram about protein. It also aimed to determine the knowledge of protein collegiate athletes already had and if their sport and year in school influenced their protein knowledge. This study surveyed Division I collegiate athletes on their social media use and knowledge of protein before and after being …

The Immediate And Persistent Effects Of Binge Ethanol Exposure On Myelin Protein Expression In Dba/2j Mice, Aminatta Z. Tejan-Kamara

Theses and Dissertations

Binge drinking is the most lethal of drinking patterns, responsible for 77% of all U.S. alcohol misuse costs. Adolescents have increased sensitivity to ethanol’s rewarding properties and decreased sensitivity to ethanol’s aversive effects likely allowing for their increased binge drinking, as compared to adults. Ethanol consumption during adolescent neurodevelopment can lead to immediate and lasting neurobiological-consequences. We have previously shown that adolescent intermittent binge ethanol exposure decreases myelin-related gene expression in DBA/2J mice. Here, we hypothesize intermittent binge ethanol exposure in adolescent DBA/2J mice will decrease myelin protein expression in the prefrontal cortex (PFC) immediately following binge ethanol and this …

Developing A Bio-Based Wood Composite Using Refined Cottonseed Protein Adhesives., Julianna Nicole Stratton

Theses and Dissertations

A growing market of environmentally-conscious consumers combined with a progression toward ‘greener’ products has caused the wood industry to investigate adhesives containing little to no formaldehyde. This study examines cottonseed proteins’ ability to bind southern yellow pine for plywood applications. Three-ply plywood panels were constructed with varying blends of cottonseed protein isolate (CSPI), soy protein, and phenol: formaldehyde adhesive. Wet and dry shear testing revealed that while the novel adhesives did not perform as well as a commercial control, the CSPI and soy adhesives generated similar shear strengths. Another set of boards were created, varying the CSPI amount added and …

Culture Techniques And Evaluation Of Snail Consumption By Black Buffalo Ictiobus Niger, Emmet Liam Guy

Theses and Dissertations

Black Buffalo Ictiobus niger are a catostomid species native to the Mississippi River Basin with declining populations. Aquaculture is a common component of recovery plans, however little information is published on catostomid culture methods. This study developed culture techniques for Black Buffalo including tank acclimation, induced spawning, larval rearing, and protein requirements. Fish needed a lengthy acclimation (> 2 months) to reduce signs of stress. After, an induced-spawning protocol was described with gonadotropin-releasing hormone analogue. Swim-up fry did not readily consume zooplankton at first feeding but began selecting for cladocerans one week after stocked in a rearing pond. The juvenile …

Comparison Of Functional, Nutritional, And Sensory Properties Of Spray-Dried And Oven-Dried Cricket (Acheta Domesticus) Powder, Fred Stephen Bassett

Theses and Dissertations

Three dried cricket products – Oven-roasted Cricket Meal (ORCM), Oven-roasted Cricket Powder (ORCP), and Spray-dried Cricket Powder (SDCP) – were prepared and compared to assess their relative nutritional, functional, and sensory properties. The range of particle sizes in SDCP was wider than that of ORCP; while SDCP had more particles in the 7.78-22.0 µm range, it also had more particles in the 176-498 µm range, which contributed to differences in functional and sensory properties. ORCM and SDCP meet the FDA definition of an "excellent source" of vitamin B₂, ORCP is a "good source" of vitamin B₂, …

A Biorelevant In Vitro Model To Characterize In Vivo Release Of Bone Morphogenetic Protein-2 (Rhbmp-2), Deblina Biswas

Theses and Dissertations

Biorelevant in vitro release/dissolution tests are designed to predict the in vivo behavior of a drug and are crucial in understanding its in vivo performance. Currently, there is no standardized compendial in vitro release testing methods or regulatory guidance’s for release/dissolution testing of implants due to their complex physiological locations.Furthermore, existing compendial methods do not capture the local release profile of ‘novel’ parenterals in physiological low fluid volume surrounding areas.

Long acting and in situ forming implants with orthobiologic proteins and peptides have increased over the past few decades due to a better understanding of genetic engineering. One of these …

Muscle Inflammatory Signaling Regulates Eccentric Contraction-Induced Protein Synthesis During Cancer Cachexia, Justin Perry Hardee

Theses and Dissertations

Cancer-related cachexia is a complex metabolic syndrome that involves the unintentional loss of skeletal muscle mass that cannot be reversed by standard nutritional intervention. Skeletal muscle mass depletion directly impacts patient survival and life quality, and therefore treatments that preserve muscle mass and function may have significant implications for cancer patient treatment and survival. Systemic and muscle inflammation has been implicated in the regulation of skeletal muscle homeostasis, and the cytokine interleukin-6 (IL-6) and muscle gp130 receptor signaling have established roles in cancer-induced muscle wasting. Resistance exercise is a nonpharmacological treatment that can improve physical function and metabolic health in …

Strength Training And Body Composition In Middle-Age Women, Rachelle Burrup

Theses and Dissertations

OBJECTIVE: The purpose of this study was to examine the relationship between strength training and body composition before and after controlling for several covariates. A cross-sectional study including 257 female subjects was conducted. METHODS: Subjects' level of involvement in strength training was determined via questionnaire. Body composition was assessed using dual energy X-ray absorptiometry (DXA). Diet was assessed using 7-d weighed food records. RESULTS: Strong linear relationships between subjects' level of involvement in strength training and body composition were identified. For each additional day of strength training reported per week, body fat was 1.32 percentage points lower (F = 14.8, …

Protein Therapy For The Treatment Of Vascular Calcification In Patients With End-Stage Renal Disease, Janice L. Cunningham

Theses and Dissertations

Premature death from cardiovascular disease is especially high in patients with chronic kidney disease (CKD). Vascular calcification is a potent risk factor for developing cardiac-related morbidity and mortality and is especially prominent within the CKD population. Deficiencies in serum levels of fetuin-A as well as inadequate production of matrix Gla protein (MGP) correlate inversely with the extent of vascular calcification and time spent on dialysis. Fetuin-A is a well-known systematic regulator of bone metabolism and MGP is a local antagonist of bone forming proteins. To meet the clinical need of at-risk patients prone to cardiac-related mortality. We propose a targeted …

Cloning, Expression And Preliminary Characterization Of A Novel Nitrilase From The Red Sea, Atlantis Ii Deep Brine Pool Using A Metagenomic Approach, Sarah Ali Sonbol

Theses and Dissertations

With the abundance of hazardous nitrile compounds in nature and with the extensive use of them in industries of fine chemicals, the attention towards nitrilases has profoundly increased as biocatalysts in different industries and in bioremediation. The use of nitrilases, which hydrolyze nitriles into carboxylic acids and ammonia, was introduced as a biological method superior to conventional chemical means. In contrast to conventional microbiological techniques using cultured microorganisms, and with the loads of genomic sequences available in the databases, identification of novel nitrilases using metagenomic approaches seems more promising. In this study, a nitrilase was isolated from the microbial metagenomic …

Probing Protein And Organothiol Interactions With Gold Nanoparticles, Karthikeshwar Vangala

Theses and Dissertations

Proteins and organothiols are known for their high binding affinity to noble metal surface including gold nanoparticles (AuNPs). Numerous reports have been dedicated to AuNP interaction with protein or organothiol alone. Competitive protein and organothiol (OT) interaction is, however, mostly an unexplored area. The research reported here focused on developing a fundamental understanding of sequential and simultaneous protein and organothiol interaction with AuNPs in which protein and OT are added either simultaneously or sequentially into the colloidal AuNP solutions. In studies of OT interactions with bovine serum albumin (BSA) stabilized AuNPs, we found that the protein coating layer is highly …

Signaling By Protease-Activated Receptors In Gastrointestinal Smooth Muscle, Wimolpak Sriwai

Theses and Dissertations

In the present study, we have examined the expression of protease-activated receptors (PARS) and characterized their signaling pathways in rabbit gastric muscle cells. Immunoblot analysis revealed expression of PARl and PAR2 but not PAR3 or PAR4 in smooth muscle. The PARl agonist TFLLR activated Gq, G12, and Gi3, but not Gil, Gi2, G13, Gs or Gz, whereas the PAR2 agonist SLIGRL activated Gq, G13, Gil, and Gi2, but not Gi3, G12, Gs, or Gz. Both PARl and PAR2 agonists stimulated PI hydrolysis and Rho kinase activity and inhibited cAMP formation. PAR1-stimulated PI hydrolysis was abolished in cells expressing Gαq minigene, …

Biochemical Investigations Of Macular Degeneration: The Significance Of Protein Oxidation Including Novel Methods For Its Study, Sarah Warburton

Theses and Dissertations

The retinal pigment epithelium (RPE) is a monolayer of cells located directly behind the photoreceptor cells in the retina. These cells are involved in a variety of functions that support the visual process in the eye, namely 1) they form a blood-retina barrier which separates the neural retina from the choroid's blood supply, 2) the apical processes of RPE cells diurnally phagocytose the outer segments of photoreceptor cells, and 3) they participate in the renewal of the photopigment 11-cis retinal. Age-related macular degneration (AMD) is the leading cause of blindness in people over the age of 50 years in North …

Fabrication Of Polymeric Microfluidic Devices For Protein Analysis, Jikun Liu

Theses and Dissertations

2-Bromoisobutyryl bromide was immobilized on poly(methyl methacrylate) (PMMA) substrates activated using an oxygen plasma. Atom-transfer radical polymerization was then performed to graft poly(ethylene glycol) (PEG) on the PMMA surface. PMMA micro capillary electrophoresis (µCE) devices made with the covalently modified surfaces exhibited substantially reduced electroosmotic flow and nonspecific adsorption of proteins. Both column efficiency and migration time reproducibility were one order of magnitude better with derivatized PMMA µCE devices compared to untreated versions. Fast, reproducible, and efficient separations of proteins and peptides were demonstrated using the PEG-grafted PMMA µCE chips. All analyses were completed in less than 60 seconds, and …

Membrane-Based Protein Preconcentration Microfluidic Devices, Yi Li

Theses and Dissertations

Interest in microchip capillary electrophoresis (CE) is growing due to the rapid analysis times provided and small sample input requirements. However, higher-concentration samples are typically needed because of the small (~pL) detection volumes in these devices. I have made membrane-based protein preconcentration systems in capillary and microchip designs to increase the detectability of low-concentration biological samples. A photopolymerized ion-permeable membrane interfaced with a microchannel in poly(methyl methacrylate) (PMMA) formed the preconcentrator. When a voltage was applied between the sample reservoir and the ionically conductive membrane in a capillary-based system, R-phycoerythrin was concentrated more than 1,000 fold, as determined by laser-induced …

Identification Of Novel Allosteric Regulators Of Human Erythrocyte Pyruvate Kinase, Shilpa S. Kharalkar

Theses and Dissertations

Erythrocyte pyruvate kinase (R-PK) is a key glycolytic enzyme catalyzing the transphosphorylation of phosphoenolpyruvate (PEP) and ADP to pyruvate and ATP respectively3,4. The substrate PEP and product pyruvate of this reaction are involved in a number of energetic and biosynthetic pathways; hence a tight regulation of R-PK activity is crucial not only for glycolysis, but also for the entire cellular metabolism. Deficiency of R-PK is one of the most common enzymatic defects of RBC, and may be caused by mutations of the PK-LR (pyruvate kinase liver red blood cell) gene31, 32. Clinically, R-PK deficiency manifests itself as a chronic life-long …

Small Molecules Binding To Serpins, Junaid Afridi

Theses and Dissertations

Serpins are a unique breed of proteins due to their enzymatic mechanism. Two systems were closely monitored during fluorescent binding studies, the ACT-CHY along with the AT:TRY interaction. Four different conformational variants of each system were studied including the native, cleaved, latent and complex forms. Three different fluorescent dyes were used to identify the conformations including ANS, TNS, and bis-ANS. SI studies and protease assays utilizing both Suc-AAPF-pNA and L-BAPNA were instrumental in determining conformations along with gel electrophoresis studies. The hydrophobic dyes bound to the different serpins with varying KD and ΔFmax due to structural variations among the conformers …

Characterization Of The Tcof1 Gene Using A Neuroblastoma Cell Line And A Mouse Model, Lin Li

Theses and Dissertations

Treacher Collins syndrome (TCS) is an autosomal dominant craniofacial development disorder and is caused by mutations in the TCOF1 gene. The TCOFl protein treacle is a nucleolar protein and may function in ribosome biogenesis.Previously, we identified downstream candidate genes using microarray analysis after manipulating Tcofl levels in a murine neuroblastoma (NB) cell line. The list of genes includes cell cycle genes as well as the transcription factors Cnbp and Tbx2, which are known to affect the cell cycle through the c-myc and p19-Mdm2-p53-p21 pathways respectively. To further characterize the cellular effects of Tcofl, stably transfected NB cell lines with overexpression …

Dual Regulation Of Telomerase Activity By Hsf1 And Its Role In Prostate Cancer Progression, Keith Douglas Ostergaard Jensen

Theses and Dissertations

It has been shown that the key components of the hsp90 chaperone complex, including hsp90, p23, hsp70, hsp40, and HOP (p60), associate with telomerase; however, their specific roles in telomerase function and tumor progression have not yet been defined. HSF1, the primary mammalian heat shock protein transcription factor, may affect telomerase activity and transformation by regulating the expression of several hsp90 chaperone complex proteins in response to stress as well as regulating the transcription of hTERT, the protein subunit of telomerase.In our in vitro model of prostate cancer progression, as cells progress from immortal but non-tumorigenic (P69) to tumorigenic (M2182) …

Identification Of The Pla2 Responsible For Prostanoid Synthesis In Response To Inflammatory Cytokines, Chaminda Fernando

Theses and Dissertations

Preliminary studies from our laboratory showed that cPLA2α may be responsible for approximately 50-60% of the PGE2 production in response to inflammatory cytokines. Thus, we hypothesized that a closely-related PLA2 is responsible for 40-50% of the PGE2 produced in response to inflammatory cytokines. To this end, we utilized RNAi technology, extensively optimized, to down regulate the expression of closely-related isoforms of phospholipase A2 in A549 cells and used an enzyme linked immuno sorbent assay (ELISA) to quantitate the PGE2 produced. These studies found that cytosolic phospholipase A2α (cPLA2α) regulated 97.7% of the prostaglandin E2 (PGE2) produced in response to inflammatory …

The Effect Of Traumatic Brain Injury On Expression Levels Of Ankyrin-G In The Corpus Callosum And Cerebral Cortex, Andrew S. Vanderveer

Theses and Dissertations

The ankyrins comprise a family of proteins serving as components of the membrane cytoskeleton, and participate in a diverse set of associations with multiple binding partners including the cytoplasmic domains of transporters, ion channels, some classes of receptors, and cell adhesion proteins. Moreover, evidence is accumulating that ankyrin participates in defining functionally distinct subcellular regions. The complex functional and structural roles of ankyrins indicate they are likely to play essential roles in the pathology of traumatic axonal injury. The current study examined changes in ankyrin-G expression following a moderate central fluid percussion injury administered to adult rats. At 1d, 3d, …

Atomic Force Microscope Conductivity Measurements Of Single Ferritin Molecules, Degao Xu

Theses and Dissertations

Conductive Atomic Force Microscope (c-AFM) was used to measure the conductivity of single horse spleen ferritin (HoSF) and azotobacter vinelandii bacterial ferritin (AvBF) molecules deposited on flat gold surfaces. A 500 micron diameter gold ball was also used as a contact probe to measure the conductivity of a thin film of ferritin molecules. The average current measured for holo HoSF was 13 and 5 times larger than that measured for apo HoSF as measured by c-AFM at 1V and gold ball at 2V and respectively, which indicates that the core of ferritin is more conductive than the protein shell and …

Identification Of Proteins Involved In Chloroplast Dna Replication, Matthew G. Lassen

Theses and Dissertations

Chapter 1

Chloroplast nucleoids (ct-nucleoids) are DNA/protein complexes involved in compacting the chloroplast genome, and may play a role in regulating DNA replication. Ct-nucleoids were isolated from young soybean plants and separated by 2-D gel electrophoresis. Gel spots were excised and analyzed by MALDI-ToF mass spectrometry, resulting in several protein identifications. The proteins identified all have functions unrelated to DNA replication. While some of these proteins may be due to contamination, it is possible that some of these proteins are dual-functional, playing direct roles in the regulation of DNA replication.

Chapter 2

A 28 kDa soybean protein was isolated by …