Biochemistry Commons^™

Biophysical Factors Affecting Habitat Suitability For Crassostrea Virginica, Jason D. Tilley

Dissertations

Oyster reefs provide a variety of important ecosystem services. However, the mortality rate of eastern oyster, Crassostrea virginica, the dominant species that produces oyster reefs in the northern Gulf of Mexico, is increasing at an alarming rate due to a variety of abiotic and biological factors. I examined how biophysical factors, including the less-studied fatty acid profiles of the suspended particulate matter on which oysters feed, influenced morphometric condition of C. virginica.

I sampled suspended particulate matter (SPM) and oysters in-situ in the western Mississippi Sound, which historically supported the majority of oyster production in Mississippi waters. Sampling …

Oligomerization Of Amyloid-Β Peptide In The Presence Of Gangliosides–Implications For Alzheimer Disease, Sydney Boyd

Honors Theses

Alzheimer disease (AD) is a neurodegenerative disorder that leads to memory impairment and cognitive dysfunction in elderly populations worldwide. A key characteristic of AD is the formation of neuritic plaques composed of extracellularly deposited aggregates of amyloid-β (Aβ), an intrinsically disordered protein. Although Aβ fibrils were once thought to be the prime initiators of the disease, research has since been shifted to consider soluble, low molecular weight Aβ oligomers as the driving force behind AD toxicity. Due to its origin as a cleavage product of amyloid precursor protein (APP), an integral membrane protein, Aβ is known to perpetually interact with …

Expression And Purification Of The Bacterial Protein Curli Csga And Its Cross-Interactions With Amyloid-B, Leah Grace Cantrell

Honors Theses

One of the main causes of neurodegenerative diseases is aggregation of amyloid proteins that are toxic to the neurons. Proteins like amyloid-β (Aβ) and α-syneuclein (α-syn) form hallmark aggregate lesions that contribute to pathological processes in the brain in Alzheimer and Parkinson’s patients, respectively. Recent ground-breaking studies have suggested a link between the microbiota of the gut and neurodegenerative diseases, called the “gut-brain axis.” It has been long known that the protein, CsgA found in many enteric bacteria, forms amyloid fibers of its own called Curli. Curli fibrils are a structural component of bacterial colonies and maintain the integrity of …

Granulins In Norm And Neurodegenerative Pathologies, Anukool Bhopatkar

Dissertations

Granulins (GRNs) are small, cysteine-rich modules produced from the proteolytic cleavage of the precursor protein called progranulin (PGRN). GRNs are present in the form of seven tandem repeats within the precursor and are known to be produced in the extracellular and in lysosomal environments. In physiology, PGRN and GRNs plays pleiotropic roles such as neuronal growth and differentiation, immunomodulation, wound healing. Recent studies have implicated pathological role for PGRN in Alzheimer disease (AD) and frontotemporal dementia (FTD) but specific mechanism(s) remains unclear. However, potential interactions between GRNs and Ab42 and TDP-43 seem like a plausible underlying mechanism. Studies presented here …

Chemoenzymatic Study Of Coa-Linked Rna In Bacteria, Krishna Sapkota

Dissertations

The ability of RNA to store genetic information and to catalyze biochemical transformations led to the speculation of the existence of RNA world before the evolution of contemporary ribonucleoprotein (RNP) world. Recent discovery of RNA molecules containing metabolic cofactors including coenzyme A and its various thioesters at their 5’ end further supported the RNA world hypothesis as these CoA-linked RNA molecules could be the molecular fossils with very ancient origin. As both RNA and Coenzyme A are believed to have co-existed since last universal common ancestor (LUCA) or even before, the CoA-RNA conjugates in current biology may reveal fundamental molecular …

Cloning, Expression, And Purification Of Fadk And Its Application In Coa-Rna Capture, Jon-Michael L. Stork

Honors Theses

Coenzyme A (CoA) is an important enzyme cofactor involved in acyl transfer reactions. Recently, CoA and its various thioesters were found to exist at the 5’ end of RNA molecules. Although the function of these molecules is still unknown due to difficulties in their isolation, their existence at the 5’ RNA end reveals potentially novel biological roles of RNA. We are exploiting the broad substrate tolerance of the medium chain fatty acid-coenzyme A ligase (FadK), hypothesizing that this enzyme will accept biotinylated medium chain fatty acid substrates, creating biotin-tagged CoA-RNA. This modified CoA-RNA can later be captured using streptavidin affinity …

Elucidating The Interplay Between Sodium Selenite On The Tick Amblyomma Maculatum Selenoprotein Gene Expression, Afnan M. Beauti

Honors Theses

Selenium (Se) is an element recognized as an essential micronutrient in eukaryote organisms. Selenoproteins contain selenium as selenocysteine, the 21st amino acid. Selenium plays a role in cell growth and functioning. At low concentrations, it can induce growth and at high concentrations, it can cause a cell to stop growing and potentially have toxic effects on the cell and organism. When selenium levels are high, oxidative stress results by the production of reactive oxidative species. Selenoproteins, however, can aid the antioxidant response in the cell. Ticks are arthropods of interest, as they are one of few that contain many selenogenes, …

Altering The Structure Of Carboxysomal Carbonic Anhydrase Csosca To Determine The Necessity Of The N-Terminal Domain In Csosca Function, Dana L. Dillistone

Honors Theses

In this project, a DNA construct was designed and developed to remove the first fifty amino acids of the CsoSCA protein in the chemolithotrophic bacterium Halothiobacillus neapolitanus. The csoS3 gene codes for a carbonic anhydrase enzyme (CsoSCA) that is unique to a structure called a carboxysome. Carboxysomes are polyhedral microcompartments where carbon fixation is housed. The carbonic anhydrase is a shell-associated protein that improves the catalytic efficiency of ribulose-bisphosphate carboxylase/oxygenase (RuBisCO), the enzyme that catalyzes the fixation of carbon. By deleting the first fifty amino acids of the carbonic anhydrase, the necessity of the amino acids in carboxysome function was …

The Preparation Of Synthetic Myod Mrna For Cellular Differentiation And Innate Immune Response Downstream Application, James D. Grenn

Honors Theses

The development of therapeutic immune responses from the manipulation of embryonic stem cells (ESCs) via induction of synthetic RNA with 5’ capped and 3’ poly(A)tailed ends would lead to development of stem cell therapy. A necessary step in attaining such a goal is to first produce an mRNA transcript from a plasmid containing the open reading frame (ORF) for a transcription factor for cellular activation.

In this research, the DNA plasmid pMD4 encoding MyoD was transcribed into synthetic mRNA. The plasmid was first amplified using Polymerase Chain Reaction (PCR), and analyzed using gel electrophoresis. The amplified template was purified via …

Characterization Of P43lgrn-3, Kelly G. Hill

Honors Theses

Alzheimer’s Disease (AD) is a progressive neurodegenerative disorder characterized by extensive memory loss and cognitive deficits, which occur due to severe neuronal loss. Two hallmark lesions, intracellular neurofibrillary tangles composed of hyperphosphorylated tau and extracellular neuritic plaques formed by the aggregation of amyloid-β (Aβ), are responsible for the progressive neuronal loss seen in AD brains. Neurotoxic Aβ aggregates are also known to cause inflammation within the brain. It has recently come to light that severe and acute inflammation, such as seen in traumatic brain injury (TBI), may also lead to AD-type dementia. This has raised the question whether some of …

Uncovering The Identity And Metabolism Of Bacterial Coa-Rna, Joseph R. Spangler

Dissertations

Coenzyme A is an indispensable molecule in all known life with roles in metabolism, gene regulation, and macromolecule synthesis. As CoA is derived from RNA itself, it’s incorporation into RNA by in vitro methods has proven useful in research probing the origin of life based on the RNA World theory. The discovery in contemporary bacteria of RNA modified with CoA, however, provided an unexpected twist to previously well-characterized bacterial systems. The identity of sequences associated with CoA-RNA has been elusive since their discovery in 2009 based on the difficulties in isolation while maintaining RNA quality. The aim of this study …

Disorder In Cysteine-Rich Granulin-3 And Its Implication In Alzheimer Disease, Gaurav Ghag

Dissertations

Granulins (GRNs) are a family of small, cysteine-rich proteins that are generated upon proteolytic cleavage of their precursor, progranulin (PGRN) during inflammation. All seven GRNs (1 – 7 or A – G) contain twelve conserved cysteines that form six intramolecular disulfide bonds, rendering this family of proteins unique. GRNs play multiple roles and are involved in a myriad of physiological as well as pathological processes. They are known to a play role in growth and embryonic development, wound healing, and signaling cascades as well as in tumorigenesis. They are also implicated in neurodegenerative diseases like frontotemporal dementia (FTD), Alzheimer disease …

Electrogenerated Chemiluminescence Study Of Semiconductor Nanoparticles Towards Sensitive Detection Of Biomolecules, Yiliyasi Wusimanjiang

Dissertations

The main focus of this dissertation is to unfold the fundamental aspects of electrogenerated chemiluminescence (ECL) generation from semiconductor nanoparticles (also known as quantum dots or QDs) within different ECL systems. The ECL and photo-physical interactions between the CdTe QDs (λ_emission= ~760 nm) and the CdSe QDs (λ_emission= ~550 nm), as well as the effects of carbon nanotubes on ECL of QDs were separately investigated. Optimum experimental conditions for peptide bond formation on an electrode surface through EDC (1-ethyl-3-(-3-dimethylaminopropyl) carbodiimide hydrochloride)/NHS (N-Hydroxysulfosuccinimide) coupling were also revealed using cyclic voltammetry technique. Based on the information …

A Polyaniline-Based Sensor Of Nucleic Acids, Partha Sengupta, Jared Gloria, Marcus K. Parker, Alex S. Flynt

Faculty Publications

Detection of nucleic acids is at the center of diagnostic technologies used in research and the clinic. Standard approaches used in these technologies rely on enzymatic modification that can introduce bias and artifacts. A critical element of next generation detection platforms will be direct molecular sensing, thereby avoiding a need for amplification or labels. Advanced nanomaterials may provide the suitable chemical modalities to realize label-free sensors. Conjugated polymers are ideal for biological sensing, possessing properties compatible with biomolecules and exhibit high sensitivity to localized environmental changes. In this article, a method is presented for detecting nucleic acids using the electroconductive …

Analysis Of The Intricacies Of Substrate Recognition Of High Mobility Group Proteins And Aminoacyl-Trna Synthetases Using Non-Cognate Substrates, Douglas Van Iverson Ii

Dissertations

The studies presented in section 1 (Chapters I-IV) focus on the design and development of nucleic acid four-way junctions (4WJs) to target a member of the high mobility group (HMG) proteins, the proinflammatory cytokine high mobility group box 1 protein (HMGB1). In the present study, hybrid PNA-DNA 4WJs based on a model DNA 4WJ were constructed to improve the thermal stability of 4WJs while maintaining strong binding affinity toward HMGB1. An electrophoretic mobility shift assay (EMSA) was used to examine the binding affinity of an isolated DNA binding domain of HMGB1, the HMGB1 b-box (HMGB1b), toward a set of PNA-DNA …

Developing A Deletion Construct Of The Halothiobacillus Neapolitanus Csos1c Gene, Ellyn K. Dunbar

Honors Theses

The purpose of this research was to develop a deletion construct for the chemoautotrophic bacterium Halothiobacillus neapolitanus, which will be used to generate a mutant lacking a carboxysome shell protein gene. The carboxysome is the location of carbon dioxide fixation. The operon that encodes the carboxysome contains three genes for CsoS1 proteins, the major components of the carboxysome shell. The small CsoS1 proteins self-assemble into hexamers with small central pores. The hexamers arrange into the facets of the icosahedral carboxysome shell. The pores are believed to be involved in selective diffusion of materials necessary for carbon dioxide fixation across the …

Evaluation Of The Protein Recognition Properties Of Peptide Nucleic Acids, Crystal Cox Serrano

Master's Theses

The objective is to evaluate the ability of aminoacyl-tRNA synthetases (aaRS) to recognize the non-standard nucleic acid, PNA (peptide nucleic acid). PNA has immense potential in biomedical applications due to its increased thermostability and nuclease resistance over natural nucleic acids. PNA represents a superior alternative to natural nucleic acids in many biomedical applications due to its specificity, strong binding, and nuclease resistance. This study is the initial data set that indicates PNA are recognized by translation enzymes.

Granulins As Inflammatory Mediators In Alzheimer Disease, Randi G. Reed

Honors Theses

Alzheimer disease (AD) is a neurodegenerative disorder characterized by severe memory deficit and cognitive decline among the elderly. This degeneration is caused by the aggregation and deposition of a protein called amyloid-β (Aβ) in the brain. Aggregation of Aβ causes neuroinflammation in addition to other toxic events. However, it is unclear whether inflammation from an external source, such as from a traumatic brain injury (TBI), could trigger Aβ aggregation. In this context, several pro-inflammatory mediators such as cytokines and chemokines have been suspects. It is now hypothesized that a group of proteins called granulins (Grns) are unique inflammatory mediators that …

Propagation Of Oligomeric Α-Synuclein And Amyloid-Β: Implications For Parkinson's And Alzheimer's Diseases, Matthew Stephen Planchard

Master's Theses

The aggregation of amyloidogenic proteins is a critical event in the pathology of a variety of neurodegenerative diseases, including Alzheimer’s disease (AD) and Parkinson’s disease (PD). The proteins α-synuclein (αS) and amyloid-β (Aβ) are involved in the formation of amyloid lesions observed in PD and AD, respectively. Both PD and AD exhibit a significant amount of co-pathology in clinical settings, and the αS and Aβ proteins have been shown to interact in vitro. Recent experimental consensus has shown oligomeric species to be significant, if not primary, sources of toxicity in these diseases. …

Cloning, Expression And Interaction Studies Of The Potential Rubisco Activase Cbbq, Salma A. Dawoud

Honors Theses

Carboxysomes are polyhedral microcompartments found in all cyanobacteria and in many chemoautotrophs. Within their shell they contain the enzyme ribulose-1, 5-bisphosphate carboxylase/oxygenase (RubisCO), which fixes CO₂. Downstream of the carboxysome operon there is another gene cluster containing several genes that may enhance carboxysome function. Two of these genes, cbbQ and cbbO, encode potential RubisCO activases. Using recombinant CbbQ and CbbO protein, and RubisCO isolated from carboxysomes the interaction between these proteins was studied. The CbbO and CbbQ proteins were both His tagged, allowing them to be purified with Ni²⁺-NTA column chromatography. Each of these tagged …

Biocompatibility Of Synthetic Poly(Ester Urethane)/Polyhedral Oligomeric Silsesquioxane Matrices With Embryonic Stem Cell Proliferation And Differentiation, Yan-Lin Guo, Wenshou Wang, Joshua U. Otaigbe

Faculty Publications

Incorporation of polyhedral oligomeric silsesquioxanes (POSS) into poly(ester urethanes) (PEU) as a building block results in a PEU/POSS hybrid polymer with increased mechanical strength and thermostability. An attractive feature of the new polymer is that it forms a porous matrix when cast in the form of a thin film, making it potentially useful in tissue engineering. In this study, we present detailed microscopic analysis of the PEU/POSS matrix and demonstrate its biocompatibility with cell culture. The PEU/POSS polymer forms a continuous porous matrix with open pores and interconnected grooves. From SEM image analysis, it is calculated that there are about …

Biochemical Characterization Of Two Yeast Paralogous Proteins Mth1 And Std1, Satish Pasula

Dissertations

Glucose is the most abundant monosaccharide and preferred carbon and energy source for most cells. Many organisms have evolved sophisticated means to sense glucose and respond to it appropriately. The budding yeast, Saccharomyces cerevisiae senses glucose through two transmembrane proteins, Snf3 and Rgt2. In the presence of extracellular glucose Snf3 and Rgt2 generate an intracellular signal that leads to the degradation of Mthl and Stdl, thereby inducing the expression of hexose transporter genes (EXT) by inhibiting the function of Rgtl, a transcriptional repressor of HXT genes. Mthl and Stdl are degraded via the Yckl/2 Kinase-SCFGrrl-26S proteasome pathway triggered by the …