Biochemistry, Biophysics, and Structural Biology Commons^™

Extraction Of Transcriptional Regulators For The Polyhydroxyalkanoate Depolymerase Gene From Streptomyces Nymphaeiformis, Kara B. Eppard, Stephen F. Baron

Honors Projects

Plastic waste has become an increasingly prevalent environmental pollutant. This problem is exacerbated by the inability of plastic to degrade under most natural conditions. In contrast, polyhydroxyalkanoates (PHAs) are biologically produced, plastic-like polymers that can be broken down and metabolized by bacteria. The bacterium Streptomyces nymphaeiformis can degrade the PHA, polyhydroxybutrate (PHB), using an extracellular PHB depolymerase, which is encoded by the phaZ gene. PHB depolymerase is synthesized only in the presence of PHB or its monomer, but not glucose, suggesting that transcription of phaZ is regulated, presumably by transcriptional regulatory proteins that bind to its promoter region. The DNA …

Host And Symbiont-Specific Patterns Of Gene Expression In Response To Cold Stress In The Temperate Coral Astrangia Poculata, Kellie Navarro

Honors Projects

The coral Astrangia poculata inhabits hard-bottom environments from the Gulf of Mexico to Massachusetts and withstands large seasonal variation in temperature (–2 to 26 °C). This thermal range and its ability to live in a facultative symbiosis makes this species an ideal model system for investigating stress responses to ocean temperature variation. Although it has been shown that aposymbiotic A. poculata upregulates more genes in response to cold stress than heat stress, the transcriptomic response of the holobiont (coral host and symbiotic algae) to stress is unknown. In this study, we characterize changes in gene expression in both the host …

Screening Anti-Pd-L2 Peptides As Antitumor Ligands Using Phage Display, Chien Tran Phuoc

Honors Projects

Cancer still remains one of the top leading causes of death in America. Recently, programmed cell death protein 1 (PD-1) blockades have been demonstrated to be highly effective against various types of cancer. By blocking PD-1 from binding with their ligands (PD-L1 and PD-L2), the “off” signal to the immune system is inhibited, hence reinvigorating the immune cells to kill tumor cells. To date, despite PD-L1 and PD-L2 both interacting with PD-1, research efforts have only been focused on developing anti-PD-L1 inhibitors. Therefore, the work of this honor project has focused on finding anti-PD-L2 peptides by phage display, with the …

Overexpression Of Foxf2 In Prostate Cancer Cells Causes An Increase In Cancer Cell Apoptosis And A Decrease In Proliferation, Vaughn Poon

Honors Projects

FOXF2 is a transcription factor that plays a crucial role in organ development, and recent studies have shown that it suppresses tumor growth and progression in mouse prostate models by attenuating the cancer-associated fibroblasts (CAF) phenotype and transcriptionally downregulating Cxcl5. However, the effects of FOXF2 overexpression in prostate cancer cells have not been extensively studied. Here, we investigate the impact of FOXF2 overexpression in prostate cancer cells and demonstrate that it leads to a significant increase in cell apoptosis and a decrease in proliferation. These findings suggest that FOXF2 may have potential as an immunotherapy drug target for prostate …

Bci Validation: Yfiler Plus Kit Validation, Chloe Koon

Honors Projects

In order to have access and use the Combined DNA Index System (CODIS), it is required by the Federal Bureau of Investigation's Quality Assurance Standards (FBI-QAS) for Forensic DNA Units to complete comprehensive validation studies on all equipment, materials, and methods used in the process of DNA analysis. This study continues validation previous done within the Ohio Bureau of Criminal Investigation (BCI). YFiler Plus is a DNA amplification kit that is used in cases where it is most useful to examine only male DNA profile(s) within a sample by amplifying loci only on the Y-Chromosome. Components of this study include …

The Role Of Elmo5 In Arabidopsis Thaliana Cell Adhesion, Isabel Kristina Ball

Honors Projects

Plant cell growth and development relies on proper cellular adhesion. As the extracellular matrix serves as the area of connection between two cells, its synthesis and maintenance are essential for cellular adhesion. The middle lamella region, the layer of the extracellular matrix between two adjacent cell walls, is diffuse with the polysaccharide pectin due to its delivery by Golgi vesicles early during cell division. A Ruthenium Red screen for cellular adhesion mutants identified the family of 5 ELMO proteins that are critical for proper cellular adhesion. To further our understanding of plant cellular adhesion and pathways of pectin synthesis and …

Characterization Of The Elmo2 Protein That Mediates Cell Adhesion In Arabidopsis Thaliana, Devaki Rajiv

Honors Projects

The binding of adjacent cells to one another, or cell adhesion, is critical for the growth and development of multicellular organisms. In plant cells, much evidence suggests that the amount and modification of pectin in the cell wall largely determines how well cell adhesion occurs. ELMO1 is a Golgi protein involved in pectin-mediated cellular adhesion, and mutations in ELMO1 lead to disrupted cell organization in Arabidopsis. ELMO1 is predicted to be a scaffold for pectin biosynthesis enzymes, and thus its absence leads to the adhesion-defective phenotype of elmo1-/- plants. There are four other ELMO homologues (ELMO2,3,4 and 5) which remain …

Genetic Analysis Of Adhesion Protein Elmo3 In Arabidopsis Thaliana, Garrison Asper

Honors Projects

The Extracellular Matrix (ECM) between plant cells is vital for structure, development, and intercellular adhesion. A pectin rich layer in between cells, the middle lamella, is largely responsible for regulating the adhesive properties of adjacent plant cells. Homogalacturonan (HG) pectin, the most common, is synthesized in the Golgi and secreted into the ECM where it undergoes calcium crosslinking, increasing its adhesive properties. Mutations in proteins essential for HG synthesis can reveal a severe adhesion defective phenotype, where the hypocotyls of dark grown Arabidopsis exhibit cell sloughing, curling, and general disorganization. A family of five ELMO proteins are suspected to act …

Alterations To The Brain Following Traumatic Brain Injury, Jacqueline Mader

Honors Projects

Traumatic brain injuries (TBIs) have been labeled as a modern-day epidemic, increasing exponentially with the advancement of technology and society. Gaining a better understanding of the cognitive paths, including the chemical and electrical signals of the brain, neural correlates, and possible interventions for TBI patients allows for the best possible outcome for every patient, and allows for the further advancement of care. By revising and reassessing the ways in which TBIs are categorized and described the prognosis for recovery paints a more realistic view for each individual patient case. The symptoms and impairments that may occur post-injury can be monitored …

Salicylic Acid Response To Simulated Herbivory In Geographically Distinct T. Heterophylla And H. Discolor Populations, Amy E. Castle

Honors Projects

It is commonly known that plants may produce salicylic acid as a chemical defense response to wounding, although the phenomenon has usually been observed with regard to insect herbivory. Stem and leaf tissue of two species, Tsuga heterophylla and Holodiscus discolor, which are often eaten by deer, were extracted in methanol and analyzed by HPLC to quantify salicylic acid concentration in experimentally wounded or control samples. No salicylic acid response was detectable in T. heterophylla, suggesting it is a less useful candidate species for future study. Some but not all H. discolorsamples had a measurable salicylic acid …

Pectin And Alginate Extraction To Treat Liquid Cafo Manure, Clare Sunderman

Honors Projects

For this project, various extraction methods were used to extract pectin from Pastinaca Sativa and alginate from Macrocystis. These extractions were then dried and used in treating 250mL of manure along with a CaCl₂ or FeCl₃ coagulant. It was found that CaCl₂ was not as effective as FeCl₃ in coagulating manure. But the results obtained suggest that pectin and alginate obtained with a simpler extraction method is just as effective as the highly purified and refined pectin and alginate produced for the food industry, in the treatment of CAFO manure. The liquid portion of the …

Semaphorin-Induced Plasticity In The Nervous System Of The Cricket, Gryllus Bimaculatus, Alicia G. Edwards

Honors Projects

The adult auditory system of the cricket, Gryllus bimaculatus, exhibits a rare example of neuronal plasticity. Upon deafferentation, we observe medial dendrites that normally respect the midline of the PTG in the central nervous system sprouting across the boundary and forming synaptic connections with the contralateral auditory afferents. The Horch Lab has investigated key molecular factors that might play a causal role in this paradigm. Specifically, the protein Sema1a.2 comes from a guidance molecule family and has a role in developmental neuronal plasticity in other organisms. In this study, I explored the role of Sema1a.2 in the neuronal plasticity of …

Genetic Analysis Of Cellular Adhesion In Arabidopsis Thaliana, Andrew Close Bolender

Honors Projects

Plant cell adhesion is mediated by the extracellular matrix (ECM) or cell wall and plays an important role in plant morphogenesis and development. The amount, modification, and cleavage of pectin in the cell wall are major contributors to the adhesive properties of the ECM. To gain a more complete picture of plant cell adhesion processes, Arabidopsis thaliana seedlings were previously mutagenized and screened for hypocotyl adhesion defects. Genomic sequencing of one plant exhibiting an adhesion defect, isolate 242, showed that two mutations, one in cellulose synthase (CesA1) and another in a sugar transporter, are candidates for the causative mutation. This …

The Kinetic Signatures Of Antibody Binding To M. Genitalium Adhesin Protein Fragments, Margaret C. Lunn

Honors Projects

Mycoplasma genitalium is a sexually-transmitted bacterial pathogen that persists in patients by adherence to cells through matrix glycoproteins and evasion of host antibodies. The MgpB and MgpC adherence proteins consist of variable and conserved regions. Variable regions undergo antigenic variation to avoid specific antibodies. However, the C-terminus (MgpB-4a) does not vary, is highly immunogenic, and antibodies to this region inhibit attachment and promote bacterial killing in vitro. To better understand how M. genitalium avoids clearance by antibodies to MgpB-4a in vivo we used surface plasmon resonance (SPR) to measure kinetic values of binding events. Binding of polyclonal rabbit antibodies (3935 …

Living Upstream: Kennebec River Influence On Nutrient Regimes And Phytoplankton Communities In Harpswell Sound, Siena Brook Ballance

Honors Projects

Phytoplankton underpin marine trophic systems and biogeochemical cycles. Estuarine and coastal phytoplankton account for 40-50% of global ocean primary productivity and carbon flux making it critical to identify sources of variability. This project focuses on the Kennebec River and Harpswell Sound, a downstream, but hydrologically connected coastal estuary, as a case study of temperate river influence on estuarine nutrient regimes and phytoplankton communities. Phytoplankton pigments and nutrients were analyzed from water samples collected monthly at 8 main-stem rivers stations (2011-2013) and weekly in Harpswell Sound (2008-2017) during ice-free months. Spatial bedrock and land use impacts on river nutrients were investigated …

Life In A Rural Emergency Department: Patients Speak To Underlying, Fundamental Disparities In Physician And Resource Allocation, Jessica Gustad

Honors Projects

In the Journal of Health Services Research and Policy, access to health care was defined as, “The availability of an adequate supply of health care services and the individual’s opportunity to obtain health care when it is wanted or needed” (MacKinney, 2014). Current trends in rural emergency departments are trying to communicate a critical message about the fundamental problems with the distribution of health care services in America today and the inability to properly meet the definition above. In rural communities, patient complaints and behaviors demonstrate the pressing concerns of inadequate access to primary care, limited specialty care, insufficient mental …

Identification Of A Tola Protein Binding Site For Bacterial Toxins, Monica Ferrante

Honors Projects

Group A colicins are proteinaceous bacteriocins encoded by plasmids that exploit the cellular envelope protein TolA to translocate the cell wall barrier and cellular envelope of the bacterium Escherichia coli. These colicins offer protocols for studying certain protein-protein interactions involved in such membrane transport functions. Previous experimentations suggest the carboxyl-terminal domain of TolA protein contains specific amino acid binding regions required for the translocation of group A colicins into E. coli. The amino acid sequence of this domain varies between E. coli and other gram-negative bacterial species. It has been suggested that this diversity could be utilized to …

Evaluation Of Cafo Dairy Manure Treatment To Reduce Nutrient Transport, Jenna Laib

Honors Projects

Excess amounts of phosphorus and nitrogen flowing into Lake Erie from agricultural fields in Northwest Ohio has led to several harmful algal blooms (HABs). One potential source of those nutrients is manure applied to fields for fertilizer. Manure from confined animal feeding operations (CAFOs) is 95-98% water with only ~3% solids and nutrients, thus physical transportation is expensive relative to the value of the agricultural nutrients. Furthermore, once manure nutrients are applied to agricultural fields, they are relatively easily mobilized to waterways by precipitation. More than 800 lab-scale tests have been used to optimize the treatment of CAFO manure with …

Assay Of The Reverse Osmosis Purified Water In The Life Science Building At Bowling Green State University, Ohio, Nicholas Mendenhall, Scott O. Rogers, Neocles B. Leontis

Honors Projects

Contaminated water sources can cause problems for scientific research and result in costly delays and failures of experiments. At Bowling Green State University, the reverse osmosis supply circulating in the Life Sciences Building has been measurably contaminated for nearly three years, corresponding to a change in servicing of the system. While servicing has been accelerated, the contamination in the system remains. The focus of this research was to identify the species of bacteria and fungi growing inside of the water system so that it might alert those servicing the system, and to begin to eliminate the contamination. Reverse osmosis water …

Examining The Role Of Grp And Lik1 In Wall Associated Kinase (Wak) Perception Of Pectin In The Plant Cell Wall, Jack Ryan Mitchell

Honors Projects

Wall associated kinases (WAKs) are cell membrane bound receptor kinases that bind pectin and pectin fragments (OGs).The binding of WAKs to pectin sends a growth signal required for cell elongation and plant development. WAKs bind OGs with higher affinity than native pectin and instead activate a stress response. Glycine rich proteins (GRPs) are secreted cell wall proteins of unknown function. Seven GRPs with 65% sequence similarity are coded on a 90kb locus of Arabidopsis chromosome 2. GRP3 and WAK1 have been shown to bind in vitro, but single null mutations have no discernible phenotype, suggesting that the GRPs are redundant. …

Purification Of A Bacteriophage Protein Involved In Host Range Specificity, Alec Brown

Honors Projects

The Escherichia coli ferric hydroxamate uptake receptor FhuA serves as the receptor for ferrichrome-Fe(III) complexes, with TonB protein energizing the active transport of the complex. The FhuA receptor is exploited by a variety of bacteriophages as a conduit into the cell. Interestingly, certain of these phages carry a gene called “Cor”, the product of which, when cloned and expressed from a plasmid, blocks transport by FhuA. In the present study, components of the cor gene from the bacteriophage ϕ80 were used to construct an IPTG-inducible MalE-Cor-His6 fusion protein, which allowed for affinity purification of the Cor protein. At 61 residues …

The Role Of Protein Kinases Rog1 And Srf6 In The Wak Stress Response Pathway, Jaepil E. Yoon

Honors Projects

No abstract provided.

Characterization Of A Novel Clade Of Transporters In Phytophthora, Stephanie Padula, Paul F. Morris Dr, Howard Casey Cromwell Dr., Menaka Ariyaratne, Andrew Wagner

Honors Projects

The oomycete Phytophthora parasitica has a worldwide distribution and is an economically important pathogen of more than 100 species⁴. RNA-seq analysis showed that one gene, PPTG_16698 has the 5^th highest level of expression of all transport proteins in the zoospore stage, and is highly conserved throughout Phytophthora species. This project attempts to characterize the important biological role that PPTG_16698 plays in P. parasitica and other oomycetes. Three strategies have been implemented to accomplish this goal: growth analysis by heterologous expression in yeast, metabolite analysis in yeast, and construction of a GFP fusion protein to enable localization of …

Annotation And Transcription Start Site Analysis Of Contig70 In Drosophila Biarmipes, Robin Wolschendorf

Honors Projects

The following is my annotation and transcription start site analysis report for contig70 in D. biarmipes to be submitted to the Genomics Education Partnership (GEP) at Washington University in St. Louis. The report was completed in accordance with the guidelines and template set forth by the GEP. There are eight complete genes found in contig70, for which I found the protein coding exon boundaries as well as the transcription start sites for all isoforms. All conclusions and reasoning for them are outlined in the report.

The GEP does genomics research, specifically comparing the genomes of all species of Drosophila. …

Split-Spinach Monitoring Of Rna Aptamer Assembly, Tucker A. Rogers, Grant Andrews

Honors Projects

As insights into RNA’s many diverse cellular roles continue to be gained, interest and applications in RNA self-assembly and dynamics remain at the forefront of structural biology. The bifurcation of functional molecules into nonfunctional fragments provides a useful strategy for controlling and monitoring cellular RNA processes and functionalities. Herein we present the bifurcation of the preexisting Spinach aptamer and demonstrate its utility as a novel split aptamer system for monitoring RNA self-assembly as well as the processing of pre-short interfering substrates. We show for the first time that the Spinach aptamer can be divided into two nonfunctional halves that, once …

The Effect Of Visual Wulst Lesions And Trigeminal Nerve Sectioning On The Discrimination Of Magnetic Inclination In The Homing Pigeon (Columba Livia), Merissa Acerbi

Honors Projects

The ability of homing pigeons to return to their loft from unknown places has fascinated scientists for centuries. It is well established that homing pigeons, like migratory birds, posses an innate magnetic inclination compass to determine direction by measuring the angle between the magnetic field vector and the Earth's surface. Recent work has indicated that the avian magnetic compass is light mediated and appears to mediate magnetic information to the brain. This occurs via a visual pathway with processing in the visual Wulst area of the forebrain. There is, however, also evidence from other avian species that magnetic direction may …

Ufd2a Isoforms With Vsp/P97 And Its Ibmpfd Mutants, Amanda St. Germain

Honors Projects

Using the Yeast2Hybrid system, the binding affinities between Ufd2a variants and VCP/p97 mutants were analyzed. Several VCP/p97 mutants are associated with Inclusion body myositis in association of Paget's disease of the bone and fronto temperal dementia (IMBPFD). It was found that Ufd2a bound strongest to the VCP disease causing mutant A232E. It was also determined that the muscle specific isoform of Ufd2a, Ufd2a III, did not interact with VCP/p97.

Determining The Composition Of The Dwelling Tubes Of Antarctic Pterobranchs, Lukasz J. Sewera

Honors Projects

Pterobranchs are a group of marine invertebrates within the Hemichordata, which share characteristics with both chordates and echinoderms. Pterobranchs live in colonies of secreted tubes, coenicia, which are composed of a gelatinous material of unknown composition. Visually, the tubes appear similar to the tunic of tunicates, a group of invertebrates within the Chordata. The nonproteinaceous tunic of tunicates is composed of cellulose, which is unusual. The goal of this study was to determine the composition of the pterobranch coenicium. Some aspects of pterobranch phylogeny are still unclear even after multiple molecular and morphological studies. Identification of any new shared characteristics …

The Effects Of Β-Mercaptoethanol On The Specific Activity Of Porphobilinogen Synthase Mutants Of The Rhodobacter Sphaeroides Protein, Trefan Archibald '05

Honors Projects

The enzyme porphobilinogen synthase (PBGS) catalyzes the conversion of two molecules of δ-aminolevulinic acid (ALA) into porphobilinogen (PBG) in the first common step of the tetrapyrrole biosynthesis pathway. A homology model of the R. sphaeroides PBGS was created by Dr. Eileen K. Jaffe at Fox Chase Cancer Center based on comparison to the crystal structure of Pseudomonas aeruginosa. The proposed structure suggests that there are four cysteines in close proximity to the active site. Three of these cysteines are not present in the highly similar R. capsulatus sequence. Under oxidizing conditions these residues can potentially participate in the formation of …

Characterizing Conformational Changes Along The Dimerization Helix Of The Global Regulator, Fnr, Using Fluorescence Spectroscopy, Melanie Zupancic '03

Honors Projects

The FNR protein is a transcription factor that allows Escherichia coli to undergo anaerobic cellular respiration. It is known to positively regulate the expression ofseveral genes required for anaerobic respiration as well as negatively regulate genes responsible for aerobic respiration. Consequently, FNR is active under anaerobic conditions and inactive under aerobic conditions. Although the tertiary structure ofFNR is unknown, previous studies have indicated that FNR is inactive in the monomeric state and active in the dimeric state. Thus, it is believed that in anaerobic conditions, FNR undergoes a confonnational change from the monomeric to dimeric state. The mechanism involved in …