Biochemistry, Biophysics, and Structural Biology Commons^™

Protein Structures Under Physiological Conditions, Karl Michael Bertrand

Legacy Theses & Dissertations (2009 - 2024)

My research focused on the evaluation of protein structures and protein dynamics inside eukaryotic cells under physiological conditions. The primary analyses of my research involved the use of in-cell Nuclear Magnetic Resonance spectroscopy using Heteronuclear Single Quantum Coherence experiments. This allowed me to visualize protein structures at an atomic resolution level, as well as, study the interactions of these proteins with small molecules.

Molecular Biochemical Study Of The Cataract-Associated Mutants In Human Gammad-Crystallin, Cindy Yeung

Legacy Theses & Dissertations (2009 - 2024)

Cataract is a vision impairment disease that leads to light scattering and opacity of the eye lens. It is recognized as the most prevalent cause of blindness. Most cataracts occur due to aging, but those that occur in childhood or in young adults are typically of genetic origin. In the young (developing) eye, post–surgical complications such as glaucoma are common after cataract surgery. Hence there is a great need to develop non-surgical intervention strategies for the treatment of cataract. Here, two cataract-associated mutants of human γD-crystallin (HGD): i) Arg76 to Ser (R76S) and ii) a truncation mutation at Trp156 (W156X …

Mutational Analysis Of An Rna Aptamer, Deblina Biswas

Legacy Theses & Dissertations (2009 - 2024)

Aptamers are oligonucleotide molecules (DNA or RNA) that are selected against specific target proteins, nucleic acids or small molecules by using the well established technique of SELEX (Systemic Evolution of Ligands by Exponential Enrichment). Their ability to bind with high affinity and specificity has opened up a wide arena of scientific applications. Aptamers have been shown to have many potential therapeutic and bioanalytical contributions such as in the treatment of diseases like cancer, cardiovascular and other major ailments. The aim of this dissertation is to study and analyze double transversion involving 4 bases in a RNA aptamer molecule: MinB4-au-flipS clamp …

Structural Biology Of The Receptor For Advanced Glycation End Products (Rage), Jing Xue

Legacy Theses & Dissertations (2009 - 2024)

The receptor for advanced glycated end products (RAGE), due to its location in the major histocompatibility complex class III (MHC III) region, suggests its involvement in immune responses. Because RAGE has been linked to complications of diabetes and chronic inflammation, the severity of Alzheimer's disease and cancer, it's significant to understanding the biological mechanism of signal transduction of RAGE.

New Tools To Study Amyloid Fibrils And Intrinsically Disordered Proteins In Vitro And In Vivo, Jacqueline D. Washington

Legacy Theses & Dissertations (2009 - 2024)

Amyloid fibrils are β-sheet-rich protein aggregates commonly found in the organs and tissues of patients with various amyloid-associated diseases. The structure of insulin fibrils was characterized by deep ultraviolet resonance Raman and Nuclear Magnetic Resonance spectroscopy combined with hydrogen-deuterium exchange. Our new approach of combining NMR and Raman spectroscopy with molecular dynamic simulations for characterizing amyloid fibrils provided exclusive knowledge about fibril structure at amino acid residue resolution.

Y-Family Dna Polymerases : Mechanism Of Single-Base Deletion And Effect Of Pcna Interaction, Yifeng Wu

Legacy Theses & Dissertations (2009 - 2024)

Translesional synthesis (TLS) by specialized Y-family DNA polymerases is a DNA damage tolerance pathway to bypass DNA lesions that have not been repaired by other DNA repair mechanisms. Despite their valuable lesion bypass ability, the Y-family DNA polymerases display a much lower fidelity upon replicating undamaged DNA. Their activity therefore needs to be well regulated in the cell. In our first project, we examined how an archaeal Y-family DNA polymerase, Dpo4, makes single-base deletions during replication. Dpo4 belongs to the DinB subfamily which is known to make single-base deletions. Although previous studies suggested that it uses a dNTP-stabilized misalignment mechanism, …

Characterizing The Effects Of Glutaraldehyde On The Ryanodine Receptor Calcium Release Channel, Joshua Dov Strauss

Legacy Theses & Dissertations (2009 - 2024)

Ryanodine receptors (RyR) are large intracellular calcium release channels, which

Structure-Activity Relationship Of 2,3-Benzodiazepin-4-Ones As Noncompetitive Ampa Receptor Antagonists, Mohammad Shaban Qneibi

Legacy Theses & Dissertations (2009 - 2024)

2,3-Benzodiazepin-4-one (BDZ-11) derivatives are a special group of 2,3-benzodiazepine compounds. These compounds have been previously synthesized as AMPA receptor inhibitors, and they are also potential drugs for the treatment of various neurological diseases involving excessive AMPA receptor activity. AMPA receptors are a subtype of glutamate ion channel proteins and are responsible for the majority of excitatory neurotransmission in the mammalian central nervous system. Specifically, they are critically involved in neuronal development and brain activities, including learning and memory. Overstimulation of AMPA receptors is associated with some neurological diseases such as ALS, stroke and Alzheimer's disease. Despite the fact that hundreds …

The Sxrna Platform : Biophysical Characterization Of Trans-Acting Rna Switches For Gene Expression Control, Christine Bazinet

Legacy Theses & Dissertations (2009 - 2024)

In the past decade, microRNAs (miRNAs) have become a hot topic in biochemical research. These very short ~ 22 nucleotide strands of RNA have been recognized as a functional RNA group and found to play a key role in mediating gene regulation along with RNA-binding proteins (RBPs). For example, by binding to the histone stem-loop (HSL) region of the target mRNA, normally in the 3'- untranslated region (UTR), gene expression can either be activated or deactivated for up or down regulation of the gene product. The majority of miRNAs have been found to down regulate gene expression upon binding to …

2'-O-Methyl Substitutions In The Yeast Telomerase Pseudoknot And Their Effects On Telomerase Activity, Katelyn Mae Jasper

Legacy Theses & Dissertations (2009 - 2024)

Telomerase is a ribonucleoprotein (RNP) complex that synthesizes telomeric repeats at the ends of linear chromosomes to form the DNA–protein complexes known as telomeres. Telomeres protect the ends of chromosomes from degradation during replication due to the end-replication problem. When replication occurs, gaps are created at the beginning of the lagging and leading strands that result in the loss of a small amount of DNA at every replication cycle. By adding thousands of copies of telomeric repeats, the repeats are lost in the replication process and not precious genetic information. The telomerase RNA varies drastically among different species both in …

(S)-Trifluoroselenomethionine : A New Non-Natural Amino Acid With Enhanced Methioninase-Induced Cytotoxicity Toward Human Colon Cancer Cells, Stephene Nathele Lodge

Legacy Theses & Dissertations (2009 - 2024)

Abstract

Voltage-Controlled Enzyme-Catalyzed Glucose–Gluconolactone Conversion Using A Field-Effect Enzymatic Detector, Siu Tung Yau, Yan Xu, Yang Song, Ye Feng, Jiapeng Wang

Chemistry Faculty Publications

The field-effect enzymatic detection (FEED) technique was used to control the kinetics of the enzymatic conversion of glucose to gluconolactone. The glucose–gluconolactone conversion occurring at an enzyme-immobilized electrode, a well-studied process, was confirmed using mass spectrometry. Electrochemical studies showed that the glucose oxidation current depends on the gating voltage VG and the ion concentration of the sample solution. Additionally, the depletion of glucose in the sample also showed a dependence on VG. FEED was used to detect H2O2 on the zepto-molar level in order to show the ultrasensitive detection capability of the technique. These results, while providing evidence for the …

Rhodotorula Minuta Fungemia In A Ewe Lamb, C. G. Chitko-Mckown, K. A. Leymaster, M. P. Heaton, D. D. Griffin, J. K. Veatch, S. A. Jones, M. L. Clawson

School of Veterinary and Biomedical Sciences: Faculty Publications

An 8-month-old crossbred ewe, normal upon physical examination, was humanely euthanized for tissue collection. After approximately 3 weeks in tissue culture, fungi began budding out of cells obtained from the choroid plexus. After an additional 3 weeks, budding was observed in kidney cell cultures and eventually in monocyte cultures as well. Serum from the lamb was submitted to the Veterinary Diagnostic Laboratory at Colorado State University for fungal diagnosis and was found negative for Aspergillus, Blastomyces, Coccidioidomycosis and Histoplasmosis. DNA was isolated from fungi collected from tissue culture supernatants and used in a set of pan-fungal PCR assays with …

Interferon Inducible Protein Ifi35 Negatively Regulates Rig-I Antiviral Signaling And Supports Vesicular Stomatitis Virus Replication, Anshuman Das, Phat X. Dinh, Debasis Panda, Asit K. Pattnaik

School of Veterinary and Biomedical Sciences: Faculty Publications

In a genome-wide siRNA screen, we recently identified the interferon (IFN) inducible protein 35 (IFI35, also known as IFP35) as a factor required for VSV infection. Studies reported here were conducted to further understand the role and requirement of IFI35 in VSV infection. Consistent with the siRNA screening data, we found that depletion of IFI35 led to reduced VSV replication at the level of viral gene expression. Although no direct interaction of IFI35 with the viral replication machinery was observed, we found that IFI35 negatively regulated the host innate immune response and rescued poly(I:C)-induced inhibition of VSV replication. Promoter-driven reporter …

Sequence-Optimized And Targeted Double-Stranded Rna As A Therapeutic Antiviral Treatment Against Infectious Myonecrosis Virus In Litopenaeus Vannamei, J. Dustin Loy, Duan S. Loy, Mark A. Mogler, Bruce Janke, Kurt Kamrud, D. L. Hank Harris, Lyric C. Bartholomay

School of Veterinary and Biomedical Sciences: Faculty Publications

Infectious myonecrosis virus (IMNV) is a significant and emerging pathogen that has a tremendous impact on the culture of the Pacific white shrimp Litopenaeus vannamei. IMNV first emerged in Brazil in 2002 and subsequently spread to Indonesia, causing large economic losses in both countries. No existing therapeutic treatments or effective interventions currently exist for IMNV. RNA interference (RNAi) is an effective technique for preventing viral disease in shrimp. Here, we describe the efficacy of a double-stranded RNA (dsRNA) applied as an antiviral therapeutic following virus challenge. The antiviral molecule is an optimized dsRNA construct that targets an IMNV sequence …

Bovine Herpes Virus 1 (Bhv-1) And Herpes Simplex Virus Type 1 (Hsv-1) Promote Survival Of Latently Infected Sensory Neurons, In Part By Inhibiting Apoptosis, Clinton Jones

School of Veterinary and Biomedical Sciences: Faculty Publications

α-Herpesvirinae subfamily members, including herpes simplex virus type 1 (HSV-1) and bovine herpes virus 1 (BHV-1), initiate infection in mucosal surfaces. BHV-1 and HSV-1 enter sensory neurons by cell-cell spread where a burst of viral gene expression occurs. When compared to non-neuronal cells, viral gene expression is quickly extinguished in sensory neurons resulting in neuronal survival and latency. The HSV-1 latency associated transcript (LAT), which is abundantly expressed in latently infected neurons, inhibits apoptosis, viral transcription, and productive infection, and directly or indirectly enhances reactivation from latency in small animal models. Three anti-apoptosis genes can be substituted for LAT, which …

Molecular Considerations In The Design Of Novel Alpha/Beta Hydrolase Inhibitors, Shana Victoria Stoddard

Electronic Theses and Dissertations

Alpha/beta hydrolases (ABHs) are a superfamily of hydrolytic enzymes that process a wide variety of substrates. A subfamily of ABHs called carboxylesterases (CEs) are important enzymes that catalyze biological detoxification, hydrolysis of certain pesticides, and metabolism of many esterified drugs. The chemotherapy drug irinotecan used for treatment of colorectal cancer is metabolized to SN-38, the active drug metabolite, by two CE isozymes CES1 (localized in the liver) and CES2 (localized in the small intestines). CES2's ability to activate irinotecan at a faster rate than CES1 creates a localization of activated SN-38 in the gut epithelium, resulting in the dose limiting …

Role Of The Endocannabinoid System In Cardiac Remodeling, Monica Gole

Electronic Theses and Dissertations

The endocannabinoid system and CB1 and CB2 cannabinoid receptors have been implicated in cardiac remodeling with CB1 inhibition and CB2 activation reported to be beneficial. However, the role of these receptors in cardiac remodeling secondary to hypertension is not known. Moreover, the functions of the CB1 and CB2 cannabinoid receptors in cardiac ventricular fibroblasts and their downstream fibrotic pathways has not yet been elucidated. This study was designed to determine the temporal myocardial expression pattern of CB1 and CB2 receptors in cardiac remodeling secondary to pressure overload (PO) induced hypertension at 3, 5, 14 and 28 days. The expression of …

The Mononuclear Metal Center Of Type-I Dihydroorotase From Aquifex Aeolicus, Brian Fp Edwards, Roshini Fernando, Philip D. Martin, Edward Grimley, Melissa Cordes, Asmita Vaishnav, Joseph S. Brunzelle, Hedeel Evans, David R. Evans

Wayne State University Associated BioMed Central Scholarship

Abstract

Background

Dihydroorotase (DHO) is a zinc metalloenzyme, although the number of active site zinc ions has been controversial. E. coli DHO was initially thought to have a mononuclear metal center, but the subsequent X-ray structure clearly showed two zinc ions, α and β, at the catalytic site. Aquifex aeolicus DHO, is a dodecamer comprised of six DHO and six aspartate transcarbamoylase (ATC) subunits. The isolated DHO monomer, which lacks catalytic activity, has an intact α-site and conserved β-site ligands, but the geometry of the second metal binding site is completely disrupted. However, the putative β-site is restored when the …

Disulfide By Design 2.0: A Web-Based Tool For Disulfide Engineering In Proteins, Douglas B. Craig, Alan A. Dombkowski

Wayne State University Associated BioMed Central Scholarship

Abstract

Background

Disulfide engineering is an important biotechnological tool that has advanced a wide range of research. The introduction of novel disulfide bonds into proteins has been used extensively to improve protein stability, modify functional characteristics, and to assist in the study of protein dynamics. Successful use of this technology is greatly enhanced by software that can predict pairs of residues that will likely form a disulfide bond if mutated to cysteines.

Results

We had previously developed and distributed software for this purpose: Disulfide by Design (DbD). The original DbD program has been widely used; however, it has a number …

Role Of Phosphoylation Of The Mycobacterium Tuberculosis Proteasome In Oxidative Stress Response And Regulation Of Homoserine Kinase (Thrb) In Corynebacterium Glutamicum, Heather Baun

Wayne State University Theses

Part 1:

One-third of the world's population is infected with the latent form of Mycobacterium tuberculosis. M. tuberculosis must be able to control this latent even in response to nitrosative and oxidative stress conferred by the host. It is not well known how M. tuberculosis controls this state.

One possibility for this control is through its eleven eukaryotic-like serine/threonine protein kinases. Our lab has focused research on two eukaryotic-like ser/thr protein kinases, PknA and PknB. It has been found that PknA expression may affect the stability or the assembly of the proteasome complex, which is essential for M. tuberculosis to …

Spliceosomal Prp24 Unwinds A Minimal U2/U6 Complex From Yeast, Chandani Manoja Warnasooriya

Wayne State University Theses

Splicing plays a major role in eukaryotic gene expression by processing pre-mRNA to form mature mRNA. Pre-mRNAs undergo splicing to remove introns, non–protein coding regions, and religate exons, protein coding regions. This process is catalyzed by the spliceosome, which consists of five small nuclear ribonucleoprotein particles (snRNPs: U1, U2, U4, U5 and U6) and numerous protein factors. Proper assembly of spliceosomal components is critical for function, and thus, defects in assembly can be lethal. Several spliceosomal proteins facilitate structural rearrangements important for spliceosomal assembly and function. Prp24 is an essential factor in U6 snRNP assembly, and it has been proposed …

Investigation Of Bacterial Rna-Directed Dna Methylation Via Dcm And Hfq, Dandan Li

Wayne State University Theses

Bacterial small RNAs and the RNA chaperone Hfq play crucial roles in post-transcriptional gene regulation, often as parts of stress-response pathways, but little is known about their roles in regulation of gene transcription. A recent report showed that changes in methylation patterns caused by DNA cytosine methyltransferase (Dcm) were linked to gene regulation occurring during the transition to stationary phase. Here, we show that Dcm involves in the stress responses under nutrient starvation and cold stress. Dcm and Hfq together mediate gene expression under cold stress. Hfq promotes Dcm-catalyzed cytosine methylation at specific sites near the rpoS promoter, which is …

Lc3i And Lc3ii As Autophagy Markers For The Development And Improvement Of Products And Techniques Used In Research, Caitlin Jane Williams

Wayne State University Theses

Abstract

LC3I and LC3II as Autphagy Markers for the Development and Improvement of Products and Techniques used in Research

By

Caitlin J. Williams

May 2013

Advisor: Dr. Robert Akins

Major: Biochemistry & Molecular Biology

Degree: Master of Science

Autophagy is an intracellular process that functions to maintain homeostasis in the cell by degrading misfolded proteins, old or nonfunctioning organelles, and outside invaders such as bacteria or viruses. This process can be split into three different types, microautophagy, chaperone mediated autophagy and macroautophagy. Macroautophagy is the most commonly studied form and is believed to be regulated by Atg proteins, as well …

Effect Of Folate Deficiency And Aging On Mtor Signaling Network In The Liver Of Dna Polymerase B Haploinsufficient Mice, Amanda Lynn Arrabi

Wayne State University Theses

EFFECT OF FOLATE DEFICIENCY AND AGING ON mTOR SIGNALING NETWORK IN THE LIVER OF DNA POLYMERASE B HAPLOINSUFFICIENT MICE

by

AMANDA ARRABI

August 2013

Advisor: Dr. Ahmad R. Heydari

Major: Nutrition and Food Science

Degree: Masters of Science

The interaction between the dietary nutrients and the human genome causes some alterations in the molecular pathways that could significantly impact the development of cancer. The main approach to such concerns is to closely observe how the diet modifies the gene expressions that control several cellular processes in the human body. Folate deficiency (FD) has established an important role in modulating the …

Cysteine Desulfurase And Isd11: A Drosophila Model, Rahul Ravindran Nair

Wayne State University Theses

ABSTRACT

Cysteine desulfurase and Isd11: A Drosophila model

Iron-sulfur clusters are cofactors with evolutionary origins that date back to the pre-biotic world. Ever since life originated, these cofactors have intermingled with proteins to play vital roles in sustaining life. My research focuses on one such protein, the cysteine desulfurase (Nfs) that has the PLP cofactor incorporated in its active site and avails of the catalytic property of PLP to provide sulphur for Iron-sulfur cluster biogenesis and assembly in a cell. Interestingly, in a eukaryotic cell, despite the versatility of PLP, cysteine desulfurase's role as a "sulphur-extractor" is incomplete without another …

Studies Of Regulated Exocytosis From Neuroendocrine Cells, Madhurima Das

Wayne State University Theses

In this thesis we study cargo release and fusion pore dilation during calcium triggered exocytosis and the co-localization of calcium sensing proteins essential for exocytosis, in neuroendocrine cells.

Pancreatic beta cells secrete several hormones, the most studied one being insulin. C-peptide is a protein which is co-stored with and secreted from the same vesicles as insulin. It is found in the soluble phase unlike insulin, which is found in the dense core. The pancreatic beta cells also secrete the Chromogranin B (CgB) which is mostly found in the dense cores of secretory vesicles. In chapter 1, we found that CgB, …

Optimization Of Labeling Techniques; Determination Of Best Parameter For Olfactory Mucosal Progenitor Cell Delivery And Study Of Effects Of Methylene Blue And Polyethylene Glycol In An Animal Model Of Spinal Cord Injury, Kiran Kumar Rokkappanavar

Wayne State University Theses

In the United States, the incidence of TSCI is about 40 per million persons per year, with approximately 250,000 living survivors of TSCI in the United States in July 2005. A number of human clinical trials (85) are ongoing using stem cells to evaluate the methods to reduce the injury following TBI. Among the stem cells, olfactory mucosal progenitor cells have several advantages with respect to ease of obtaining, fate of cells, etc. Our study was intended to optimize the labeling technique and to determine the best parameter to deliver the stem cells to the site of injury. We found …

Characterizing Cyclin J By Identifying Conserved Protein-Protein Interactions, Phillip Jacob Selman

Wayne State University Theses

Cyclins are proteins that bind to Cyclin-dependent kinases, or Cdks, through a conserved domain called the Cyclin Box. Many Cyclins regulate the cell cycle. A few Cyclins impact cellular processes outside of the cell cycle. Also, a few Cyclins have poorly understood functions.

Cyclin J is a member of the Cyclin superfamily of proteins. Cyclin J is conserved among all metazoans, but is presently not well understood. All the research done on Cyclin J has been done in Drosophila.

Its mRNA is present in the early embryo, then disappears, only to reappear in adult females. When probing protein extracts with …

Histone Deacetylation As A Mechanism Of Ypel3 Down-Regulation In Er-A Positive Breast Cancer Cell Lines, Remah Ali

Browse all Theses and Dissertations

YPEL3 is a growth inhibitory gene that was established by our laboratory to trigger senescence in both primary and tumor cells. We recently reported YPEL3 to be directly repressed by estrogen in estrogen receptor-a positive (ER-a +ve) breast cancer cell lines. Here, we set out to determine whether the repression of YPEL3 involves histone deacetylation and, if so, elucidate the molecular mechanism of this activity. MCF-7, T-47D, and ZR-75.1 (all ER-a +ve) breast cancer cell lines were treated with varying doses of the global histone deacetylase inhibitors (HDACIs), trichostatin A (TSA) or suberoyl anilide hydroxamic acid (SAHA), for 24, 48 …