Biochemistry Commons^™

Neuronal Gq Structures In Neurodegeneration, Damian S. Mcaninch

Electronic Theses and Dissertations

This study investigates protein nucleic acid interactions between various proteins and G quadruplex (GQ) forming messenger RNAs (mRNAs) in human neurological disorders. GQ structures are formed in DNA/RNA, when four guanine residues form planar tetrads stabilized by Hoogsteen base pairing, that stack forming a GQ structure stabilized by potassium ions. These GQ structures are targeted by the arginine-glycine-glycine (RGG) repeat domain containing RNA-binding domain.

Three RGG domain containing RNA-binding proteins, all of which have been implicated in neurological disorders, and their interactions with GQ forming mRNAs, were investigated in this study: fused in sarcoma (FUS), fragile X mental retardation protein …

Elucidating Structure, Function, And Small Molecular Interactions Of Human Immunodeficiency Virus And Chikungunya Virus, Kristin Nicole Slater

Wayne State University Theses

Abstract HIV-1:

Human immunodeficiency virus-1 (HIV-1) is a widespread, incurable retrovirus known to cause

immunodeficiency and a shortened life span. Despite successful treatment methods, HIV-1

frequently mutates, resulting in antiviral resistance. Many therapies target the HIV-1 protease

(PR), which is responsible for cleaving the viral polyprotein essential for its life cycle. HIV-1 PR

often evades treatment by way of mutations and less commonly through residue insertions. We

have identified a clinical isolate with a five residue insertion between residues 28 and 29.

Through molecular dynamics simulations we analyzed the protease protein structure and

determined that the residue insertion created a …

An Effective Computational Method Incorporating Multiple Secondary Structure Predictions In Topology Determination For Cryo-Em Images, Abhishek Biswas, Desh Ranjan, Mohammad Zubair, Stephanie Zeil, Kamal Al Nasr, Jing He

Computer Science Faculty Publications

A key idea in de novo modeling of a medium-resolution density image obtained from cryo-electron microscopy is to compute the optimal mapping between the secondary structure traces observed in the density image and those predicted on the protein sequence. When secondary structures are not determined precisely, either from the image or from the amino acid sequence of the protein, the computational problem becomes more complex. We present an efficient method that addresses the secondary structure placement problem in presence of multiple secondary structure predictions and computes the optimal mapping. We tested the method using 12 simulated images from alpha-proteins and …

Interaction Of Spliceosomal U2 Snrnp Protein P14 With Its Branch Site Rna Target, William Perea Vargas

Dissertations, Theses, and Capstone Projects

Newly transcribed precursor messenger RNA (pre-mRNA) molecules contain coding sequences (exons) interspersed with non-coding intervening sequences (introns). These introns must be removed in order to generate a continuous coding sequence prior to translation of the message into protein. The mechanism through which these introns are removed is known as pre-mRNA splicing, a two-step reaction catalyzed be a large macromolecular machine, the spliceosome, located in the nucleus of eukaryotic cells. The spliceosome is a protein-directed ribozyme composed of small nuclear RNAs (snRNA) and hundreds of proteins that assemble in a very dynamic process. One of these snRNAs, the U2 snRNA, is …

Mutant Study Of Sinorhizobium Meliloti Proline Utilization A (Puta), Jacob E. Wilkinson, John J. Tanner, Donald F. Becker

UCARE: Research Products

The purpose of this project is to purify and characterize the reaction kinetics of mutant versions the enzyme Proline Utilization A (PutA) in Sinorhizobium meliloti. The enzyme catalyzes the first step in proline metabolism. It has two active sites. The first is proline dehydrogenase (PRODH) which converts proline to pyrroline-5-carboxylate (P5C). The second is P5C dehydrogenase (P5CDH) which converts P5C to glutamate. Although many bacterial organisms have PutA, there are still significant interspecies variations, resulting in an entire family of PutA enzymes. The main difference is the length of the amino acid sequence. This affects the protein’s structure or …

Structural Elucidation Of Aggr-Activated Regulator, Aar, In Enteroaggregative Escherichia Coli, Andrew Heindel

Senior Honors Projects, 2010-2019

Travelers’ Diarrhea is the number one cause of childhood death in the world. Enteroaggregative Escherichia coli (EAEC) is one of the main causes of this disease. EAEC adhere to the surface of the intestine and stack in a brick-like pattern. Via an unstudied quorum-sensing mechanism, these bacteria express a variety of virulence factors that lead to diarrhea. The long-term goal of this research is to elucidate the mechanism by which EAEC changes from benign to virulent. A previously-unstudied open-reading frame in EAEC, AggR activated repressor (Aar), has recently been hypothesized to act as one of the major transcription factors influencing …

Exploring The Effect Of Climate Change On Biological Systems, Nardos Sori

Chemistry & Biochemistry Theses & Dissertations

The present and potential future effect of global warming on the ecosystem has brought climate change to the forefront of scientific inquiry and discussion. For our investigation, we selected two organisms, one from cyanobacteria and one from a cereal plant to determine how climate change may impact these biological systems. The study involved understanding the physiological and adaptive responses at both the genetic and protein function levels to counteract environmental stresses. An increase in atmospheric carbon dioxide is a key factor in global climate change and can lead to alterations in ocean chemistry. Cyanobacteria are important, ancient and ubiquitous organisms …

Cloning And Expression For The Future Characterization Of The Air2 Protein, Emily Sue Frampton

Undergraduate Honors Capstone Projects

Air2 is a eukaryotic protein involved in multiple biological processes including protein-protein interactions as well as RNA binding. Air2 plays a critical role in RNA quality control and also helps regulate post-translational modification of various proteins. Although previous studies have revealed information regarding Air2's roles within a cell, the molecular and structural basis for Air2 function is unclear. Using a codon-optimized version of the Air2 gene, various constructs were created that improved the expression and solubility of Air2. Additionally a co-expression complex of Air2 with a PRMTI mutant, K13S, was made to obtain the Air2 protein with a native binding …

Design And Study Of The Efflux Function Of The Egfp Fused Mexab-Oprm Membrane Transporter In Pseudomonas Aeruginosa Using Spectroscopy, Feng Ding, Kerry J. Lee, Ardeschir Vahedi-Faridi, Hiroshi Yoneyama, Christopher J. Osgood, Xiao-Hong Nancy Xu

Biological Sciences Faculty Publications

Multidrug membrane transporters (efflux pumps) can selectively extrude a variety of structurally and functionally diverse substrates (e.g., chemotoxics, antibiotics), leading to multidrug resistance (MDR) and ineffective treatment of a wide variety of diseases. In this study, we have designed and constructed a fusion gene (egfp-mexB) of N-terminal mexB with C-terminal egfp, inserted it into a plasmid vector (pMMB67EH), and successfully expressed it in the Δ MexB (MexB deletion) strain of Pseudomonas aeruginosato create a new strain that expresses MexA-(EGFP-MexB)-OprM. We characterized the fusion gene using gel electrophoresis and DNA sequencing, and determined its expression in live …

Biochemical And Biophysical Studies Of Novel Features Of Ras-Related Protein Interactions, Kyla Marie Morinini Morris

Graduate Theses and Dissertations

The Ras superfamily of G-proteins are of great research interest for structure-function relationships among proteins as they act as molecular switches in the regulation of various biochemical reactions in the cell. They are regulated by protein-protein interactions targeted to the highly flexible switch regions. Mutations in G-proteins or their effectors may cause alterations in structure and/or function that can lead to overactivity.

The Ras-related protein Cell division cycle 42 (Cdc42) is important in regulating cell-signaling processes. The T35A mutation in Cdc42 leads to a decrease in flexibility of the Switch I region responsible for effector binding. The kinetics of the …

Effect Of Anthraquinone Dyes On The Carbohydrate, Protein And Lipid Content In The Muscle Of Channa Punctatus And Cyprinus Carpio, Rajee Olaganathan, Jamila Patterson

Publications

Channa punctatus and Cyprinus carpio was exposed to sub lethal concentrations viz., 6.15 and 6.60 mg/l of vat blue 4 and vat green 1 respectively for a period of thirty days. The cumulative effect of these two dyes on the total carbohydrate, protein and lipid levels in the muscle of both the fishes were assayed on 7th, 18th and 30th day of exposure. The mean carbohydrate content of C. punctatus (both control and treated fish) collected on 18th and 30th day showed a significant difference (P < 0.01 to 0.04). In the case of control and treated C. carpio the data collected on 30th day showed significant variation (P < 0.04) in the carbohydrate content of the muscle. Student’s ‘t’ test analysis conducted on the protein content of C. punctatus during different occasions registered a significant decrease; in case of lipid content, except between the 1st and 30th day in all other cases the ‘t’ value is not significant. In case of C. carp protein and lipid content were observed to increase during the 7th day and thereafter a marked decrease was recorded on the 18th and 30th day of exposure. When the data was subjected to student’s ‘t’ test, both in protein and lipid, significant difference could not be recorded.

How Long Is A Piece Of Loop?, Yoonjoo Choi, Sumeet Agarwal, Charlotte M. Deane

Dartmouth Scholarship

Loops are irregular structures which connect two secondary structure elements in proteins. They often play important roles in function, including enzyme reactions and ligand binding. Despite their importance, their structure remains difficult to predict. Most protein loop structure prediction methods sample local loop segments and score them. In particular protein loop classifications and database search methods depend heavily on local properties of loops. Here we examine the distance between a loop's end points (span). We find that the distribution of loop span appears to be independent of the number of residues in the loop, in other words the separation between …

Ring Domain, David J. Hall

Protein Domains

Ring domain #1CHC. The RING finger is a specialized type of Zn finger consisting of 40–60 residues that binds two atoms of zinc, and is involved in mediating protein—protein interactions. Many zinc fingers bind nucleic acids. The presence of a RING finger domain is a characteristic of RING-class E3 ubiquitin protein ligases capable of transferring ubiquitin from an E2 enzyme to a substrate protein.

Sh2 Domain, David J. Hall

Protein Domains

SH2 domain #1BFJ. Src-homology 2 (SH2) domains are modules of ~100 amino acids that bind to specific phospho tyrosine (pY) containing peptide motifs. Conventional SH2 domains have a conserved pocket that recognizes pY, and a more variable pocket that binds 3-6 residues C-terminal to the pY and confers specificity.

Sh3 Domain, David J. Hall

Protein Domains

SH3 domain #1NEB. Src-homology 3 (SH3) domains bind to Pro-rich peptides that form a left-handed poly-Pro type II helix, with the minimal consensus Pro-X-X-Pro. Each Pro is usually preceeded by an aliphatic residue. Each in the aliphatic-Pro pair binds to a hydrophobic pocket on the SH3 domain.

Ig Domain, David J. Hall

Protein Domains

Ig domain #2CKN. This particular domain is named for the first protein in which it was found, the immunoglobulin. An immunoglobulin is a antibody. Antibodies are generated by our immune system to recognize the specific size, shape and charge of pathogens. This domain is also found on the extracellular portion of many receptors including the interleukin-1 family of receptors.

Beta Barrel, David J. Hall

Protein Domains

Beta barrel (cyan fluorescent protein) #4AR7. This fluorescent protein is a variation of green fluorescent protein from a jellyfish and is the only domain that is a complete protein. The protein is routinely used to visualize a variety of biological processes. The beta barrel domain is a beta sheet wrapped around the fluorescent active site to provide structure.

Helix Turn Helix Domain, David J. Hall

Protein Domains

Helix turn helix domain #3V1A. The helix-turn helix is a DNA-binding domain. The two alpha helices are the reading or recognition helices, which bind in a groove in the DNA and recognize specific gene regulatory sequences in the DNA.

Biochemical, Structural, And Drug Design Studies Of Multi-Drug Resistant Hiv-1 Therapeutic Targets, Tamaria Grace Dewdney

Wayne State University Dissertations

Protein point mutations acquired as a mechanism of survival against therapeutics cause structural changes that effect protein function and inhibitor binding. This work investigates the structural mechanisms that lead to multi-drug resistance to HIV-1 protease and integrase inhibitors.

Proper proteolytic processing of the HIV-1 Gag/Pol polyprotein is required for HIV infection and viral replication. This feature has made HIV-1 protease an attractive target for antiretroviral drug design for the treatment of HIV-1 infected patients, thus the development of drug resistance has arisen as a major therapeutic and drug design challenge. To understand the molecular mechanisms leading to drug resistance we …

Dimerization And Heme Binding Are Conserved In Amphibian And Starfish Homologues Of The Microrna Processing Protein Dgcr8., Rachel Senturia, Arthur Laganowsky, Ian Barr, Brooke D. Scheidemantle, Feng Guo

Natural Sciences and Mathematics | Faculty Scholarship

Human DiGeorge Critical Region 8 (DGCR8) is an essential microRNA (miRNA) processing factor that is activated via direct interaction with Fe(III) heme. In order for DGCR8 to bind heme, it must dimerize using a dimerization domain embedded within its heme-binding domain (HBD). We previously reported a crystal structure of the dimerization domain from human DGCR8, which demonstrated how dimerization results in the formation of a surface important for association with heme. Here, in an attempt to crystallize the HBD, we search for DGCR8 homologues and show that DGCR8 from Patiria miniata (bat star) also binds heme. The extinction coefficients (ε) …

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 …

In-Cell And In Vitro Studies Of Disease Related Protein-Protein Interactions Using Nmr-Spectroscopy, Andres Yudiel Maldonado

Legacy Theses & Dissertations (2009 - 2024)

The receptor for advanced glycation end products (RAGE) is a multiligand cell surface macromolecule that plays a central role in the etiology of diabetes, inflammation, and neurodegeneration. The cytoplasmic domain of RAGE, ctRAGE, is critical for RAGE-dependent signal transduction. As the most membrane proximal event, mDia1 binds to ctRAGE and is essential for RAGE ligand-stimulated phosphorylation of AKT and cell proliferation/migration. We show that ctRAGE contains an unusual alpha-turn that mediates the mDia1-ctRAGE interaction and is required for RAGE dependent signaling. The results establish a novel mechanism through which an extracellular signal initiated by RAGE ligands regulates RAGE signaling in …

Biophysical Investigations Of The Molecular Basis Of Cataract Associated With The R76s Mutation In Human Gammad-Crystallin, Vurghun Ahmadov

Legacy Theses & Dissertations (2009 - 2024)

Cataract disease results when the eye lens becomes opaque and scatters a significant part of the incoming light into the eye. The lens contains very high concentrations of the lens proteins, called crystallins, which are present at concentrations comparable to those found in protein crystals (about 400-600 mg/mL). Chemical modifications of the crystallins, such as oxidation and deamidation, or genetic mutations are known to result in increased light-scattering in vitro, and are implicated in cataract formation in vivo. Here we present the in vitro work on a mutant protein of human gammaD crystallin (HGD), namely R76S (i.e. Arg 76 to …

Mimosa: A System For Minimotif Annotation, Jay Vyas, Ronald J. Nowling, Thomas Meusburger, David P. Sargeant, Krishna Kadaveru, Michael R. Gryk, Vamsi Kundeti, Sanguthevar Rajasekaran, Martin Schiller

Life Sciences Faculty Research

BACKGROUND:

Minimotifs are short peptide sequences within one protein, which are recognized by other proteins or molecules. While there are now several minimotif databases, they are incomplete. There are reports of many minimotifs in the primary literature, which have yet to be annotated, while entirely novel minimotifs continue to be published on a weekly basis. Our recently proposed function and sequence syntax for minimotifs enables us to build a general tool that will facilitate structured annotation and management of minimotif data from the biomedical literature.

RESULTS:

We have built the MimoSA application for minimotif annotation. The application supports management of …

Elucidating The Structure Of Protein Aggregates By Raman Spectroscopy, Ludmila A. Popova

Legacy Theses & Dissertations (2009 - 2024)

The structures and properties of amyloid fibrils are of considerable interest due to their associations with numerous neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease and transmissible spongiform encephalopaties (prion diseases). Understanding fibrillogenesis at a molecular level requires detailed structural characterization of amyloid fibrils. However amyloid fibrils are difficult objects to study due to their non-crystalline and insoluble nature. These properties make the application of classical tools of structural biology, such as X-Ray crystallography and solution Nuclear Magnetic Resonance spectroscopy, impractical for structural characterization of protein fibrils.

Light-Regulated Sampling Of Protein Kinase Activity (Ii), Collaborative Project

Dyson College- Seidenberg School of CSIS : Collaborative Projects and Presentations

This entry adheres to the use of the quad chart template to provide a succinct description only of the current research project undertaken by the participants. It provides for the following information

1. Participants and Affiliations
2. Overall Project Goals
3. Illustrative picture
4. Specific research/artistic/pedagogic foci

The Stator Complex Of The A1a0-Atp Synthase--Structural Characterization Of The E And H Subunits., Erik Kish-Trier, Lee-Ann K Briere, Stanley D Dunn, Stephan Wilkens

Biochemistry Publications

Archaeal ATP synthase (A-ATPase) is the functional homolog to the ATP synthase found in bacteria, mitochondria and chloroplasts, but the enzyme is structurally more related to the proton-pumping vacuolar ATPase found in the endomembrane system of eukaryotes. We have cloned, overexpressed and characterized the stator-forming subunits E and H of the A-ATPase from the thermoacidophilic Archaeon, Thermoplasma acidophilum. Size exclusion chromatography, CD, matrix-assisted laser desorption ionization time-of-flight mass spectrometry and NMR spectroscopic experiments indicate that both polypeptides have a tendency to form dimers and higher oligomers in solution. However, when expressed together or reconstituted, the two individual polypeptides interact with …

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 …