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Structural Determinants For The Interactions Of Chemically Modified Nucleic Acids With The Stabilin‑2 Clearance Receptor, Hans Gaus, Colton M. Miller, Punit P. Seth, Edward N. Harris 2018 Ionis Pharmaceuticals, Carlsbad, California

Structural Determinants For The Interactions Of Chemically Modified Nucleic Acids With The Stabilin‑2 Clearance Receptor, Hans Gaus, Colton M. Miller, Punit P. Seth, Edward N. Harris

Biochemistry -- Faculty Publications

The Stabilin receptors are systemic clearance receptors for some classes of chemically modified nucleic acid therapeutics. In this study, the recombinant human secreted ecto-domain of the small isoform of Stabilin-2 (s190) was purified from cell culture and evaluated for direct binding with a multitude of antisense oligonucleotides (ASOs) using a fluorescence polarizationbased assay. The tested ASOs varied in their backbone composition, modification of the ribose 2′ position, overall length of the oligo, and sequence of the nucleotide bases. A fully phosphorothioate (PS) ASO with a 5−10−5 pattern of flanking 2′-O-methoxyethyl modifications was then used to test ...


The Role Of The Metallochaperone Hypa In The Acid Survival And Activities Of Nickel Enzymes In Helicobacter Pylori, Heidi Hu 2018 University of Massachusetts Amherst

The Role Of The Metallochaperone Hypa In The Acid Survival And Activities Of Nickel Enzymes In Helicobacter Pylori, Heidi Hu

Doctoral Dissertations

Helicobacter pylori is a bacterium that has colonized the human gastric mucosa of over 50% of the world population. Persistent infection can cause gastritis, peptic ulcers, and cancers. The ability of H. pylori to colonize the acidic environment of the human stomach is dependent on the activity of the nickel containing enzymes, urease and NiFe-hydrogenase. The nickel metallochaperone, HypA, was previously shown to be required for the full activity of both enzymes. In addition to a Ni-binding site, HypA also contains a structural Zn site, which has been characterized to alter its averaged structure depending on pH and the presence ...


Curcumin Potentiates The Function Of Human Α7-Nicotinic Acetylcholine Receptors Expressed In Sh-Ep1 Cells, Eslam El Nebrisi, Lina T. Al Kury, Keun-Hang Susan Yang, Petrilla Jayaprakash, Frank Christopher Howarth, Nadine Kabbani, Murat Oz 2018 UAE University

Curcumin Potentiates The Function Of Human Α7-Nicotinic Acetylcholine Receptors Expressed In Sh-Ep1 Cells, Eslam El Nebrisi, Lina T. Al Kury, Keun-Hang Susan Yang, Petrilla Jayaprakash, Frank Christopher Howarth, Nadine Kabbani, Murat Oz

Mathematics, Physics, and Computer Science Faculty Articles and Research

Effects of curcumin, a biologically active ingredient of turmeric, were tested on the Ca2+transients induced by the activation of α7 subunit of the human nicotinic acetylcholine (α7nACh) receptor expressed in SH-EP1 cells. Curcumin caused a significant potentiation of choline (1 mM)-induced Ca2+ transients with an EC50 value of 133 nM. The potentiating effect of curcumin was not observed in Ca2+ transients induced by high K+ (60 mM) containing solutions or activation of α4β2 nACh receptors and the extent of curcumin potentiation was not altered in the presence of ...


Mutations In Rhodopsin Transmembrane Helix 5 And Dimerization Interface, Margaret Pinkevitch 2018 The University of Akron

Mutations In Rhodopsin Transmembrane Helix 5 And Dimerization Interface, Margaret Pinkevitch

Williams Honors College, Honors Research Projects

G protein coupled receptors (GPCRs) are membrane proteins with many functional roles. This family of receptors is expansive, with several subfamilies. Here, a member of subfamily A, rhodopsin (Rho) is studied. Mutations in rhodopsin have been linked to the disease retinitis pigmentosa (RP). Some of these mutations have been found to disrupt the dimerization/oligomerization of Rho at the TM4/TM5 proposed dimerization interface, causing pathogenesis of RP. In this study, two mutations of transmembrane helix 5 were investigated for changes in dimerization and stability, cell trafficking, and signaling. It was found that the V209M and F220C mutations did not ...


Improving The Detection Limit Of Tau Aggregates For Use With Biological Samples, Emily Rickman Hager 2018 University of Denver

Improving The Detection Limit Of Tau Aggregates For Use With Biological Samples, Emily Rickman Hager

Electronic Theses and Dissertations

The protein Tau is found in neurofibrillary tangles in Alzheimer's disease and over 20 other neurodegenerative diseases. An assay has been developed to detect minute amounts of fibrils from human brain tissue. This assay subjects brain tissue extract and recombinant Tau to several rounds of sonication and incubation. Incubation allows recombinant Tau to add itself to the ends of the existing fibrils in brain tissue extract. Sonication breaks the existing fibrils in the brain tissue extract offering more ends for Tau to add onto. Cycles of sonication and incubation have been shown to allow for amplification of Tau fibrils ...


Structural Features And Domain Movements Controlling Substrate Binding And Cofactor Specificity In Class Ii Hmg-Coa Reductase, Bradley R. Miller, Yan Kung 2018 Bryn Mawr College

Structural Features And Domain Movements Controlling Substrate Binding And Cofactor Specificity In Class Ii Hmg-Coa Reductase, Bradley R. Miller, Yan Kung

Chemistry Faculty Research and Scholarship

The key mevalonate pathway enzyme 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase (HMGR) uses the cofactor NAD(P)H to reduce HMG-CoA to mevalonate in the production of countless metabolites and natural products. Although inhibition of HMGR by statin drugs is well-understood, several mechanistic details of HMGR catalysis remain unresolved, and the structural basis for the wide range of cofactor specificity for either NADH or NADPH among HMGRs from different organisms is also unknown. Here, we present crystal structures of HMGR from Streptococcus pneumoniae (SpHMGR) alongside kinetic data of the enzyme’s cofactor preferences. Our structure of SpHMGR bound with its kinetically ...


Recently Discovered Benzo(A)Pyrene Induced Health Effects And Their Threat To The Inland Empire, Quinn Navarro 2018 University of Redlands

Recently Discovered Benzo(A)Pyrene Induced Health Effects And Their Threat To The Inland Empire, Quinn Navarro

Armacost Library Undergraduate Research Award (ALURA)

This study investigated recent discoveries of Benzo(a)Pyrene (BaP) in order to determine the current threat BaP poses to residents of Southern California. Literature reviews comprise the research done on BaP-induced health effects. Primary articles reviewed include “Adverse Effect of Sub-Chronic Exposure to Benzo(a)Pyrene and Protective Effect of Butylated Hydroxyanisole on Learning and Memory Ability in Male

Sprague-Dawley Rat” by Xiao Liang (2014), and “Bioaccumulation and oxidative damage in juvenile scallop Chlamys farreri exposed to benzo[a]Pyrene, benzo[b]Fluoranthene and chrysene” by Meng Xiu (2014). These studies conclude that BaP potentially impairs memory and learning ...


Reactive Oxygen Species In Chronic Obstructive Pulmonary Disease, Samia Boukhenouna, Mark A. Wilson, Karim Bahmed, Beata Kosmider 2018 Temple University

Reactive Oxygen Species In Chronic Obstructive Pulmonary Disease, Samia Boukhenouna, Mark A. Wilson, Karim Bahmed, Beata Kosmider

Biochemistry -- Faculty Publications

Chronic obstructive pulmonary disease (COPD) includes chronic bronchitis and emphysema. Environmental exposure, primarily cigarette smoking, can cause high oxidative stress and is the main factor of COPD development. Cigarette smoke also contributes to the imbalance of oxidant/antioxidant due to exogenous reactive oxygen species (ROS). Moreover, endogenously released ROS during the inflammatory process and mitochondrial dysfunction may contribute to this disease progression. ROS and reactive nitrogen species (RNS) can oxidize different biomolecules such as DNA, proteins, and lipids leading to epithelial cell injury and death. Various detoxifying enzymes and antioxidant defense systems can be involved in ROS removal. In this ...


Impact Of Phosphoinositides On Regulation Of K-Atp By Atp And Hydrogen Sulfide, Tyler Hendon 2018 Virginia Commonwealth University

Impact Of Phosphoinositides On Regulation Of K-Atp By Atp And Hydrogen Sulfide, Tyler Hendon

Theses and Dissertations

Hydrogen sulfide (H2S) reduces ischemia reperfusion (IR) injury by stimulating adenosine triphosphate (ATP) sensitive potassium channels (KATP) [1-5]. Demonstrating H2S stimulation is unique to KATP, as other inwardly rectifying potassium (Kir) channels demonstrate inhibition or are unaffected [6]. We recently showed that H2S inhibits Kir2 and Kir3 by decreasing channel sensitivity to phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2 or PIP2) [6]. Here, we test the hypothesis that H2S regulation of Kir6.2, a pore-forming subunit of the KATP channel, is also dependent on PIP2. Using whole-cell patch-clamp we show that H2S increases the activity of Kir6.2 channels expressed ...


Rates And Patterns Of Evolution In A Duplicated Genome In The Family Catostomidae, Megann Michelle Schmidt 2018 University of Northern Iowa

Rates And Patterns Of Evolution In A Duplicated Genome In The Family Catostomidae, Megann Michelle Schmidt

Honors Program Theses

Whole genome duplication (WGD) is a process in which the entire genome of an organism is duplicated, making redundant genes which are subject to unique evolutionary forces. Various modes of selection create different genetic fates such as retention of ancestral function, development of new function, or loss of function. Because of these differing fates, WGD is hypothesized to be a major driving force behind diversification. In this project, DNA sequences from fish species in the family Catostomidae were examined to observe patterns of evolution following a known WGD. Gene trees were generated for 179 loci to determine the amount of ...


Total Mercury Determination In Muscle And Liver Tissue Samples From Brazilian Amazon Fish Using Slurry Sampling, João Vitor de Queiroz, José Cavalcante Souza Vieira, Izabela da Cunha Bataglioli, Alis Correia Bittarello, Camila Pereira Braga, Grasieli de Oliveira, Cilene do Carmo Federici Padilha, Pedro de Magalhães Padilha 2018 São Paulo State University

Total Mercury Determination In Muscle And Liver Tissue Samples From Brazilian Amazon Fish Using Slurry Sampling, João Vitor De Queiroz, José Cavalcante Souza Vieira, Izabela Da Cunha Bataglioli, Alis Correia Bittarello, Camila Pereira Braga, Grasieli De Oliveira, Cilene Do Carmo Federici Padilha, Pedro De Magalhães Padilha

Biochemistry -- Faculty Publications

This paper presents a slurry sampling method for total mercury determination by graphite furnace atomic absorption spectrometry (GFAAS) in tissue of fish from the Amazon. The tissue samples were lyophilized and macerated, and then the slurry samples were prepared by putting 20 mg of tissue, added to a solution containing Triton X-100, Suprapur HNO3, and zirconium nitrate directly in sampling vials of a spectrometer. Mercury standard solutions were prepared under the same conditions as the slurry samples. The slurry samples and the mercury standard solutions were sonicated for 20 s. Twenty microliters of slurry samples were injected into the ...


Cancerdiscover: An Integrative Pipeline For Cancer Biomarker And Cancer Class Prediction From High-Throughput Sequencing Data, Akram Mohammed, Greyson Biegert, Jiri Adamec, Tomáš Helikar 2018 University of Nebraska-Lincoln

Cancerdiscover: An Integrative Pipeline For Cancer Biomarker And Cancer Class Prediction From High-Throughput Sequencing Data, Akram Mohammed, Greyson Biegert, Jiri Adamec, Tomáš Helikar

Biochemistry -- Faculty Publications

Accurate identification of cancer biomarkers and classification of cancer type and subtype from High Throughput Sequencing (HTS) data is a challenging problem because it requires manual processing of raw HTS data from various sequencing platforms, quality control, and normalization, which are both tedious and timeconsuming. Machine learning techniques for cancer class prediction and biomarker discovery can hasten cancer detection and significantly improve prognosis. To date, great research efforts have been taken for cancer biomarker identification and cancer class prediction. However, currently available tools and pipelines lack flexibility in data preprocessing, running multiple feature selection methods and learning algorithms, therefore, developing ...


The Colomoto Interactive Notebook: Accessible And Reproducible Computational Analyses For Qualitative Biological Networks, Aurelien Naldi, Celine Hernandez, Nicolas Levy, Gautier Stoll, Pedro T. Monteiro, Claudine Chaouiya, Tomáš Helikar, Andrei Zinovyev, Laurence Calzone, Sarah Cohen-Boulakia, Denis Thieffry, Loic Pauleve 2018 PSL Université

The Colomoto Interactive Notebook: Accessible And Reproducible Computational Analyses For Qualitative Biological Networks, Aurelien Naldi, Celine Hernandez, Nicolas Levy, Gautier Stoll, Pedro T. Monteiro, Claudine Chaouiya, Tomáš Helikar, Andrei Zinovyev, Laurence Calzone, Sarah Cohen-Boulakia, Denis Thieffry, Loic Pauleve

Biochemistry -- Faculty Publications

Analysing models of biological networks typically relies on workflows in which different software tools with sensitive parameters are chained together,many times with additional manual steps. The accessibility and reproducibility of such workflows is challenging, as publications often overlook analysis details, and because some of these tools may be difficult to install, and/or have a steep learning curve. The CoLoMoTo Interactive Notebook provides a unified environment to edit, execute, share, and reproduce analyses of qualitative models of biological networks. This framework combines the power of different technologies to ensure repeatability and to reduce users’ learning curve of these technologies ...


Simulation Of Stimulation: Cytokine Dosage And Cell Cycle Crosstalk Driving Timing-Dependent T Cell Differentiation, Matteo Barberis, Tomáš Helikar, Paul Verbruggen 2018 University of Amsterdam

Simulation Of Stimulation: Cytokine Dosage And Cell Cycle Crosstalk Driving Timing-Dependent T Cell Differentiation, Matteo Barberis, Tomáš Helikar, Paul Verbruggen

Biochemistry -- Faculty Publications

Triggering an appropriate protective response against invading agents is crucial to the effectiveness of human innate and adaptive immunity. Pathogen recognition and elimination requires integration of a myriad of signals from many different immune cells. For example, T cell functioning is not qualitatively, but quantitatively determined by cellular and humoral signals. Tipping the balance of signals, such that one of these is favored or gains advantage on another one, may impact the plasticity of T cells. This may lead to switching their phenotypes and, ultimately, modulating the balance between proliferating and memory T cells to sustain an appropriate immune response ...


Towards The Synthetic Design Of Camelina Oil Enriched In Tailored Acetyl-Triacylglycerols With Medium-Chain Fatty Acids, Sunil Bansal, Hae Jin Kim, GunNam Na, Megan E. Hamilton, Edgar B. Cahoon, Chaofu Lu, Timothy P. Durrett 2018 Lovely Professional University

Towards The Synthetic Design Of Camelina Oil Enriched In Tailored Acetyl-Triacylglycerols With Medium-Chain Fatty Acids, Sunil Bansal, Hae Jin Kim, Gunnam Na, Megan E. Hamilton, Edgar B. Cahoon, Chaofu Lu, Timothy P. Durrett

Biochemistry -- Faculty Publications

The ability to manipulate expression of key biosynthetic enzymes has allowed the development of genetically modified plants that synthesise unusual lipids that are useful for biofuel and industrial applications. By taking advantage of the unique activities of enzymes from different species, tailored lipids with a targeted structure can be conceived. In this study we demonstrate the successful implementation of such an approach by metabolically engineering the oilseed crop Camelina sativa to produce 3-acetyl-1,2-diacyl-sn-glycerols (acetyl-TAGs) with medium-chain fatty acids (MCFAs). Different transgenic camelina lines that had been genetically modified to produce MCFAs through the expression of MCFA-specific thioesterases ...


A Mechanistic Computational Model Reveals That Plasticity Of Cd4+ T Cell Differentiation Is A Function Of Cytokine Composition And Dosage, Bhanwar Lal Puniya, Robert G. Todd, Akram Mohammed, Deborah M. Brown, Matteo Barberis, Tomáš Helikar 2018 University of Nebraska-Lincoln

A Mechanistic Computational Model Reveals That Plasticity Of Cd4+ T Cell Differentiation Is A Function Of Cytokine Composition And Dosage, Bhanwar Lal Puniya, Robert G. Todd, Akram Mohammed, Deborah M. Brown, Matteo Barberis, Tomáš Helikar

Biochemistry -- Faculty Publications

CD4+ T cells provide cell-mediated immunity in response to various antigens. During an immune response, naïve CD4+ T cells differentiate into specialized effector T helper (Th1, Th2, and Th17) cells and induced regulatory (iTreg) cells based on a cytokine milieu. In recent studies, complex phenotypes resembling more than one classical T cell lineage have been experimentally observed. Herein, we sought to characterize the capacity of T cell differentiation in response to the complex extracellular environment. We constructed a comprehensive mechanistic (logical) computational model of the signal transduction that regulates T cell differentiation. The model’s dynamics were characterized and analyzed ...


Computational Systems Biology Approach For The Study Of Rheumatoid Arthritis: From A Molecular Map To A Dynamical Model, Vidisha Singh, Marek Ostaszewski, George D. Kalliolias, Gilles Chiocchia, Robert Olaso, Elisabeth Petit-Teixeira, Tomáš Helikar, Anna Niarakis 2018 Universite Paris-Saclay

Computational Systems Biology Approach For The Study Of Rheumatoid Arthritis: From A Molecular Map To A Dynamical Model, Vidisha Singh, Marek Ostaszewski, George D. Kalliolias, Gilles Chiocchia, Robert Olaso, Elisabeth Petit-Teixeira, Tomáš Helikar, Anna Niarakis

Biochemistry -- Faculty Publications

In this work we present a systematic effort to summarize current biological pathway knowledge concerning Rheumatoid Arthritis (RA). We are constructing a detailed molecular map based on exhaustive literature scanning, strict curation criteria, re-evaluation of previously published attempts and most importantly experts’ advice. The RA map will be web-published in the coming months in the form of an interactive map, using the MINERVA platform, allowing for easy access, navigation and search of all molecular pathways implicated in RA, serving thus, as an on line knowledgebase for the disease. Moreover the map could be used as a template for Omics data ...


Single-Molecule Unbinding Forces Between The Polysaccharide Hyaluronan And Its Binding Proteins, Fouzia Bano, Markku I. Tammi, David W. Kang, Edward N. Harris, Ralf P. Richter 2018 University of Leeds

Single-Molecule Unbinding Forces Between The Polysaccharide Hyaluronan And Its Binding Proteins, Fouzia Bano, Markku I. Tammi, David W. Kang, Edward N. Harris, Ralf P. Richter

Biochemistry -- Faculty Publications

The extracellular polysaccharide hyaluronan (HA) is ubiquitous in all vertebrate tissues, where its various functions are encoded in the supramolecular complexes and matrices that it forms with HA-binding proteins (hyaladherins). In tissues, these supramolecular architectures are frequently subjected to mechanical stress, yet how this affects the intermolecular bonding is largely unknown. Here, we used a recently developed single-molecule force spectroscopy platform to analyze and compare the mechanical strength of bonds between HA and a panel of hyaladherins from the Link module superfamily, namely the complex of the proteoglycan aggrecan and cartilage link protein, the proteoglycan versican, the inflammation-associated protein TSG-6 ...


Purification Of Hepatocytes And Sinusoidal Endothelial Cells From Mouse Liver Perfusion, Fatima Cabral, Colton M. Miller, Katrina M. Kudrna, Blake E. Hass, Jocelyn G. Daubendiek, Brianna M. Kellar, Edward N. Harris 2018 University of Nebraska - Lincoln

Purification Of Hepatocytes And Sinusoidal Endothelial Cells From Mouse Liver Perfusion, Fatima Cabral, Colton M. Miller, Katrina M. Kudrna, Blake E. Hass, Jocelyn G. Daubendiek, Brianna M. Kellar, Edward N. Harris

Biochemistry -- Faculty Publications

This protocol demonstrates a method for obtaining high yield and viability for mouse hepatocytes and sinusoidal endothelial cells (SECs) suitable for culturing or for obtaining cell lysates. In this protocol, the portal vein is used as the site for catheterization, rather than the vena cava, as this limits contamination of other possible cell types in the final liver preparation. No special instrumentation is required throughout the procedure. A water bath is used as a source of heat to maintain the temperature of all the buffers and solutions. A standard peristaltic pump is used to drive the fluid, and a refrigerated ...


Endosomal Escape Of Antisense Oligonucleotides Internalized By Stabilin Receptors Is Regulated By Rab5c And Eea1 During Endosomal Maturation, Colton M. Miller, W. Brad Wan, Punit P. Seth, Edward N. Harris 2018 University of Nebraska - Lincoln

Endosomal Escape Of Antisense Oligonucleotides Internalized By Stabilin Receptors Is Regulated By Rab5c And Eea1 During Endosomal Maturation, Colton M. Miller, W. Brad Wan, Punit P. Seth, Edward N. Harris

Biochemistry -- Faculty Publications

Second-generation (Gen 2) Antisense oligonucleotides (ASOs) show increased nuclease stability and affinity for their RNA targets, which has translated to improved potency and therapeutic index in the clinic. Gen 2 ASOs are typically modified using the phosphorothioate (PS) backbone modification, which enhances ASO interactions with plasma, cell surface, and intracellular proteins. This facilitates ASO distribution to peripheral tissues and also promotes cellular uptake after injection into animals. Previous work identified that Stabilin receptors specifically internalize PS-ASOs in the sinusoidal endothelial cells of the liver and the spleen. By modulating expression of specific proteins involved in the trafficking and maturation of ...


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