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2014

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Articles 1 - 17 of 17

Full-Text Articles in Nucleic Acids, Nucleotides, and Nucleosides

Effect Of Hydrogen Peroxide On The Biosynthesis Of Heme And Proteins: Potential Implications For The Partitioning Of Glu-TrnaGlu Between These Pathways, Carolina Farah, Gloria Levicán, Michael Ibba, Omar Orellana Dec 2014

Effect Of Hydrogen Peroxide On The Biosynthesis Of Heme And Proteins: Potential Implications For The Partitioning Of Glu-TrnaGlu Between These Pathways, Carolina Farah, Gloria Levicán, Michael Ibba, Omar Orellana

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Glutamyl-tRNA (Glu-tRNAGlu) is the common substrate for both protein translation and heme biosynthesis via the C5 pathway. Under normal conditions, an adequate supply of this aminoacyl-tRNA is available to both pathways. However, under certain circumstances, Glu-tRNAGlu can become scarce, resulting in competition between the two pathways for this aminoacyl-tRNA. In Acidithiobacillus ferrooxidans, glutamyl-tRNA synthetase 1 (GluRS1) is the main enzyme that synthesizes Glu-tRNAGlu. Previous studies have shown that GluRS1 is inactivated in vitro by hydrogen peroxide (H2O2). This raises the question as to whether H2O2 negatively affects …


Mrp4-Dependent Regulation Of Fibroblast Migration, Chandrima Sinha Dec 2014

Mrp4-Dependent Regulation Of Fibroblast Migration, Chandrima Sinha

Theses and Dissertations (ETD)

Roles of cyclic nucleotides and cyclic nucleotide-dependent signaling molecules in regulating several signaling pathways including cell migration have long been known. However, the new and revolutionary concept is that it is not just the absence or presence of cyclic nucleotides, but a highly coordinated balance between these molecules regulates cell migration. Multi-drug resistance protein 4 (MRP4), is a member of the large family of ATP binding cassette (ABC) transporter proteins, that localizes to the plasma membrane and functions as a nucleotide efflux transporter and thus plays a pivotal role in the regulation of intracellular cyclic nucleotide dynamics. In our study …


Nucleoside Reverse Transcriptase Inhibitors Possess Intrinsic Anti-Inflammatory Activity, Benjamin J. Fowler, Bradley D. Gelfand, Younghee Kim, Nagaraj Kerur, Valeria Tarallo, Yoshio Hirano, Shoba Amarnath, Daniel H. Fowler, Marta Radwan, Mark T. Young, Keir Pittman, Paul Kubes, Hitesh Agarwal, Keykavous Parang, David R. Hinton, Ana Bastos-Carvalho, Shengjian Li, Testuhiro Yasuma, Takeshi Mizutani, Reo Yasuma, Charles Wright, Jayakrishna Ambati Nov 2014

Nucleoside Reverse Transcriptase Inhibitors Possess Intrinsic Anti-Inflammatory Activity, Benjamin J. Fowler, Bradley D. Gelfand, Younghee Kim, Nagaraj Kerur, Valeria Tarallo, Yoshio Hirano, Shoba Amarnath, Daniel H. Fowler, Marta Radwan, Mark T. Young, Keir Pittman, Paul Kubes, Hitesh Agarwal, Keykavous Parang, David R. Hinton, Ana Bastos-Carvalho, Shengjian Li, Testuhiro Yasuma, Takeshi Mizutani, Reo Yasuma, Charles Wright, Jayakrishna Ambati

Pharmacy Faculty Articles and Research

Nucleoside reverse transcriptase inhibitors (NRTIs) are mainstay therapeutics for HIV that block retrovirus replication. Alu (an endogenous retroelement that also requires reverse transcriptase for its life cycle)–derived RNAs activate P2X7 and the NLRP3 inflammasome to cause cell death of the retinal pigment epithelium in geographic atrophy, a type of age-related macular degeneration. We found that NRTIs inhibit P2X7-mediated NLRP3 inflammasome activation independent of reverse transcriptase inhibition. Multiple approved and clinically relevant NRTIs prevented caspase-1 activation, the effector of the NLRP3 inflammasome, induced by Alu RNA. NRTIs were efficacious in mouse models of geographic atrophy, choroidal neovascularization, graft-versus-host disease, and sterile …


The Non-Canonical Hydroxylase Structure Of Yfcm Reveals A Metal Ion-Coordination Motif Required For Ef-P Hydroxylation, Kan Kobayashi, Assaf Katz, Andrei Rajkovic, Ryohei Ishii, Owen E. Branson, Michael A. Freitas, Ryuichiro Ishitani, Michael Ibba, Osamu Nureki Oct 2014

The Non-Canonical Hydroxylase Structure Of Yfcm Reveals A Metal Ion-Coordination Motif Required For Ef-P Hydroxylation, Kan Kobayashi, Assaf Katz, Andrei Rajkovic, Ryohei Ishii, Owen E. Branson, Michael A. Freitas, Ryuichiro Ishitani, Michael Ibba, Osamu Nureki

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

EF-P is a bacterial tRNA-mimic protein, which accelerates the ribosome-catalyzed polymerization of poly-prolines. In Escherichia coli, EF-P is post-translationally modified on a conserved lysine residue. The post-translational modification is performed in a two-step reaction involving the addition of a β-lysine moiety and the subsequent hydroxylation, catalyzed by PoxA and YfcM, respectively. The β-lysine moiety was previously shown to enhance the rate of poly-proline synthesis, but the role of the hydroxylation is poorly understood. We solved the crystal structure of YfcM and performed functional analyses to determine the hydroxylation mechanism. In addition, YfcM appears to be structurally distinct from any …


Mistranslation Of The Genetic Code, Adil Moghal, Kyle Mohler, Michael Ibba Sep 2014

Mistranslation Of The Genetic Code, Adil Moghal, Kyle Mohler, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

During mRNA decoding at the ribosome, deviations from stringent codon identity, or “mistranslation,” are generally deleterious and infrequent. Observations of organisms that decode some codons ambiguously, and the discovery of a compensatory increase in mistranslation frequency to combat environmental stress have changed the way we view “errors” in decoding. Modern tools for the study of the frequency and phenotypic effects of mistranslation can provide quantitative and sensitive measurements of decoding errors that were previously inaccessible. Mistranslation with non‐protein amino acids, in particular, is an enticing prospect for new drug therapies and the study of molecular evolution.


Relaxed Substrate Specificity Leads To Extensive Trna Mischarging By Streptococcus Pneumoniae Class I And Class Ii Aminoacyl-Trna Synthetases, Jennifer Shepherd, Michael Ibba Sep 2014

Relaxed Substrate Specificity Leads To Extensive Trna Mischarging By Streptococcus Pneumoniae Class I And Class Ii Aminoacyl-Trna Synthetases, Jennifer Shepherd, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacyl-tRNA synthetases provide the first step in protein synthesis quality control by discriminating cognate from noncognate amino acid and tRNA substrates. While substrate specificity is enhanced in many instances by cis- and trans-editing pathways, it has been revealed that in organisms such as Streptococcus pneumoniae some aminoacyl-tRNA synthetases display significant tRNA mischarging activity. To investigate the extent of tRNA mischarging in this pathogen, the aminoacylation profiles of class I isoleucyl-tRNA synthetase (IleRS) and class II lysyl-tRNA synthetase (LysRS) were determined. Pneumococcal IleRS mischarged tRNAIle with both Val, as demonstrated in other bacteria, and Leu in a tRNA sequence-dependent …


Translation Initiation Rate Determines The Impact Of Ribosome Stalling On Bacterial Protein Synthesis, Steven J. Hersch, Sara Elgamal, Assaf Katz, Michael Ibba, William Wiley Navarre Aug 2014

Translation Initiation Rate Determines The Impact Of Ribosome Stalling On Bacterial Protein Synthesis, Steven J. Hersch, Sara Elgamal, Assaf Katz, Michael Ibba, William Wiley Navarre

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Ribosome stalling during translation can be caused by a number of characterized mechanisms. However, the impact of elongation stalls on protein levels is variable, and the reasons for this are often unclear. To investigate this relationship, we examined the bacterial translation elongation factor P (EF-P), which plays a critical role in rescuing ribosomes stalled at specific amino acid sequences including polyproline motifs. In previous proteomic analyses of both Salmonella and Escherichia coli efp mutants, it was evident that not all proteins containing a polyproline motif were dependent on EF-P for efficient expression in vivo . The α- and β-subunits of …


Ef-P Dependent Pauses Integrate Proximal And Distal Signals During Translation, Sara Elgamal, Assaf Katz, Steven J. Hersch, David Newsom, Peter White, William Wiley Navarre, Michael Ibba Aug 2014

Ef-P Dependent Pauses Integrate Proximal And Distal Signals During Translation, Sara Elgamal, Assaf Katz, Steven J. Hersch, David Newsom, Peter White, William Wiley Navarre, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Elongation factor P (EF-P) is required for the efficient synthesis of proteins with stretches of consecutive prolines and other motifs that would otherwise lead to ribosome pausing. However, previous reports also demonstrated that levels of most diprolyl-containing proteins are not altered by the deletion of efp. To define the particular sequences that trigger ribosome stalling at diprolyl (PPX) motifs, we used ribosome profiling to monitor global ribosome occupancy in Escherichia coli strains lacking EF-P. Only 2.8% of PPX motifs caused significant ribosomal pausing in the Δefp strain, with up to a 45-fold increase in ribosome density observed at …


A Novel Label-Free Colorimetric Assay For Dna Concentration In Solution, Molla R. Islam, Michael J. Serpe Jun 2014

A Novel Label-Free Colorimetric Assay For Dna Concentration In Solution, Molla R. Islam, Michael J. Serpe

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Optical devices were fabricated by sandwiching a “monolithic” poly(N-isopropylacrylamide-co-N-(3-aminopropyl) methacrylamide hydrochloride) (pNIPAm-co-APMAH) microgel layer between two semitransparent Au layers. These devices, referred to as etalons, exhibit characteristic multipeak reflectance spectra, and the position of the peaks in the spectra primarily depends on the distance between the Au surfaces mediated by the microgel layer thickness. Here, we show that the positively charged microgel layer can collapse in the presence of negatively charged single stranded DNA (ssDNA) due to ssDNA induced microgel crosslinking. The collapse results in a change in the etalon's optical properties, which can be used to detect target DNA …


Trnas As Regulators Of Biological Processes, Medha Raina, Michael Ibba Jun 2014

Trnas As Regulators Of Biological Processes, Medha Raina, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Transfer RNAs (tRNA) are best known for their role as adaptors during translation of the genetic code. Beyond their canonical role during protein biosynthesis, tRNAs also perform additional functions in both prokaryotes and eukaryotes for example in regulating gene expression. Aminoacylated tRNAs have also been implicated as substrates for non-ribosomal peptide bond formation, post-translational protein labeling, modification of phospholipids in the cell membrane, and antibiotic biosyntheses. Most recently tRNA fragments, or tRFs, have also been recognized to play regulatory roles. Here, we examine in more detail some of the new functions emerging for tRNA in a variety of cellular processes …


Oxidation Of Cellular Amino Acid Pools Leads To Cytotoxic Mistranslation Of The Genetic Code, Tammy J. Bullwinkle, Noah M. Reynolds, Medha Raina, Adil Moghal, Eleftheria Matsa, Andrei Rajkovic, Huseyin Kayadibi, Farbod Fazlollahi, Christopher Ryan, Nathaniel Howitz, Kym F. Faull, Beth A. Lazazzera, Michael Ibba Jun 2014

Oxidation Of Cellular Amino Acid Pools Leads To Cytotoxic Mistranslation Of The Genetic Code, Tammy J. Bullwinkle, Noah M. Reynolds, Medha Raina, Adil Moghal, Eleftheria Matsa, Andrei Rajkovic, Huseyin Kayadibi, Farbod Fazlollahi, Christopher Ryan, Nathaniel Howitz, Kym F. Faull, Beth A. Lazazzera, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacyl-tRNA synthetases use a variety of mechanisms to ensure fidelity of the genetic code and ultimately select the correct amino acids to be used in protein synthesis. The physiological necessity of these quality control mechanisms in different environments remains unclear, as the cost vs benefit of accurate protein synthesis is difficult to predict. We show that in Escherichia coli, a non-coded amino acid produced through oxidative damage is a significant threat to the accuracy of protein synthesis and must be cleared by phenylalanine-tRNA synthetase in order to prevent cellular toxicity caused by mis-synthesized proteins. These findings demonstrate how stress …


Reduced Amino Acid Specificity Of Mammalian Tyrosyl-Trna Synthetase Is Associated With Elevated Mistranslation Of Tyr Codons, Medha Raina, Adil Moghal, Amanda Kano, Mathew Jerums, Paul D. Schnier, Shun Luo, Rohini Deshpande, Pavel D. Bondarenko, Henry Lin, Michael Ibba May 2014

Reduced Amino Acid Specificity Of Mammalian Tyrosyl-Trna Synthetase Is Associated With Elevated Mistranslation Of Tyr Codons, Medha Raina, Adil Moghal, Amanda Kano, Mathew Jerums, Paul D. Schnier, Shun Luo, Rohini Deshpande, Pavel D. Bondarenko, Henry Lin, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Quality control operates at different steps in translation to limit errors to approximately one mistranslated codon per 10,000 codons during mRNA-directed protein synthesis. Recent studies have suggested that error rates may actually vary considerably during translation under different growth conditions. Here we examined the misincorporation of Phe at Tyr codons during synthesis of a recombinant antibody produced in tyrosine-limited Chinese hamster ovary (CHO) cells. Tyr to Phe replacements were previously found to occur throughout the antibody at a rate of up to 0.7% irrespective of the identity or context of the Tyr codon translated. Despite this comparatively high mistranslation rate, …


Structural And Biophysical Characterization Of Staphylococcus Aureus Samazf Shows Conservation Of Functional Dynamics, Valentina Zorzini, Lieven Buts, Mike Sleutel, Abel Garcia-Pino, Ariel Talavera, Sarah Haesaerts, Henri De Greve, Ambrose Cheung, Nico A. J. Van Nuland, Remy Loris Mar 2014

Structural And Biophysical Characterization Of Staphylococcus Aureus Samazf Shows Conservation Of Functional Dynamics, Valentina Zorzini, Lieven Buts, Mike Sleutel, Abel Garcia-Pino, Ariel Talavera, Sarah Haesaerts, Henri De Greve, Ambrose Cheung, Nico A. J. Van Nuland, Remy Loris

Dartmouth Scholarship

The Staphylococcus aureus genome contains three toxin-antitoxin modules, including one mazEF module, SamazEF. Using an on-column separation protocol we are able to obtain large amounts of wild-type SaMazF toxin. The protein is well-folded and highly resistant against thermal unfolding but aggregates at elevated temperatures. Crystallographic and nuclear magnetic resonance (NMR) solution studies show a well-defined dimer. Differences in structure and dynamics between the X-ray and NMR structural ensembles are found in three loop regions, two of which undergo motions that are of functional relevance. The same segments also show functionally relevant dynamics in the distantly related CcdB family despite divergence …


Design And Synthesis Of 4-N-Alkanoyl And 4-N-Alkyl Gemcitabine Analogues Suitable For Positron Emission Tomography, Jesse E. Pulido Mar 2014

Design And Synthesis Of 4-N-Alkanoyl And 4-N-Alkyl Gemcitabine Analogues Suitable For Positron Emission Tomography, Jesse E. Pulido

FIU Electronic Theses and Dissertations

Gemcitabine is a highly potent chemotherapeutic nucleoside agent used in the treatment of several cancers and solid tumors. However, it is therapeutically limitated because of toxicity to normal cells and its rapid intracellular deamination by cytidine deaminase into the inactive uracil derivative. Modification at the 4-(N) position of gemcitabine's exocyclic amine to an -amide functionality is a well reported prodrug strategy which has been that confers a resistance to intracellular deamination while also altering pharmacokinetics of the parent drug. Coupling of gemcitabine to carboxylic acids with varying terminal moieties afforded the 4-N-alkanoylgemcitabines whereas reaction of 4-N …


The Abcs Of The Ribosome, Kurt Fredrick, Michael Ibba Feb 2014

The Abcs Of The Ribosome, Kurt Fredrick, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

An ABC protein that binds the ribosomal exit site suggests a new mechanism for direct regulation of translation in response to changing ATP levels in the cell.


Perspectives On: Cyclic Nucleotide Microdomains And Signaling Specificity, Jeffrey W. Karpen Jan 2014

Perspectives On: Cyclic Nucleotide Microdomains And Signaling Specificity, Jeffrey W. Karpen

Touro College of Osteopathic Medicine (Middletown) Publications and Research

The author provides an introduction to a special issue on cyclic nucleotides concerning four areas in cAMP compartmentation.


The Control Of Lipid Metabolism By Mrna Splicing In Drosophila, R. M. Gingras, M. E. Warren, A. A. Nagengast, J. R. Diangelo Jan 2014

The Control Of Lipid Metabolism By Mrna Splicing In Drosophila, R. M. Gingras, M. E. Warren, A. A. Nagengast, J. R. Diangelo

Journal Articles

The storage of lipids is an evolutionarily conserved process that is important for the survival of organisms during shifts in nutrient availability. Triglycerides are stored in lipid droplets, but the mechanisms of how lipids are stored in these structures are poorly understood. Previous in vitro RNAi screens have implicated several components of the spliceosome in controlling lipid droplet formation and storage, but the in vivo relevance of these phenotypes is unclear. In this study, we identify specific members of the splicing machinery that are necessary for normal triglyceride storage in the Drosophila fat body. Decreasing the expression of the splicing …