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Justin Hall, Amy Brault, Fabien Vincent, Shawn Weng, Hong Wang, Darren Dumlao, Ann Aulabaugh, Dikran Aivazian, Dana Castro, Ming Chen, Jeffrey Culp, Ken Dower, Joseph Gardner, Steven Hawrylik, Douglas Golenbock, David Hepworth, Mark Horn, Lyn Jones, Peter Jones, Eicke Latz, Jing Li, Lih-Ling Lin, Wen Lin, David Lin, Frank Lovering, Nootaree Niljanskul, Ryan Nistler, Betsy Pierce, Olga Plotnikova, Daniel Schmitt, Suman Shanker, James Smith, William Snyder, Timothy Subashi, John Trujillo, Edyta Tyminski, Guoxing Wang, Jimson Wong, Bruce Lefker, Leslie Dakin, Karen Leach
Cyclic GMP-AMP synthase (cGAS) initiates the innate immune system in response to cytosolic dsDNA. After binding and activation from dsDNA, cGAS uses ATP and GTP to synthesize 2', 3' -cGAMP (cGAMP), a cyclic dinucleotide second messenger with mixed 2'-5' and 3'-5' phosphodiester bonds. Inappropriate stimulation of cGAS has been implicated in autoimmune disease such as systemic lupus erythematosus, thus inhibition of cGAS may be of therapeutic benefit in some diseases; however, the size and polarity of the cGAS active site makes it a challenging target for the development of conventional substrate-competitive inhibitors...
2017: PloS One
Marie K Schwinn, Thomas Machleidt, Kris Zimmerman, Christopher T Eggers, Andrew S Dixon, Robin Hurst, Mary P Hall, Lance P Encell, Brock F Binkowski, Keith V Wood
Intracellular signaling pathways are mediated by changes in protein abundance and post-translational modifications. A common approach for investigating signaling mechanisms and the effects induced by synthetic compounds is through overexpression of recombinant reporter genes. Genome editing with CRISPR/Cas9 offers a means to better preserve native biology by appending reporters directly onto the endogenous genes. An optimal reporter for this purpose would be small to negligibly influence intracellular processes, be readily linked to the endogenous genes with minimal experimental effort, and be sensitive enough to detect low expressing proteins...
February 16, 2018: ACS Chemical Biology
Kevin D Hall, Kong Y Chen, Juen Guo, Yan Y Lam, Rudolph L Leibel, Laurel Es Mayer, Marc L Reitman, Michael Rosenbaum, Steven R Smith, B Timothy Walsh, Eric Ravussin
BACKGROUND: The carbohydrate-insulin model of obesity posits that habitual consumption of a high-carbohydrate diet sequesters fat within adipose tissue because of hyperinsulinemia and results in adaptive suppression of energy expenditure (EE). Therefore, isocaloric exchange of dietary carbohydrate for fat is predicted to result in increased EE, increased fat oxidation, and loss of body fat. In contrast, a more conventional view that "a calorie is a calorie" predicts that isocaloric variations in dietary carbohydrate and fat will have no physiologically important effects on EE or body fat...
August 2016: American Journal of Clinical Nutrition
Carmen Trigo, Kurt A Spokas, Kathleen E Hall, Lucia Cox, William C Koskinen
Addition of organic amendments such as biochar to soils can influence pesticide sorption-desorption processes and, in turn, the amount of pesticide readily availability for transport and biodegradation. Sorption-desorption processes are affected by both the physical and chemical properties of soils and pesticides, as well as soil-pesticide contact time, or aging. Changes in sorption-desorption of metolachlor with aging in soil amended with three macadamia nut shell biochars aged 0 (BCmac-fr), 1 year (BCmac-1yr), and 2 years (BCmac-2yr) and two wood biochars aged 0 (BCwood-fr) and 5 years (BCwood-5yr) were determined...
April 27, 2016: Journal of Agricultural and Food Chemistry
M Gupta, D Neavin, D Liu, J Biernacka, D Hall-Flavin, W V Bobo, M A Frye, M Skime, G D Jenkins, A Batzler, K Kalari, W Matson, S S Bhasin, H Zhu, T Mushiroda, Y Nakamura, M Kubo, L Wang, R Kaddurah-Daouk, R M Weinshilboum
Millions of patients suffer from major depressive disorder (MDD), but many do not respond to selective serotonin reuptake inhibitor (SSRI) therapy. We used a pharmacometabolomics-informed pharmacogenomics research strategy to identify genes associated with metabolites that were related to SSRI response. Specifically, 306 MDD patients were treated with citalopram or escitalopram and blood was drawn at baseline, 4 and 8 weeks for blood drug levels, genome-wide single nucleotide polymorphism (SNP) genotyping and metabolomic analyses...
December 2016: Molecular Psychiatry
Andrew S Dixon, Marie K Schwinn, Mary P Hall, Kris Zimmerman, Paul Otto, Thomas H Lubben, Braeden L Butler, Brock F Binkowski, Thomas Machleidt, Thomas A Kirkland, Monika G Wood, Christopher T Eggers, Lance P Encell, Keith V Wood
Protein-fragment complementation assays (PCAs) are widely used for investigating protein interactions. However, the fragments used are structurally compromised and have not been optimized nor thoroughly characterized for accurately assessing these interactions. We took advantage of the small size and bright luminescence of NanoLuc to engineer a new complementation reporter (NanoBiT). By design, the NanoBiT subunits (i.e., 1.3 kDa peptide, 18 kDa polypeptide) weakly associate so that their assembly into a luminescent complex is dictated by the interaction characteristics of the target proteins onto which they are appended...
February 19, 2016: ACS Chemical Biology
Yeon Sun Lee, Sara M Hall, Cyf Ramos-Colon, Michael Remesic, David Rankin, Todd W Vanderah, Frank Porreca, Josephine Lai, Victor J Hruby
Dynorphin A (Dyn A) is an endogenous opioid ligand that possesses neuroinhibitory (antinociceptive) effects via μ, δ, and κ opioid receptors. However, under chronic pain conditions, up-regulated spinal Dyn A can also interact with bradykinin receptors (BRs) to promote hyperalgesia through a neuroexcitatory(pronociceptive) effect. These excitatory effects cannot be blocked by an opioid antagonist, and thus are non-opioid in nature. On the basis of the structural dissimilarity between Dyn A and endogenous BR ligands, bradykinin(BK) and kallidin (KD), Dyn A's interaction with BRs could not be predicted, and provided an opportunity to identify a novel potential neuroexcitatory target...
2015: Receptors & Clinical Investigation
Andrew K Urick, Laura M L Hawk, Melissa K Cassel, Neeraj K Mishra, Shuai Liu, Neeta Adhikari, Wei Zhang, Camila O dos Santos, Jennifer L Hall, William C K Pomerantz
Bromodomain-containing protein dysregulation is linked to cancer, diabetes, and inflammation. Selective inhibition of bromodomain function is a newly proposed therapeutic strategy. We describe a (19)F NMR dual screening method for small molecule discovery using fluorinated tryptophan resonances on two bromodomain-containing proteins. The chemical shift dispersion of (19)F resonances within fluorine-labeled proteins enables the simultaneous analysis of two fluorinated bromodomains by NMR. A library of 229 small molecules was screened against the first bromodomain of Brd4 and the BPTF bromodomain...
October 16, 2015: ACS Chemical Biology
Yuan Liu, Marty W Mayo, Aizhen Xiao, Emily H Hall, Elianna B Amin, Kyuichi Kadota, Prasad S Adusumilli, David R Jones
Breast cancer metastasis suppressor 1 (BRMS1) is downregulated in non-small cell lung cancer (NSCLC), and its reduction correlates with disease progression. Herein, we investigate the mechanisms through which loss of the BRMS1 gene contributes to epithelial-to-mesenchymal transition (EMT). Using a short hairpin RNA (shRNA) system, we show that loss of BRMS1 promotes basal and transforming growth factor beta-induced EMT in NSCLC cells. NSCLC cells expressing BRMS1 shRNAs (BRMS1 knockdown [BRMS1(KD)]) display mesenchymal characteristics, including enhanced cell migration and differential regulation of the EMT markers...
January 2015: Molecular and Cellular Biology
James Hall, Stefan Reschke, Hongnan Cao, Silke Leimkühler, Russ Hille
The kinetic properties of an E232Q variant of the xanthine dehydrogenase from Rhodobacter capsulatus have been examined to ascertain whether Glu(232) in wild-type enzyme is protonated or unprotonated in the course of catalysis at neutral pH. We find that kred, the limiting rate constant for reduction at high [xanthine], is significantly compromised in the variant, a result that is inconsistent with Glu(232) being neutral in the active site of the wild-type enzyme. A comparison of the pH dependence of both kred and kred/Kd from reductive half-reaction experiments between wild-type and enzyme and the E232Q variant suggests that the ionized Glu(232) of wild-type enzyme plays an important role in catalysis by discriminating against the monoanionic form of substrate, effectively increasing the pKa of substrate by two pH units and ensuring that at physiological pH the neutral form of substrate predominates in the Michaelis complex...
November 14, 2014: Journal of Biological Chemistry
Yeon Sun Lee, Dhanasekaran Muthu, Sara M Hall, Cyf Ramos-Colon, David Rankin, Jackie Hu, Alexander J Sandweiss, Milena De Felice, Jennifer Yanhua Xie, Todd W Vanderah, Frank Porreca, Josephine Lai, Victor J Hruby
We hypothesized that under chronic pain conditions, up-regulated dynorphin A (Dyn A) interacts with bradykinin receptors (BRs) in the spinal cord to promote hyperalgesia through an excitatory effect, which is opposite to the well-known inhibitory effect of opioid receptors. Considering the structural dissimilarity between Dyn A and endogenous BR ligands, bradykinin (BK) and kallidin (KD), this interaction could not be predicted, but it allowed us to discover a potential neuroexcitatory target. Well-known BR ligands, BK, [des-Arg(10), Leu(9)]-kallidin (DALKD), and HOE140 showed different binding profiles at rat brain BRs than that previously reported...
May 7, 2014: Journal of the American Chemical Society
D Brent Halling, Sophia A Kenrick, Austen F Riggs, Richard W Aldrich
Ca(2+) activates SK Ca(2+)-activated K(+) channels through the protein Ca(2+) sensor, calmodulin (CaM). To understand how SK channels operate, it is necessary to determine how Ca(2+) regulates CaM binding to its target on SK. Tagless, recombinant SK peptide (SKp), was purified for binding studies with CaM at low and high Ca(2+) concentrations. Composition gradient multi-angle light scattering accurately measures the molar mass, stoichiometry, and affinity of protein complexes. In 2 mM Ca(2+), SKp and CaM bind with three different stoichiometries that depend on the molar ratio of SKp:CaM in solution...
February 2014: Journal of General Physiology
C J Thomas, A R Smith, M A Hall
The ethylene-binding site (EBS) from Phaseolus vulgaris cv. Canadian Wonder cotyledons can be solubilised from 96,000 g pelleted material by Triton X-100 or sodium cholate. Extraction of 96,000 g pellets with acetone, butanol or butanol and ether results in a total loss of ethylene-binding activity. Like the membrane-bound form, the solubilised EBS has an apparent KD(liquid) of 10(-10) M at a concentration of 32 pmol EBS per gram tissue fresh weight. Propylene and acetylene act as competitive inhibitors, carbon dioxide appears to promote ethylene binding and ethane has no significant effect...
April 1984: Planta
I O Sanders, A R Smith, M A Hall
Using in-vivo assays at least two classes of ethylene-binding site are shown to exist in pea epicotyls. These classes appear to have identical affinities for ethylene with a KD of 6 · 10(-11)-8 · 10(-11) M; the fast associating class has high and appropriate affinities for physiologically active analogues of ethylene, and CO2 is without effect upon bfinding. Experiments involving suppression of endogenous ethylene production demonstrate that previous work may have underestimated both the amount of binding sites present and the affinity of such sites for ethylene...
January 1991: Planta
G G Anskjær, K A Krogh, B Halling-Sørensen
Equilibrium dialysis experiments, a novel approach for conducting soil/water distribution experiments in environmental samples, were found to be applicable for assessing pH-dependent partitioning and to quantify the sorption of three sulfonamides, sulfadiazine, sulfadoxine, and sulfacetamide. Clay fractions from two agricultural soils including both particulate and dissolved soil matter were used in the experiments to achieve a high sorption capacity when varying pH in a relevant environmental range. Stabilizing and controlling pH was done by using organic buffers...
January 2014: Chemosphere
Siew-Na Lim, Wenlong Huang, Jodie C E Hall, Adina T Michael-Titus, John V Priestley
In this study we have characterised the locomotor recovery, and temporal profile of cell loss, in a novel thoracic compression spinal cord injury (SCI) in the mouse. We have also shown that treatment with docosahexaenoic acid (DHA) is neuroprotective in this model of SCI, strengthening the growing literature demonstrating that omega-3 polyunsaturated fatty acids are neuroprotective after SCI. Compression SCI in C57BL/6 mice was produced by placing a 10 g weight for 5 min onto a 2 mm × 1.5 mm platform applied to the dura at vertebral level T12...
January 2013: Experimental Neurology
Celeste Bolin, Mohammed-Tayyib Boudra, Marie Fernet, Laurence Vaslin, Vincent Pennaneach, Tomasz Zaremba, Denis Biard, Fabrice P Cordelières, Vincent Favaudon, Frédérique Mégnin-Chanet, Janet Hall
Cyclin-dependent kinase 5 (Cdk5) has been identified as a determinant of sensitivity to poly(ADP-ribose) polymerase (PARP) inhibitors. Here, the consequences of its depletion on cell survival, PARP activity, the recruitment of base excision repair (BER) proteins to DNA damage sites, and overall DNA single-strand break (SSB) repair were investigated using isogenic HeLa stably depleted (KD) and Control cell lines. Synthetic lethality achieved by disrupting PARP activity in Cdk5-deficient cells was confirmed, and the Cdk5(KD) cells were also found to be sensitive to the killing effects of ionizing radiation (IR) but not methyl methanesulfonate or neocarzinostatin...
March 2012: Cellular and Molecular Life Sciences: CMLS
Cathinka Halle, Malin Lando, Debbie Hege Svendsrud, Trevor Clancy, Marit Holden, Kolbein Sundfør, Gunnar B Kristensen, Ruth Holm, Heidi Lyng
PURPOSE: We compared the prognostic significance of ectodomain isoforms of the epidermal growth factor receptor (EGFR), which lack the tyrosine kinase (TK) domain, with that of the full-length receptor and its autophosphorylation status in cervical cancers treated with conventional chemoradiotherapy. EXPERIMENTAL DESIGN: Expression of EGFR isoforms was assessed by immunohistochemistry in a prospectively collected cohort of 178 patients with squamous cell cervical carcinoma, and their detection was confirmed with Western blotting and reverse transcriptase PCR...
August 15, 2011: Clinical Cancer Research: An Official Journal of the American Association for Cancer Research
Mona Dvir-Ginzberg, Viktoria Gagarina, Eun Jin Lee, Richard Booth, Odile Gabay, David J Hall
OBJECTIVE: The protein deacetylase SirT1 positively regulates cartilage-specific gene expression, while the proinflammatory cytokine tumor necrosis factor α (TNFα) negatively regulates these same genes. This study was undertaken to test the hypothesis that SirT1 is adversely affected by TNFα, resulting in altered gene expression. METHODS: Cartilage-specific gene expression, SirT1 activity, and results of chromatin immunoprecipitation analysis at the α2(I) collagen enhancer site were determined in RNA, protein extracts, and nuclei of human osteoarthritic chondrocytes left untreated or treated with TNFα...
August 2011: Arthritis and Rheumatism
Ping-Ke Fang, Brian H Raphael, Susan E Maslanka, Shuowei Cai, Bal Ram Singh
BACKGROUND: Type A1 Clostridium botulinum strains are a group of Gram-positive, spore-forming anaerobic bacteria that produce a genetically, biochemically, and biophysically indistinguishable 150 kD protein that causes botulism. The genomes of three type A1 C. botulinum strains have been sequenced and show a high degree of synteny. The purpose of this study was to characterize differences among these genomes and compare these differentiating features with two additional unsequenced strains used in previous studies...
2010: BMC Genomics
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