WorldWideScience

Sample records for sam methyltransferase cfr

  1. Identification of 8-methyladenosine as the modification catalyzed by the radical SAM methyltransferase Cfr that confers antibiotic resistance in bacteria

    DEFF Research Database (Denmark)

    Giessing, Anders; Jensen, Søren Skov; Rasmussen, Anette

    2009-01-01

    ,8-dimethyladenosine. The mutation of single conserved cysteine residues in the radical SAM motif CxxxCxxC of Cfr abolishes its activity, lending support to the notion that the Cfr modification reaction occurs via a radical-based mechanism. Antibiotic susceptibility data confirm that the antibiotic resistance...

  2. Insights into the structure, function and evolution of the radical-SAM 23S rRNA methyltransferase Cfr that confers antibiotic resistance in bacteria

    DEFF Research Database (Denmark)

    Karminska, K. H.; Purta, E.; Hansen, L .H.

    2010-01-01

    of a 4Fe-4S cluster, a SAM molecule coordinated to the iron-sulfur cluster (SAM1) and a SAM molecule that is the putative methyl group donor (SAM2). All mutations at predicted functional sites affect Cfr activity significantly as assayed by antibiotic susceptibility testing and primer extension analysis....... The investigation has identified essential amino acids and Cfr variants with altered reaction mechanisms and represents a first step towards understanding the structural basis of Cfr activity....

  3. Insights into the structure, function and evolution of the radical-SAM 23S rRNA methyltransferase Cfr that confers antibiotic resistance in bacteria

    DEFF Research Database (Denmark)

    Karminska, K. H.; Purta, E.; Hansen, L .H.

    2010-01-01

    The Cfr methyltransferase confers combined resistance to five classes of antibiotics that bind to the peptidyl tranferase center of bacterial ribosomes by catalyzing methylation of the C-8 position of 23S rRNA nucleotide A2503. The same nucleotide is targeted by the housekeeping methyltransferase...

  4. The U6 snRNA m6A Methyltransferase METTL16 Regulates SAM Synthetase Intron Retention.

    Science.gov (United States)

    Pendleton, Kathryn E; Chen, Beibei; Liu, Kuanqing; Hunter, Olga V; Xie, Yang; Tu, Benjamin P; Conrad, Nicholas K

    2017-05-18

    Maintenance of proper levels of the methyl donor S-adenosylmethionine (SAM) is critical for a wide variety of biological processes. We demonstrate that the N6-adenosine methyltransferase METTL16 regulates expression of human MAT2A, which encodes the SAM synthetase expressed in most cells. Upon SAM depletion by methionine starvation, cells induce MAT2A expression by enhanced splicing of a retained intron. Induction requires METTL16 and its methylation substrate, a vertebrate conserved hairpin (hp1) in the MAT2A 3' UTR. Increasing METTL16 occupancy on the MAT2A 3' UTR is sufficient to induce efficient splicing. We propose that, under SAM-limiting conditions, METTL16 occupancy on hp1 increases due to inefficient enzymatic turnover, which promotes MAT2A splicing. We further show that METTL16 is the long-unknown methyltransferase for the U6 spliceosomal small nuclear RNA (snRNA). These observations suggest that the conserved U6 snRNA methyltransferase evolved an additional function in vertebrates to regulate SAM homeostasis. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Distinction between the Cfr Methyltransferase Conferring Antibiotic Resistance and the Housekeeping RlmN Methyltransferase

    DEFF Research Database (Denmark)

    Atkinson, Gemma C; Hansen, Lykke H; Tenson, Tanel

    2013-01-01

    The cfr gene encodes the Cfr methyltransferase that primarily methylates C-8 in A2503 of 23S rRNA in the peptidyl transferase region of bacterial ribosomes. The methylation provides resistance to six classes of antibiotics of clinical and veterinary importance. The rlmN gene encodes the RlmN meth...

  6. An intact SAM-dependent methyltransferase fold is encoded by the human endothelin-converting enzyme-2 gene

    Energy Technology Data Exchange (ETDEWEB)

    Tempel, W.; Wu, H.; Dombrovsky, L.; Zeng, H.; Loppnau, P.; Zhu, H.; Plotnikov, A.N.; Bochkarev, A.; (Toronto)

    2010-08-17

    A recent survey of protein expression patterns in patients with Alzheimer's disease (AD) has identified ece2 (chromosome: 3; Locations: 3q27.1) as the most significantly downregulated gene within the tested group. ece2 encodes endothelin-converting enzyme ECE2, a metalloprotease with a role in neuropeptide processing. Deficiency in the highly homologous ECE1 has earlier been linked to increased levels of AD-related {beta}-amyloid peptide in mice, consistent with a role for ECE in the degradation of that peptide. Initially, ECE2 was presumed to resemble ECE1, in that it comprises a single transmembrane region of {approx}20 residues flanked by a small amino-terminal cytosolic segment and a carboxy-terminal lumenar peptidase domain. The carboxy-terminal domain has significant sequence similarity to both neutral endopeptidase, for which an X-ray structure has been determined, and Kell blood group protein. After their initial discovery, multiple isoforms of ECE1 and ECE2 were discovered, generated by alternative splicing of multiple exons. The originally described ece2 transcript, RefSeq NM{_}174046, contains the amino-terminal cytosolic portion followed by the transmembrane region and peptidase domain (Fig. 1, isoform B). Another ece2 transcript, available from the Mammalian Gene Collection under MGC2408 (Fig. 1, isoform C), RefSeq accession NM{_}032331, is predicted to be translated into a 255 residue peptide with low but detectable sequence similarity to known S-adenosyl-L-methionine (SAM)-dependent methyltransferases (SAM-MTs), such as the hypothetical protein TT1324 from Thermus thermophilis, PDB code 2GS9, which shares 30% amino acid sequence identity with ECE2 over 138 residues of the sequence. Intriguingly, another 'elongated' ece2 transcript (Fig. 1, isoform A) (RefSeq NM{_}014693) contains an amino-terminal portion of the putative SAM-MT domain, the transmembrane domain, and the protease domain. This suggests the possibility for coexistence of

  7. The crystal structure of Nep1 reveals an extended SPOUT-class methyltransferase fold and a pre-organized SAM-binding site

    OpenAIRE

    Taylor, Alexander B.; Meyer, Britta; Leal, Belinda Z.; Kötter, Peter; Schirf, Virgil; Demeler, Borries; Hart, P. John; Entian, Karl-Dieter; Wöhnert, Jens

    2008-01-01

    Ribosome biogenesis in eukaryotes requires the participation of a large number of ribosome assembly factors. The highly conserved eukaryotic nucleolar protein Nep1 has an essential but unknown function in 18S rRNA processing and ribosome biogenesis. In Saccharomyces cerevisiae the malfunction of a temperature-sensitive Nep1 protein (nep1-1ts) was suppressed by the addition of S-adenosylmethionine (SAM). This suggests the participation of Nep1 in a methyltransferase reaction during ribosome bi...

  8. The crystal structure of Nep1 reveals an extended SPOUT-class methyltransferase fold and a pre-organized SAM-binding site

    OpenAIRE

    Taylor, Alexander B.; Meyer, Britta; Leal, Belinda Z.; Kötter, Peter; Schirf, Virgil; Demeler, Borries; Hart, P. John; Entian, Karl-Dieter; Wöhnert, Jens

    2008-01-01

    Ribosome biogenesis in eukaryotes requires the participation of a large number of ribosome assembly factors. The highly conserved eukaryotic nucleolar protein Nep1 has an essential but unknown function in 18S rRNA processing and ribosome biogenesis. In Saccharomyces cerevisiae the malfunction of a temperature-sensitive Nep1 protein (nep1-1ts ) was suppressed by the addition of S-adenosylmethionine (SAM). This suggests the participation of Nep1 in a methyltransferase reaction during ribosome b...

  9. A New Structural Form in the SAM/Metal-Dependent O;#8209;Methyltransferase Family: MycE from the Mycinamicin Biosynthetic Pathway

    Energy Technology Data Exchange (ETDEWEB)

    Akey, David L.; Li, Shengying; Konwerski, Jamie R.; Confer, Laura A.; Bernard, Steffen M.; Anzai, Yojiro; Kato, Fumio; Sherman, David H.; Smith, Janet L. (Michigan); (Toho)

    2012-08-01

    O-linked methylation of sugar substituents is a common modification in the biosynthesis of many natural products and is catalyzed by multiple families of S-adenosyl-l-methionine (SAM or AdoMet)-dependent methyltransferases (MTs). Mycinamicins, potent antibiotics from Micromonospora griseorubida, can be methylated at two positions on a 6-deoxyallose substituent. The first methylation is catalyzed by MycE, a SAM- and metal-dependent MT. Crystal structures were determined for MycE bound to the product S-adenosyl-l-homocysteine (AdoHcy) and magnesium, both with and without the natural substrate mycinamicin VI. This represents the first structure of a natural product sugar MT in complex with its natural substrate. MycE is a tetramer of a two-domain polypeptide, comprising a C-terminal catalytic MT domain and an N-terminal auxiliary domain, which is important for quaternary assembly and for substrate binding. The symmetric MycE tetramer has a novel MT organization in which each of the four active sites is formed at the junction of three monomers within the tetramer. The active-site structure supports a mechanism in which a conserved histidine acts as a general base, and the metal ion helps to position the methyl acceptor and to stabilize a hydroxylate intermediate. A conserved tyrosine is suggested to support activity through interactions with the transferred methyl group from the SAM methyl donor. The structure of the free enzyme reveals a dramatic order-disorder transition in the active site relative to the S-adenosyl-L-homocysteine complexes, suggesting a mechanism for product/substrate exchange through concerted movement of five loops and the polypeptide C-terminus.

  10. A SAM-dependent methyltransferase cotranscribed with arsenate reductase alters resistance to peptidyl transferase center-binding antibiotics in Azospirillum brasilense Sp7.

    Science.gov (United States)

    Singh, Sudhir; Singh, Chhaya; Tripathi, Anil Kumar

    2014-05-01

    The genome of Azospirillum brasilense harbors a gene encoding S-adenosylmethionine-dependent methyltransferase, which is located downstream of an arsenate reductase gene. Both genes are cotranscribed and translationally coupled. When they were cloned and expressed individually in an arsenate-sensitive strain of Escherichia coli, arsenate reductase conferred tolerance to arsenate; however, methyltransferase failed to do so. Sequence analysis revealed that methyltransferase was more closely related to a PrmB-type N5-glutamine methyltransferase than to the arsenate detoxifying methyltransferase ArsM. Insertional inactivation of prmB gene in A. brasilense resulted in an increased sensitivity to chloramphenicol and resistance to tiamulin and clindamycin, which are known to bind at the peptidyl transferase center (PTC) in the ribosome. These observations suggested that the inability of prmB:km mutant to methylate L3 protein might alter hydrophobicity in the antibiotic-binding pocket of the PTC, which might affect the binding of chloramphenicol, clindamycin, and tiamulin differentially. This is the first report showing the role of PrmB-type N5-glutamine methyltransferases in conferring resistance to tiamulin and clindamycin in any bacterium.

  11. The Cfr rRNA methyltransferase confers resistance to Phenicols, Lincosamides, Oxazolidinones, Pleuromutilins, and Streptogramin A antibiotics

    DEFF Research Database (Denmark)

    Long, K. S.; Poehlsgaard, Jacob; Kehrenberg, C.

    2006-01-01

    A novel multidrug resistance phenotype mediated by the Cfr rRNA methyltransferase is observed in Staphylococcus aureus and Escherichia coli. The cfr gene has previously been identified as a phenicol and lincosamide resistance gene on plasmids isolated from Staphylococcus spp. of animal origin...

  12. Molecular Basis of Substrate Promiscuity for the SAM-Dependent O-Methyltransferase NcsB1, Involved in the Biosynthesis of the Enediyne Antitumor Antibiotic Neocarzinostatin

    Energy Technology Data Exchange (ETDEWEB)

    Cooke, H.; Guenther, E; Luo, Y; Shen, B; Bruner, S

    2009-01-01

    The small molecule component of chromoprotein enediyne antitumor antibiotics is biosynthesized through a convergent route, incorporating amino acid, polyketide, and carbohydrate building blocks around a central enediyne hydrocarbon core. The naphthoic acid moiety of the enediyne neocarzinostatin plays key roles in the biological activity of the natural product by interacting with both the carrier protein and duplex DNA at the site of action. We have previously described the in vitro characterization of an S-adenosylmethionine-dependent O-methyltransferase (NcsB1) in the neocarzinostatin biosynthetic pathway [Luo, Y., Lin, S., Zhang, J., Cooke, H. A., Bruner, S. D., and Shen, B. (2008) J. Biol. Chem. 283, 14694-14702]. Here we provide a structural basis for NcsB1 activity, illustrating that the enzyme shares an overall architecture with a large family of S-adenosylmethionine-dependent proteins. In addition, NcsB1 represents the first enzyme to be structurally characterized in the biosynthetic pathway of neocarzinostatin. By cocrystallizing the enzyme with various combinations of the cofactor and substrate analogues, details of the active site structure have been established. Changes in subdomain orientation were observed via comparison of structures in the presence and absence of substrate, suggesting that reorientation of the enzyme is involved in binding of the substrate. In addition, residues important for substrate discrimination were predicted and probed through site-directed mutagenesis and in vitro biochemical characterization.

  13. Combined Effect of the Cfr Methyltransferase and Ribosomal Protein L3 Mutations on Resistance to Ribosome-Targeting Antibiotics

    DEFF Research Database (Denmark)

    Pakula, Kevin K; Hansen, Lykke H; Vester, Birte

    2017-01-01

    Several groups of antibiotics inhibit bacterial growth by binding to bacterial ribosomes. Mutations in ribosomal protein L3 have been associated with resistance to linezolid and tiamulin, which both bind at the peptidyl transferase center in the ribosome. Resistance to these and other antibiotics...... also occurs through methylation of 23S rRNA at position A2503 by the methyltransferase Cfr. The mutations in L3 and the cfr gene have been found together in clinical isolates, raising the question of whether they have a combined effect on antibiotic resistance or growth. We transformed a plasmid...... seen. This study underscores the complex interplay between various resistance mechanisms and cross-resistance, even from antibiotics with overlapping binding sites....

  14. SAM in a Nutshell.

    Science.gov (United States)

    Givens, Larry R.

    2000-01-01

    Explains what the Association of Higher Education Facilities Officers' Strategic Assessment Model (SAM) is and how to use it to achieve organizational excellence through continuous improvement. Showing features of both the Malcolm Baldrige programs and the Balanced Scorecard, the SAM components are described along with an explanation of the four…

  15. SAM EMP (SEMP) model

    Energy Technology Data Exchange (ETDEWEB)

    Thatcher, R.M.

    1984-05-01

    The Surface-To-Air Missile (SAM) Electro-Magnetic-Pulse (EMP) (SEMP) model simulates the illumination of an entire SAM brigade with an EMP weapon. It computes probability distributions of SAM brigade performance levels after an EMP attack has occurred. Brigade performance is determined by the combination of components that survive the EMP. Accordingly, the SEMP model is separated into the component failure model and the condition model. The component failure model computes the failure probability of each component in the brigade from data supplied by two input data files. The condition model converts component failure probabilities into brigade performance in the form of missile availability probability tables.

  16. SAM Photovoltaic Model Technical Reference

    Energy Technology Data Exchange (ETDEWEB)

    Gilman, P. [National Renewable Energy Laboratory (NREL), Golden, CO (United States)

    2015-05-27

    This manual describes the photovoltaic performance model in the System Advisor Model (SAM). The U.S. Department of Energy’s National Renewable Energy Laboratory maintains and distributes SAM, which is available as a free download from https://sam.nrel.gov. These descriptions are based on SAM 2015.1.30 (SSC 41).

  17. Congressmember Sam Farr

    OpenAIRE

    Farmer, Ellen

    2010-01-01

    United States Congressmember Sam Farr, one of the political heroes of the sustainable agriculture movement, was interviewed by Ellen Farmer on August 23, 2007. A fifth-generation Californian, Farr was born in 1941. He is the son of California State Senator Fred Farr, who sponsored a law requiring toilets in the fields for farm workers, as well as other landmark environmental legislation. Sam Farr began his career in public service in 1964, in the Peace Corps in Colombia. Before his e...

  18. SAM Theory Manual

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Rui [Argonne National Lab. (ANL), Argonne, IL (United States)

    2017-03-01

    The System Analysis Module (SAM) is an advanced and modern system analysis tool being developed at Argonne National Laboratory under the U.S. DOE Office of Nuclear Energy’s Nuclear Energy Advanced Modeling and Simulation (NEAMS) program. SAM development aims for advances in physical modeling, numerical methods, and software engineering to enhance its user experience and usability for reactor transient analyses. To facilitate the code development, SAM utilizes an object-oriented application framework (MOOSE), and its underlying meshing and finite-element library (libMesh) and linear and non-linear solvers (PETSc), to leverage modern advanced software environments and numerical methods. SAM focuses on modeling advanced reactor concepts such as SFRs (sodium fast reactors), LFRs (lead-cooled fast reactors), and FHRs (fluoride-salt-cooled high temperature reactors) or MSRs (molten salt reactors). These advanced concepts are distinguished from light-water reactors in their use of single-phase, low-pressure, high-temperature, and low Prandtl number (sodium and lead) coolants. As a new code development, the initial effort has been focused on modeling and simulation capabilities of heat transfer and single-phase fluid dynamics responses in Sodium-cooled Fast Reactor (SFR) systems. The system-level simulation capabilities of fluid flow and heat transfer in general engineering systems and typical SFRs have been verified and validated. This document provides the theoretical and technical basis of the code to help users understand the underlying physical models (such as governing equations, closure models, and component models), system modeling approaches, numerical discretization and solution methods, and the overall capabilities in SAM. As the code is still under ongoing development, this SAM Theory Manual will be updated periodically to keep it consistent with the state of the development.

  19. Structural basis for substrate recognition in the salicylic acid carboxyl methyltransferase family.

    Science.gov (United States)

    Zubieta, Chloe; Ross, Jeannine R; Koscheski, Paul; Yang, Yue; Pichersky, Eran; Noel, Joseph P

    2003-08-01

    Recently, a novel family of methyltransferases was identified in plants. Some members of this newly discovered and recently characterized methyltransferase family catalyze the formation of small-molecule methyl esters using S-adenosyl-L-Met (SAM) as a methyl donor and carboxylic acid-bearing substrates as methyl acceptors. These enzymes include SAMT (SAM:salicylic acid carboxyl methyltransferase), BAMT (SAM:benzoic acid carboxyl methyltransferase), and JMT (SAM:jasmonic acid carboxyl methyltransferase). Moreover, other members of this family of plant methyltransferases have been found to catalyze the N-methylation of caffeine precursors. The 3.0-A crystal structure of Clarkia breweri SAMT in complex with the substrate salicylic acid and the demethylated product S-adenosyl-L-homocysteine reveals a protein structure that possesses a helical active site capping domain and a unique dimerization interface. In addition, the chemical determinants responsible for the selection of salicylic acid demonstrate the structural basis for facile variations of substrate selectivity among functionally characterized plant carboxyl-directed and nitrogen-directed methyltransferases and a growing set of related proteins that have yet to be examined biochemically. Using the three-dimensional structure of SAMT as a guide, we examined the substrate specificity of SAMT by site-directed mutagenesis and activity assays against 12 carboxyl-containing small molecules. Moreover, the utility of structural information for the functional characterization of this large family of plant methyltransferases was demonstrated by the discovery of an Arabidopsis methyltransferase that is specific for the carboxyl-bearing phytohormone indole-3-acetic acid.

  20. Zika Virus Methyltransferase: Structure and Functions for Drug Design Perspectives.

    Science.gov (United States)

    Coutard, Bruno; Barral, Karine; Lichière, Julie; Selisko, Barbara; Martin, Baptiste; Aouadi, Wahiba; Lombardia, Miguel Ortiz; Debart, Françoise; Vasseur, Jean-Jacques; Guillemot, Jean Claude; Canard, Bruno; Decroly, Etienne

    2017-03-01

    The Flavivirus Zika virus (ZIKV) is the causal agent of neurological disorders like microcephaly in newborns or Guillain-Barre syndrome. Its NS5 protein embeds a methyltransferase (MTase) domain involved in the formation of the viral mRNA cap. We investigated the structural and functional properties of the ZIKV MTase. We show that the ZIKV MTase can methylate RNA cap structures at the N-7 position of the cap, and at the 2'-O position on the ribose of the first nucleotide, yielding a cap-1 structure. In addition, the ZIKV MTase methylates the ribose 2'-O position of internal adenosines of RNA substrates. The crystal structure of the ZIKV MTase determined at a 2.01-Å resolution reveals a crystallographic homodimer. One chain is bound to the methyl donor ( S -adenosyl-l-methionine [SAM]) and shows a high structural similarity to the dengue virus (DENV) MTase. The second chain lacks SAM and displays conformational changes in the αX α-helix contributing to the SAM and RNA binding. These conformational modifications reveal a possible molecular mechanism of the enzymatic turnover involving a conserved Ser/Arg motif. In the second chain, the SAM binding site accommodates a sulfate close to a glycerol that could serve as a basis for structure-based drug design. In addition, compounds known to inhibit the DENV MTase show similar inhibition potency on the ZIKV MTase. Altogether these results contribute to a better understanding of the ZIKV MTase, a central player in viral replication and host innate immune response, and lay the basis for the development of potential antiviral drugs. IMPORTANCE The Zika virus (ZIKV) is associated with microcephaly in newborns, and other neurological disorders such as Guillain-Barre syndrome. It is urgent to develop antiviral strategies inhibiting the viral replication. The ZIKV NS5 embeds a methyltransferase involved in the viral mRNA capping process, which is essential for viral replication and control of virus detection by innate immune

  1. Sam and Wahua (24)

    African Journals Online (AJOL)

    DELL

    2017-10-27

    Oct 27, 2017 ... This study investigated the anatomical features, nutraceutical potentials and heavy metal compositions in two varieties of .... variety (Plates 1-6). Sam and Wahua: Comparative Anatomy, Nutraceutical Potentials and Heavy Metal Composition .... High content of tannin decreases protein quality by decreasing ...

  2. Structure-Activity Relationships of Diverse Oxazolidinones for Linezolid-Resistant Staphylococcus aureus Strains Possessing the cfr Methyltransferase Gene or Ribosomal Mutations▿

    Science.gov (United States)

    Locke, Jeffrey B.; Finn, John; Hilgers, Mark; Morales, Gracia; Rahawi, Shahad; G. C., Kedar; Picazo, Juan José; Im, Weonbin; Shaw, Karen Joy; Stein, Jeffrey L.

    2010-01-01

    Staphylococcal resistance to linezolid (LZD) is mediated through ribosomal mutations (23S rRNA or ribosomal proteins L3 and L4) or through methylation of 23S rRNA by the horizontally transferred Cfr methyltransferase. To investigate the structural basis for oxazolidinone activity against LZD-resistant (LZDr) strains, we compared structurally diverse, clinically relevant oxazolidinones, including LZD, radezolid (RX-1741), TR-700 (torezolid), and a set of TR-700 analogs (including novel CD-rings and various A-ring C-5 substituents), against a panel of laboratory-derived and clinical LZDr Staphylococcus aureus strains possessing a variety of resistance mechanisms. Potency against all strains was correlated with optimization of C- and D-rings, which interact with more highly conserved regions of the peptidyl transferase center binding site. Activity against cfr strains was retained with either hydroxymethyl or 1,2,3-triazole C-5 groups but was reduced by 2- to 8-fold in compounds with acetamide substituents. LZD, which possesses a C-5 acetamide group and lacks a D-ring substituent, demonstrated the lowest potency against all strains tested, particularly against cfr strains. These data reveal key features contributing to oxazolidinone activity and highlight structural tradeoffs between potency against susceptible strains and potency against strains with various resistance mechanisms. PMID:20837751

  3. Biochemical and Computational Analysis of the Substrate Specificities of Cfr and RlmN Methyltransferases

    DEFF Research Database (Denmark)

    Ntokou, Eleni; Hansen, Lykke Haastrup; Kongsted, Jacob

    2015-01-01

    interchangeability between Cfr and RlmN we constructed various combinations of their genes. The function of the mixed genes was investigated by RNA primer extension analysis to reveal methylation at 23S rRNA position A2503 and by MIC analysis to reveal antibiotic resistance. The catalytic site is expected...... to be responsible for the C2/C8 specificity and most of the combinations involve interchanging segments at this site. Almost all replacements showed no function in the primer extension assay, apart from a few that had a weak effect. Thus Cfr and RlmN appear to be much less similar than expected from their sequence......-ray structure of RlmN. We used a trinucleotide as target sequence and assessed its positioning at the active site for methylation. The calculations are in accordance with different poses of the trinucleotide in the two enzymes indicating major evolutionary changes to shift the C2/C8 specificities. To explore...

  4. Structural mechanism of S-adenosyl methionine binding to catechol O-methyltransferase.

    Directory of Open Access Journals (Sweden)

    Douglas Tsao

    Full Text Available Methyltransferases possess a homologous domain that requires both a divalent metal cation and S-adenosyl-L-methionine (SAM to catalyze its reactions. The kinetics of several methyltransferases has been well characterized; however, the details regarding their structural mechanisms have remained unclear to date. Using catechol O-methyltransferase (COMT as a model, we perform discrete molecular dynamics and computational docking simulations to elucidate the initial stages of cofactor binding. We find that COMT binds SAM via an induced-fit mechanism, where SAM adopts a different docking pose in the absence of metal and substrate in comparison to the holoenzyme. Flexible modeling of the active site side-chains is essential for observing the lowest energy state in the apoenzyme; rigid docking tools are unable to recapitulate the pose unless the appropriate side-chain conformations are given a priori. From our docking results, we hypothesize that the metal reorients SAM in a conformation suitable for donating its methyl substituent to the recipient ligand. The proposed mechanism enables a general understanding of how divalent metal cations contribute to methyltransferase function.

  5. CareSam

    DEFF Research Database (Denmark)

    Liveng, Anne; Christensen, Jonas

    2016-01-01

    This article presents findings and discussions generated on the basis of the Danish-Swedish development project CareSam. The article will on the one hand focus on how work in groups consisting of representatives from different levels in the elderly care sector at one time served as learning spaces...... and cultural encounters in which established notions of older people and elderly care were challenged and discussed. Inspired by action research these challenges were brought forth through discussions of and through insight in practical experiences. On the other hand it will focus on the tendencies to narrow...... in the project-groups this paper will ask whether it is possible to represent care work for elderly people with all the ambiguities it holds: How can we as researchers represent both meaningfulness and straining dimensions of care work? Can we avoid either supporting Florence Nightingale-ideals or cementing...

  6. Nucleosome Binding Alters the Substrate Bonding Environment of Histone H3 Lysine 36 Methyltransferase NSD2.

    Science.gov (United States)

    Poulin, Myles B; Schneck, Jessica L; Matico, Rosalie E; Hou, Wangfang; McDevitt, Patrick J; Holbert, Marc; Schramm, Vern L

    2016-06-01

    Nuclear receptor-binding SET domain protein 2 (NSD2) is a histone H3 lysine 36 (H3K36)-specific methyltransferase enzyme that is overexpressed in a number of cancers, including multiple myeloma. NSD2 binds to S-adenosyl-l-methionine (SAM) and nucleosome substrates to catalyze the transfer of a methyl group from SAM to the ε-amino group of histone H3K36. Equilibrium binding isotope effects and density functional theory calculations indicate that the SAM methyl group is sterically constrained in complex with NSD2, and that this steric constraint is released upon nucleosome binding. Together, these results show that nucleosome binding to NSD2 induces a significant change in the chemical environment of enzyme-bound SAM.

  7. An Arabidopsis thaliana methyltransferase Capable of Methylating Farnesoic Acid

    Energy Technology Data Exchange (ETDEWEB)

    Yang,Y.; Yuan, J.; Ross, J.; Noel, J.; Pichersky, E.

    2006-01-01

    We previously reported the identification of a new family of plant methyltransferases (MTs), named the SABATH family, that use S-adenosyl-l-methionine (SAM) to methylate a carboxyl moiety or a nitrogen-containing functional group on a diverse array of plant compounds. The Arabidopsis genome alone contains 24 distinct SABATH genes. To identify the catalytic specificities of members of this protein family in Arabidopsis, we screened recombinantly expressed and purified enzymes with a large number of potential substrates. Here, we report that the Arabidopsis thaliana gene At3g44860 encodes a protein with high catalytic specificity towards farnesoic acid (FA). Under steady-state conditions, this farnesoic acid carboxyl methyltransferase (FAMT) exhibits K{sub M} values of 41 and 71 {mu}M for FA and SAM, respectively. A three-dimensional model of FAMT constructed based upon similarity to the experimentally determined structure of Clarkia breweri salicylic acid methyltransferase (SAMT) suggests a reasonable model for FA recognition in the FAMT active site. In plants, the mRNA levels of At3g44860 increase in response to the exogenous addition of several compounds previously shown to induce plant defense responses at the transcriptional level. Although methyl farnesoate (MeFA) has not yet been detected in Arabidopsis, the presence of a FA-specific carboxyl methyltransferase in Arabidopsis capable of producing MeFA, an insect juvenile hormone made by some plants as a presumed defense against insect herbivory, suggests that MeFA or chemically similar compounds are likely to serve as new specialized metabolites in Arabidopsis.

  8. Combined Effect of the Cfr Methyltransferase and Ribosomal Protein L3 Mutations on Resistance to Ribosome-Targeting Antibiotics

    DEFF Research Database (Denmark)

    Pakula, Kevin K; Hansen, Lykke H; Vester, Birte

    2017-01-01

    . The presence of Cfr has a very minor influence on the growth rate. The resistance of the transformants to linezolid, tiamulin, florfenicol, and Synercid (a combination of quinupristin and dalfopristin [Q-D]) was measured by MIC assays. The resistance from Cfr was, in all cases, stronger than the effects...... of the L3 mutations, but various effects were obtained with the combinations of Cfr and L3 mutations ranging from a synergistic to an antagonistic effect. Linezolid and tiamulin susceptibility varied greatly among the L3 mutations, while no significant effects on florfenicol and Q-D susceptibility were...

  9. System for Award Management (SAM) API

    Data.gov (United States)

    General Services Administration — The SAM API is a RESTful method of retrieving public information about the businesses, organizations, or individuals (referred to as entities) within the SAM entity...

  10. Uncle Sam vs. John Bull.

    Science.gov (United States)

    Mellini, Peter

    1990-01-01

    Compares John Bull and Uncle Sam as iconographic symbols, respectively personifying male images of the British and United States national characters. Recounts their origins, evolutions, and representative values, and includes cartoons depicting the evolution. Describes female counterparts: Britannia and Columbia/Liberty. (CH)

  11. Influence of thiopurine methyltransferase gene polymorphism on ...

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Genetics; Volume 96; Issue 6. Influence of thiopurine methyltransferase gene polymorphism on Egyptian children with acute lymphoblastic leukaemia. AZZA A. G. ... thiopurine methyltransferase gene polymorphism; acute lymphoblastic leukaemia; Egyptian children; thiopurine methyltransferase.

  12. Structural insights into substrate selectivity of ribosomal RNA methyltransferase RlmCD.

    Directory of Open Access Journals (Sweden)

    Yiyang Jiang

    Full Text Available RlmCD has recently been identified as the S-adenosyl methionine (SAM-dependent methyltransferase responsible for the formation of m5U at U747 and U1939 of 23S ribosomal RNA in Streptococcus pneumoniae. In this research, we determine the high-resolution crystal structures of apo-form RlmCD and its complex with SAH. Using an in-vitro methyltransferase assay, we reveal the crucial residues for its catalytic functions. Furthermore, structural comparison between RlmCD and its structural homologue RumA, which only catalyzes the m5U1939 in Escherichia coli, implicates that a unique long linker in the central domain of RlmCD is the key factor in determining its substrate selectivity. Its significance in the enzyme activity of RlmCD is further confirmed by in-vitro methyltransferase assay.

  13. Histone methyltransferases in cancer

    DEFF Research Database (Denmark)

    Albert, Mareike; Helin, Kristian

    2009-01-01

    Cancer is perceived as a heterogeneous group of diseases that is characterized by aberrant patterns of gene expression. In the last decade, an increasing amount of data has pointed to a key role for epigenetic alterations in human cancer. In this review, we focus on a subclass of epigenetic...... regulators, namely histone methyltransferases (HMTs). Several HMTs have been linked to different types of cancer; however, in most cases we only have limited knowledge regarding the molecular mechanisms by which the HMTs contribute to disease development. We summarize the current knowledge regarding some...

  14. Samsø Energy Vision 2030

    DEFF Research Database (Denmark)

    Mathiesen, Brian Vad; Hansen, Kenneth; Ridjan, Iva

    The purpose of this report is to investigate potential scenarios for converting Samsø into 100% renewable energy supply in 2030 with focus on local electricity and biomass resources. Firstly, a 2013 reference scenario is established to investigate whether Samsø is 100% renewable today. Next, scen...

  15. Radical SAM-Mediated Methylation of Ribosomal RNA

    Science.gov (United States)

    Stojkovic, Vanja; Fujimori, Danica Galonić

    2015-01-01

    Post-transcriptional modifications of RNA play an important role in a wide range of biological processes. In ribosomal RNA (rRNA), methylation of nucleotide bases is the predominant modification. In recent years, methylation of adenosine 2503 (A2503) in bacterial 23S rRNA has attracted significant attention due to both the unusual regioselectivity of the methyl group incorporation, as well as the pathophysiological roles of the resultant methylations. Specifically, A2503 is methylated at the C2 and C8 positions of the adenine ring, and the introduced modifications have a profound impact on translational fidelity and antibiotic resistance, respectively. These modifications are performed by RlmN and Cfr, two members, of the recently discovered class of radical S-adenosylmethionine (radical SAM) methylsynthases. Here, we present several methods that can be used to evaluate the activity of these enzymes, under both in vivo and in vitro conditions. PMID:26253978

  16. Elongation factor methyltransferase 3--a novel eukaryotic lysine methyltransferase.

    Science.gov (United States)

    Zhang, Lelin; Hamey, Joshua J; Hart-Smith, Gene; Erce, Melissa A; Wilkins, Marc R

    2014-08-22

    Here we describe the discovery of Saccharomycescerevisiae protein YJR129Cp as a new eukaryotic seven-beta-strand lysine methyltransferase. An immunoblotting screen of 21 putative methyltransferases showed a loss in the methylation of elongation factor 2 (EF2) on knockout of YJR129C. Mass spectrometric analysis of EF2 tryptic peptides localised this loss of methylation to lysine 509, in peptide LVEGLKR. In vitro methylation, using recombinant methyltransferases and purified EF2, validated YJR129Cp as responsible for methylation of lysine 509 and Efm2p as responsible for methylation at lysine 613. Contextualised on previously described protein structures, both sites of methylation were found at the interaction interface between EF2 and the 40S ribosomal subunit. In line with the recently discovered Efm1 and Efm2 we propose that YJR129C be named elongation factor methyltransferase 3 (Efm3). The human homolog of Efm3 is likely to be the putative methyltransferase FAM86A, according to sequence homology and multiple lines of literature evidence. Copyright © 2014 Elsevier Inc. All rights reserved.

  17. Colorimetric activity measurement of a recombinant putrescine N-methyltransferase from Datura stramonium.

    Science.gov (United States)

    Biastoff, Stefan; Teuber, Michael; Zhou, Zhaohui Sunny; Dräger, Birgit

    2006-10-01

    Putrescine N-methyltransferase (PMT, EC 2.1.1.53) catalyses the S-adenosyl- L-methionine (SAM or AdoMet)-dependent methylation of putrescine to N-methylputrescine within the biosynthetic pathways of calystegines, nicotine, and tropane alkaloids in medicinal plants and produces S-adenosyl- L-homocysteine (SAH or AdoHcy). Determination of PMT activity was time-consuming and hardly reproducible in the past because it required tedious separation steps after chemical derivatisation or radioactive labelling of N-methylputrescine. A convenient and accurate enzyme-coupled colorimetric assay is based on the conversion of SAH to homocysteine by 5'-methylthioadenosine/ S-adenosylhomocysteine nucleosidase (MTAN/SAHN, EC 3.2.2.9) and S-ribosylhomocysteine lyase (LuxS, EC 4.4.1.21). Homocysteine is quantified by 5,5'-dithiobis-2-nitrobenzoic acid. Putrescine was shown not to interfere with MTAN or LuxS. The colorimetric assay was validated by HPLC analysis. K(m) values determined by the assay, 108 microM for putrescine and 42 microM for SAM, are lower than the previously reported values, due to alleviation of PMT inhibition by SAH. DTNB:5,5'-dithiobis-2-nitrobenzoic acid LuxS: S-ribosylhomocysteine lyase MTAN:5'-methylthioadenosine nucleosidase PMT:putrescine N-methyltransferase SAH: S-adenosyl- L-homocysteine SAM: S-adenosyl- L-methionine TNB:2-nitro-5-thiobenzoic acid.

  18. Dynamics and reactivity in Thermus aquaticus N6-adenine methyltransferase.

    Science.gov (United States)

    Aranda, Juan; Zinovjev, Kirill; Roca, Maite; Tuñón, Iñaki

    2014-11-19

    M.TaqI is a DNA methyltransferase from Thermus aquaticus that catalyzes the transfer of a methyl group from S-adenosyl-L-methionine to the N6 position of an adenine, a process described only in prokaryotes. We have used full atomistic classical molecular dynamics simulations to explore the protein-SAM-DNA ternary complex where the target adenine is flipped out into the active site. Key protein-DNA interactions established by the target adenine in the active site are described in detail. The relaxed structure was used for a combined quantum mechanics/molecular mechanics exploration of the reaction mechanism using the string method. According to our free energy calculations the reaction takes place through a stepwise mechanism where the methyl transfer precedes the abstraction of the proton from the exocyclic amino group. The methyl transfer is the rate-determining step, and the obtained free energy barrier is in good agreement with the value derived from the experimental rate constant. Two possible candidates to extract the leftover proton have been explored: a water molecule found in the active site and Asn105, a residue activated by the hydrogen bonds formed through the amide hydrogens. The barrier for the proton abstraction is smaller when Asn105 acts as a base. The reaction mechanisms can be different in other N6-DNA-methyltransferases, as determined from the exploration of the reaction mechanism in the Asn105Asp M.TaqI mutant.

  19. Quality Control Guidelines for SAM Chemical Methods

    Science.gov (United States)

    Learn more about quality control guidelines and recommendations for the analysis of samples using the chemistry methods listed in EPA's Selected Analytical Methods for Environmental Remediation and Recovery (SAM).

  20. Quality Control Guidelines for SAM Biotoxin Methods

    Science.gov (United States)

    Learn more about quality control guidelines and recommendations for the analysis of samples using the pathogen methods listed in EPA's Selected Analytical Methods for Environmental Remediation and Recovery (SAM).

  1. Quality Control Guidelines for SAM Pathogen Methods

    Science.gov (United States)

    Learn more about quality control guidelines and recommendations for the analysis of samples using the biotoxin methods listed in EPA's Selected Analytical Methods for Environmental Remediation and Recovery (SAM).

  2. Quality Control Guidelines for SAM Radiochemical Methods

    Science.gov (United States)

    Learn more about quality control guidelines and recommendations for the analysis of samples using the radiochemistry methods listed in EPA's Selected Analytical Methods for Environmental Remediation and Recovery (SAM).

  3. Crystal structures of the methyltransferase and helicase from the ZIKA 1947 MR766 Uganda strain

    Energy Technology Data Exchange (ETDEWEB)

    Bukrejewska, Malgorzata; Derewenda, Urszula; Radwanska, Malwina; Engel, Daniel A.; Derewenda, Zygmunt S.

    2017-08-15

    Two nonstructural proteins encoded byZika virusstrain MR766 RNA, a methyltransferase and a helicase, were crystallized and their structures were solved and refined at 2.10 and 2.01 Å resolution, respectively. The NS5 methyltransferase contains a boundS-adenosyl-L-methionine (SAM) co-substrate. The NS3 helicase is in the apo form. Comparison with published crystal structures of the helicase in the apo, nucleotide-bound and single-stranded RNA (ssRNA)-bound states suggests that binding of ssRNA to the helicase may occur through conformational selection rather than induced fit.

  4. Validation of SAM 2 and SAGE satellite

    Science.gov (United States)

    Kent, G. S.; Wang, P.-H.; Farrukh, U. O.; Yue, G. K.

    1987-01-01

    Presented are the results of a validation study of data obtained by the Stratospheric Aerosol and Gas Experiment I (SAGE I) and Stratospheric Aerosol Measurement II (SAM II) satellite experiments. The study includes the entire SAGE I data set (February 1979 - November 1981) and the first four and one-half years of SAM II data (October 1978 - February 1983). These data sets have been validated by their use in the analysis of dynamical, physical and chemical processes in the stratosphere. They have been compared with other existing data sets and the SAGE I and SAM II data sets intercompared where possible. The study has shown the data to be of great value in the study of the climatological behavior of stratospheric aerosols and ozone. Several scientific publications and user-oriented data summaries have appeared as a result of the work carried out under this contract.

  5. Residues in human arsenic (+3 oxidation state) methyltransferase forming potential hydrogen bond network around S-adenosylmethionine.

    Science.gov (United States)

    Li, Xiangli; Cao, Jing; Wang, Shuping; Geng, Zhirong; Song, Xiaoli; Hu, Xin; Wang, Zhilin

    2013-01-01

    Residues Tyr59, Gly78, Ser79, Met103, Gln107, Ile136 and Glu137 in human arsenic (+3 oxidation state) methyltransferase (hAS3MT) were deduced to form a potential hydrogen bond network around S-adenosylmethionine (SAM) from the sequence alignment between Cyanidioschyzon merolae arsenite S-adenosylmethyltransferase (CmArsM) and hAS3MT. Herein, seven mutants Y59A, G78A, S79A, M103A, Q107A, I136A and E137A were obtained. Their catalytic activities and conformations were characterized and models were built. Y59A and G78A were completely inactive. Only 7.0%, 10.6% and 13.8% inorganic arsenic (iAs) was transformed to monomethylated arsenicals (MMA) when M103A, Q107A and I136A were used as the enzyme. The Vmax (the maximal velocity of the reaction) values of M103A, Q107A, I136A and E137A were decreased to 8%, 22%, 15% and 50% of that of WT-hAS3MT, respectively. The KM(SAM) (the Michaelis constant for SAM) values of mutants M103A, I136A and E137A were 15.7, 8.9 and 5.1 fold higher than that of WT-hAS3MT, respectively, indicating that their affinities for SAM were weakened. The altered microenvironment of SAM and the reduced capacity of binding arsenic deduced from KM(As) (the Michaelis constant for iAs) value probably synergetically reduced the catalytic activity of Q107A. The catalytic activity of S79A was higher than that of WT despite of the higher KM(SAM) , suggesting that Ser79 did not impact the catalytic activity of hAS3MT. In short, residues Tyr59 and Gly78 significantly influenced the catalytic activity of hAS3MT as well as Met103, Ile136 and Glu137 because they were closely associated with SAM-binding, while residue Gln107 did not affect SAM-binding regardless of affecting the catalytic activity of hAS3MT. Modeling and our experimental results suggest that the adenine ring of SAM is sandwiched between Ile136 and Met103, the amide group of SAM is hydrogen bonded to Gly78 in hAS3MT and SAM is bonded to Tyr59 with van der Waals, cation-π and hydrogen bonding

  6. Residues in human arsenic (+3 oxidation state methyltransferase forming potential hydrogen bond network around S-adenosylmethionine.

    Directory of Open Access Journals (Sweden)

    Xiangli Li

    Full Text Available Residues Tyr59, Gly78, Ser79, Met103, Gln107, Ile136 and Glu137 in human arsenic (+3 oxidation state methyltransferase (hAS3MT were deduced to form a potential hydrogen bond network around S-adenosylmethionine (SAM from the sequence alignment between Cyanidioschyzon merolae arsenite S-adenosylmethyltransferase (CmArsM and hAS3MT. Herein, seven mutants Y59A, G78A, S79A, M103A, Q107A, I136A and E137A were obtained. Their catalytic activities and conformations were characterized and models were built. Y59A and G78A were completely inactive. Only 7.0%, 10.6% and 13.8% inorganic arsenic (iAs was transformed to monomethylated arsenicals (MMA when M103A, Q107A and I136A were used as the enzyme. The Vmax (the maximal velocity of the reaction values of M103A, Q107A, I136A and E137A were decreased to 8%, 22%, 15% and 50% of that of WT-hAS3MT, respectively. The KM(SAM (the Michaelis constant for SAM values of mutants M103A, I136A and E137A were 15.7, 8.9 and 5.1 fold higher than that of WT-hAS3MT, respectively, indicating that their affinities for SAM were weakened. The altered microenvironment of SAM and the reduced capacity of binding arsenic deduced from KM(As (the Michaelis constant for iAs value probably synergetically reduced the catalytic activity of Q107A. The catalytic activity of S79A was higher than that of WT despite of the higher KM(SAM , suggesting that Ser79 did not impact the catalytic activity of hAS3MT. In short, residues Tyr59 and Gly78 significantly influenced the catalytic activity of hAS3MT as well as Met103, Ile136 and Glu137 because they were closely associated with SAM-binding, while residue Gln107 did not affect SAM-binding regardless of affecting the catalytic activity of hAS3MT. Modeling and our experimental results suggest that the adenine ring of SAM is sandwiched between Ile136 and Met103, the amide group of SAM is hydrogen bonded to Gly78 in hAS3MT and SAM is bonded to Tyr59 with van der Waals, cation-π and hydrogen bonding

  7. PERANANTAN SAM CAI KONGDALAM SEJARAH CIREBON THE ROLE OF TAN SAM CAI KONGIN CIREBON HISTORY

    Directory of Open Access Journals (Sweden)

    Rusyanti Rusyanti

    2016-06-01

    ABSTRAK Keberadaan makam dan inskripsi padanisan kuno sebagai bukti arkeologis memiliki informasi yang penting dan berhubungan dengan banyak hal. Tulisan ini bertujuan melacak tokoh dan peran seorang China yang bernamaTan Sam Cai Kongyang makamnya berada di Sukalila, Cirebon. Melalui analisis arkeologi dan sejarah diketahui bahwa Tan Sam Cai Kongmerupakan tokoh sejarah yang hidup pada sekitar abad ke-17M. Gelar sebagai Tumenggung Aria Wira Cula yang tertulis pada prasasti menandakan perananya sebagai orang penting dan dekat dengan lingkungan keraton.   Kata Kunci: Cirebon, Tan Sam CaiKong, China.

  8. Floral Benzenoid Carboxyl Methyltransferases: From in Vitro to in Planta Function

    Energy Technology Data Exchange (ETDEWEB)

    Effmert,U.; Saschenbrecker, S.; Ross, J.; Negre, F.; Fraser, C.; Noel, J.; Dudareva, N.; Piechulla, B.

    2005-01-01

    Benzenoid carboxyl methyltransferases synthesize methyl esters (e.g., methyl benzoate and methyl salicylate), which are constituents of aromas and scents of many plant species and play important roles in plant communication with the surrounding environment. Within the past five years, eleven such carboxyl methyltransferases were isolated and most of them were comprehensively investigated at the biochemical, molecular and structural level. Two types of enzymes can be distinguished according to their substrate preferences: the SAMT-type enzymes isolated from Clarkia breweri, Stephanotis floribunda, Antirrhinum majus, Hoya carnosa, and Petunia hybrida, which have a higher catalytic efficiency and preference for salicylic acid, while BAMT-type enzymes from A. majus, Arabidopsis thaliana, Arabidopsis lyrata, and Nicotiana suaveolens prefer benzoic acid. The elucidation of C. breweri SAMT's three-dimensional structure allowed a detailed modelling of the active sites of the carboxyl methyltransferases and revealed that the SAM binding pocket is highly conserved among these enzymes while the methyl acceptor binding site exhibits some variability, allowing a classification into SAMT-type and BAMT-type enzymes. The analysis of expression patterns coupled with biochemical characterization showed that these carboxyl methyltransferases are involved either in floral scent biosynthesis or in plant defense responses. While the latter can be induced by biotic or abiotic stress, the genes responsible for floral scent synthesis exhibit developmental and rhythmic expression pattern. The nature of the product and efficiency of its formation in plants depend on the availability of substrates, the catalytic efficiency of the enzyme toward benzoic acid and/or salicylic acid, and the transcriptional, translational, and post-translational regulation at the enzyme level. The biochemical properties of benzenoid carboxyl methyltransferases suggest that the genes involved in plant defenses

  9. Samâ’ dalam Tradisi Tasawuf

    Directory of Open Access Journals (Sweden)

    Said Aqil Siradj

    2014-01-01

    Full Text Available Samâ‘ in Tasawuf has been a very important element in the dissemination of this spiritual dimension of Islam. Yet, it has received very little both from the practitioners of Tasawuf and its intellectuals. This paper tries to expose this simply in a hope to make it heard in the academic and popular circle. Here, samâ‘ is not only understood as a form of music, as many would do, but also as an art of listening of which music is certainly part. The paper will explore the meaning and definition of this term, putting emphasis on its many-faceted function in the formation and development of one’s soul and spirituality. It is argued that soul is musical and artistic. Using art and music to talk to soul is therefore the proper way and means. The paper will also try to show that samâ‘ is also an indispensable part of spiritual method to reach and know God. Knowledge of God in other words, can be gained through this practice. Hence, samâ‘ is treated not only as a form of entertainment, but also a kind of practical epistemology.

  10. Essay: Physical Review Letters; Sam Goudsmit's Vision.

    Science.gov (United States)

    Adair, Robert K

    2008-01-18

    Sam Goudsmit implemented his vision of converting the Letters section of Physical Review into a distinct journal fifty years ago. Physical Review Letters was designed to publish "only papers that really deserve rapid communication." The new journal became so successful with physicists throughout the world that Physical Review Letters now publishes 3500 Letters per year.

  11. Roles of DNA methyltransferases in Arabidopsis development

    African Journals Online (AJOL)

    Jane

    2010-12-13

    Dec 13, 2010 ... RNA-dependent tRNA polymerase; ROS1, repressor of silencing 1; SET, histone methyltransferase; SDC, suppressor of drm1/2 cmt3; siRNA, small interfering RNA; SUP, superman;. SUVH4, su(var)3-9 homolog 4; TRDMT1, tRNA aspartic acid methyltransferase 1; TGS, transcriptional gene silencing; UBA,.

  12. Roles of DNA methyltransferases in Arabidopsis development ...

    African Journals Online (AJOL)

    Mutations that cause severe loss of DNA methylation often leads to abnormal development. In the present review, we summarized recent findings of the three major DNA methyltransferases mutants playing vital role in development of Arabidopsis thaliana. Keywords: DNA methylation, epigenetics, methyltransferase, mutant ...

  13. Metal/SAM/metal junctions studied from first principles

    Energy Technology Data Exchange (ETDEWEB)

    Kucera, Jan; Gross, Axel [Institute for Theoretical Chemistry, Ulm University, D-89069 Ulm (Germany)

    2010-07-01

    There is currently a strong interest in sandwich structures made of self-assembled monolayers (SAMs) between two metal layers (M/SAM/M) because of their potential in future nanoelectronics. Recently, a M/SAM/M/SAM/M ''double decker'' junction was realized with two 4-mercaptopyridine SAMs sandwiched between an Au electrode and Pd and Pt monolayers. We have performed periodic electronic structure calculations based on density function theory (DFT) to describe various phenomena playing a role in hybrid systems such as Au/SAM, Au/SAM/Pd, and Au/SAM/Pd/SAM/Pt with the SAMs consisting of the bifunctional thioaromatics 4-aminothiophenol or 4-mercaptopyridine. Based on the calculated local density of states (LDOS) of the metal layers, possible explanations for experimental results will be provided which show a reduced DOS at the Fermi energy. In addition, the role of van der Waals forces on the structure and stability of the SAMs has been addressed by performing DFT calculations with semi-empirical corrections for dispersion effects.

  14. Serotonin-Induced Hypersensitivity via Inhibition of Catechol O-Methyltransferase Activity

    Science.gov (United States)

    2012-01-01

    The subcutaneous and systemic injection of serotonin reduces cutaneous and visceral pain thresholds and increases responses to noxious stimuli. Different subtypes of 5-hydroxytryptamine (5-HT) receptors are suggested to be associated with different types of pain responses. Here we show that serotonin also inhibits catechol O-methyltransferase (COMT), an enzyme that contributes to modultion the perception of pain, via non-competitive binding to the site bound by catechol substrates with a binding affinity comparable to the binding affinity of catechol itself (Ki = 44 μM). Using computational modeling, biochemical tests and cellular assays we show that serotonin actively competes with the methyl donor S-adenosyl-L-methionine (SAM) within the catalytic site. Binding of serotonin to the catalytic site inhibits the access of SAM, thus preventing methylation of COMT substrates. The results of in vivo animal studies show that serotonin-induced pain hypersensitivity in mice is reduced by either SAM pretreatment or by the combined administration of selective antagonists for β2- and β3-adrenergic receptors, which have been previously shown to mediate COMT-dependent pain signaling. Our results suggest that inhibition of COMT via serotonin binding contributes to pain hypersensitivity, providing additional strategies for the treatment of clinical pain conditions. PMID:22500608

  15. Serotonin-Induced Hypersensitivity via Inhibition of Catechol O-Methyltransferase Activity

    Directory of Open Access Journals (Sweden)

    Tsao Douglas

    2012-04-01

    Full Text Available Abstract The subcutaneous and systemic injection of serotonin reduces cutaneous and visceral pain thresholds and increases responses to noxious stimuli. Different subtypes of 5-hydroxytryptamine (5-HT receptors are suggested to be associated with different types of pain responses. Here we show that serotonin also inhibits catechol O-methyltransferase (COMT, an enzyme that contributes to modultion the perception of pain, via non-competitive binding to the site bound by catechol substrates with a binding affinity comparable to the binding affinity of catechol itself (Ki = 44 μM. Using computational modeling, biochemical tests and cellular assays we show that serotonin actively competes with the methyl donor S-adenosyl-L-methionine (SAM within the catalytic site. Binding of serotonin to the catalytic site inhibits the access of SAM, thus preventing methylation of COMT substrates. The results of in vivo animal studies show that serotonin-induced pain hypersensitivity in mice is reduced by either SAM pretreatment or by the combined administration of selective antagonists for β2- and β3-adrenergic receptors, which have been previously shown to mediate COMT-dependent pain signaling. Our results suggest that inhibition of COMT via serotonin binding contributes to pain hypersensitivity, providing additional strategies for the treatment of clinical pain conditions.

  16. On LAM's and SAM's for Halley's rotation

    Science.gov (United States)

    Peale, Stanton J.

    1992-01-01

    Non principal axis rotation for comet Halley is inferred from dual periodicities evident in the observations. The modes where the spin axis precesses around the axis of minimum moment of inertia (long axis mode or LAM) and where it precesses around the axis of maximum moment of inertia (short axis mode or SAM) are described from an inertial point of view. The currently favored LAM model for Halley's rotation state satisfies observational and dynamical constraints that apparently no SAM can satisfy. But it cannot reproduce the observed post perihelion brightening through seasonal illumination of localized sources on the nucleus, whereas a SAM can easily produce post or pre perihelion brightening by this mechanism. However, the likelihood of a LAM rotation for elongated nuclei of periodic comets such as Halley together with Halley's extreme post perihelion behavior far from the Sun suggest that Halley's post perihelion brightening may be due to effects other than seasonal illumination of localized sources, and therefore such brightening may not constrain its rotation state.

  17. A comprehensive evaluation of SAM, the SAM R-package and a simple modification to improve its performance

    Directory of Open Access Journals (Sweden)

    Zhang Shunpu

    2007-06-01

    Full Text Available Abstract Background The Significance Analysis of Microarrays (SAM is a popular method for detecting significantly expressed genes and controlling the false discovery rate (FDR. Recently, it has been reported in the literature that the FDR is not well controlled by SAM. Due to the vast application of SAM in microarray data analysis, it is of great importance to have an extensive evaluation of SAM and its associated R-package (sam2.20. Results Our study has identified several discrepancies between SAM and sam2.20. One major difference is that SAM and sam2.20 use different methods for estimating FDR. Such discrepancies may cause confusion among the researchers who are using SAM or are developing the SAM-like methods. We have also shown that SAM provides no meaningful estimates of FDR and this problem has been corrected in sam2.20 by using a different formula for estimating FDR. However, we have found that, even with the improvement sam2.20 has made over SAM, sam2.20 may still produce erroneous and even conflicting results under certain situations. Using an example, we show that the problem of sam2.20 is caused by its use of asymmetric cutoffs which are due to the large variability of null scores at both ends of the order statistics. An obvious approach without the complication of the order statistics is the conventional symmetric cutoff method. For this reason, we have carried out extensive simulations to compare the performance of sam2.20 and the symmetric cutoff method. Finally, a simple modification is proposed to improve the FDR estimation of sam2.20 and the symmetric cutoff method. Conclusion Our study shows that the most serious drawback of SAM is its poor estimation of FDR. Although this drawback has been corrected in sam2.20, the control of FDR by sam2.20 is still not satisfactory. The comparison between sam2.20 and the symmetric cutoff method reveals that the relative performance of sam2.20 to the symmetric cutff method depends on the

  18. System Advisor Model, SAM 2014.1.14: General Description

    Energy Technology Data Exchange (ETDEWEB)

    Blair, Nate [National Renewable Energy Lab. (NREL), Golden, CO (United States); Dobos, Aron P. [National Renewable Energy Lab. (NREL), Golden, CO (United States); Freeman, Janine [National Renewable Energy Lab. (NREL), Golden, CO (United States); Neises, Ty [National Renewable Energy Lab. (NREL), Golden, CO (United States); Wagner, Michael [National Renewable Energy Lab. (NREL), Golden, CO (United States); Ferguson, Tom [Global Resources, Northbrook, IL (United States); Gilman, Paul [National Renewable Energy Lab. (NREL), Golden, CO (United States); Janzou, Steven [Janzou Consulting, Idaho Springs, CO (United States)

    2014-02-01

    This document describes the capabilities of the U.S. Department of Energy and National Renewable Energy Laboratory's System Advisor Model (SAM), Version 2013.9.20, released on September 9, 2013. SAM is a computer model that calculates performance and financial metrics of renewable energy systems. Project developers, policy makers, equipment manufacturers, and researchers use graphs and tables of SAM results in the process of evaluating financial, technology, and incentive options for renewable energy projects. SAM simulates the performance of photovoltaic, concentrating solar power, solar water heating, wind, geothermal, biomass, and conventional power systems. The financial model can represent financial structures for projects that either buy and sell electricity at retail rates (residential and commercial) or sell electricity at a price determined in a power purchase agreement (utility). SAM's advanced simulation options facilitate parametric and sensitivity analyses, and statistical analysis capabilities are available for Monte Carlo simulation and weather variability (P50/P90) studies. SAM can also read input variables from Microsoft Excel worksheets. For software developers, the SAM software development kit (SDK) makes it possible to use SAM simulation modules in their applications written in C/C++, C#, Java, Python, and MATLAB. NREL provides both SAM and the SDK as free downloads at http://sam.nrel.gov. Technical support and more information about the software are available on the website.

  19. Genetics Home Reference: guanidinoacetate methyltransferase deficiency

    Science.gov (United States)

    ... literature. Of these, approximately one-third are of Portuguese origin. Related Information What information about a genetic ... require large amounts of energy, especially the brain. Learn more about the gene associated with guanidinoacetate methyltransferase ...

  20. General Quality Control (QC) Guidelines for SAM Methods

    Science.gov (United States)

    Learn more about quality control guidelines and recommendations for the analysis of samples using the methods listed in EPA's Selected Analytical Methods for Environmental Remediation and Recovery (SAM).

  1. A histone H3K9M mutation traps histone methyltransferase Clr4 to prevent heterochromatin spreading

    Energy Technology Data Exchange (ETDEWEB)

    Shan, Chun-Min; Wang, Jiyong; Xu, Ke; Chen, Huijie; Yue, Jia-Xing; Andrews, Stuart; Moresco, James J.; Yates, John R.; Nagy, Peter L.; Tong, Liang; Jia, Songtao

    2016-09-20

    Histone lysine-to-methionine (K-to-M) mutations are associated with multiple cancers, and they function in a dominant fashion to block the methylation of corresponding lysines on wild type histones. However, their mechanisms of function are controversial. Here we show that in fission yeast, introducing the K9M mutation into one of the three histone H3 genes dominantly blocks H3K9 methylation on wild type H3 across the genome. In addition, H3K9M enhances the interaction of histone H3 tail with the H3K9 methyltransferase Clr4 in a SAM (S-adenosyl-methionine)-dependent manner, and Clr4 is trapped at nucleation sites to prevent its spreading and the formation of large heterochromatin domains. We further determined the crystal structure of an H3K9M peptide in complex with human H3K9 methyltransferase G9a and SAM, which reveales that the methionine side chain had enhanced van der Waals interactions with G9a. Therefore, our results provide a detailed mechanism by which H3K9M regulates H3K9 methylation.

  2. Storifying Samsøs Renewable Energy Transition

    DEFF Research Database (Denmark)

    Papazu, Irina

    2017-01-01

    Through a joint community effort Denmark’s Renewable Energy Island Samsø became self-sufficient with renewable energy over a period of 10 years from 1997 to 2007. Today, the story about Samsø’s successful energy transition has become a global export and a widely known model of community building......, public participation and shared ownership in renewable energy technologies and transition processes. What has allowed the Samsø narrative to travel so widely has been the effective ‘transition story’ created about the islanders’ efforts. This transition story, however, has become fixed with the years...... of further local energy transitions, as challenging elements of the process are downplayed to strengthen the narrative power of the story. Ethnographic stories about Samsø complicate the island’s transition narrative and add nuance to the Samsø story, highlighting its discrepancies and problematizing...

  3. Providing output of DIRAC-SAM jobs to the IT-based SAM-Nagios framework

    CERN Document Server

    Volkl, Valentin

    2013-01-01

    Information available on LHCb grid sites in the SAM-Nagios monitor- ing framework - gathered mainly through functional tests - has been sup- plemented with results from LHCbDIRAC SAMJobs published by means of message client newly integrated in LHCbDIRAC. These are displayed as a new metric org.lhcb.DiracTest-lhcb giving additional debug in- formation to system administrators and influencing reports on grid site performances in the future

  4. Catalysis in glycine N-methyltransferase: testing the electrostatic stabilization and compression hypothesis.

    Science.gov (United States)

    Soriano, Alejandro; Castillo, Raquel; Christov, Christo; Andrés, Juan; Moliner, Vicente; Tuñón, Iñaki

    2006-12-19

    Glycine N-methyltransferase (GNMT) is an S-adenosyl-l-methionine dependent enzyme that catalyzes glycine transformation to sarcosine. Here, we present a hybrid quantum mechanics/molecular mechanics (QM/MM) computational study of the reaction compared to the counterpart process in water. The process takes place through an SN2 mechanism in both media with a transition state in which the transferring methyl group is placed in between the donor (SAM) and the acceptor (the amine group of glycine). Comparative analysis of structural, electrostatic, and electronic characteristics of the in-solution and enzymatic transition states allows us to get a deeper insight into the origins of the enzyme's catalytic power. We found that the enzyme is able to stabilize the substrate in its more active basic form by means of a positively charged residue (Arg175) placed in the active site. However, the maximum stabilization is attained for the transition state. In this case, the enzyme is able to form stronger hydrogen bonds with the positively charged amine group. Finally, we show that in agreement with previous computational studies on other methyltransferases, there is no computational evidence for the compression hypothesis, as was formulated by Schowen (Hegazi, M. F., Borchardt, R. T., and Schowen, R. L. (1979) J. Am. Chem. Soc. 101, 4359-4365).

  5. Structural Characterization of the Mitomycin 7-O-Methyltransferase MmcR

    Science.gov (United States)

    Singh, Shanteri; Chang, Aram; Goff, Randal D.; Bingman, Craig A.; Grüschow, Sabine; Sherman, David H.; Phillips, George N.; Thorson, Jon S.

    2011-01-01

    Mitomycins are quinone-containing antibiotics, widely used as anti-tumor drugs in chemotherapy. Mitomycin-7-O-methyltransferase (MmcR), a key tailoring enzyme involved in the biosynthesis of mitomycin in Streptomyces lavendulae, catalyzes the 7-O-methylation of both C9β- and C9α-configured 7-hydroxymitomycins. We have determined the crystal structures of the MmcR–S-adenosylhomocysteine (SAH) binary complex and MmcR–SAH-Mitomycin A (MMA) ternary complex at resolutions of 1.9 and 2.3 Å, respectively. The study revealed MmcR to adopt a common SAM-dependent O-MTase fold and the presence of a structurally-conserved active site general acid-base pair is consistent with a proton assisted methyltransfer common to most methyltransferases. Given the importance of C7 alkylation to modulate mitomycin redox potential this study may also present a template toward the future engineering of catalysts to generate uniquely bioactive mitomycins. PMID:21538548

  6. An In Silico Approach for Characterization of an Aminoglycoside Antibiotic-Resistant Methyltransferase Protein from Pyrococcus furiosus (DSM 3638).

    Science.gov (United States)

    Oany, Arafat Rahman; Jyoti, Tahmina Pervin; Ahmad, Shah Adil Ishtiyaq

    2014-01-01

    Pyrococcus furiosus is a hyperthermophilic archaea. A hypothetical protein of this archaea, PF0847, was selected for computational analysis. Basic local alignment search tool and multiple sequence alignment (MSA) tool were employed to search for related proteins. Both the secondary and tertiary structure prediction were obtained for further analysis. Three-dimensional model was assessed by PROCHECK and QMEAN6 programs. To get insights about the physical and functional associations of the protein, STRING network analysis was performed. Binding of the SAM (S-adenosyl-l-methionine) ligand with our protein, fetched from an antibiotic-related methyltransferase (PDB code: 3P2K: D), showed high docking energy and suggested the function of the protein as methyltransferase. Finally, we tried to look for a specific function of the proposed methyltransferase, and binding of the geneticin bound to the eubacterial 16S rRNA A-site (PDB code: 1MWL) in the active site of the PF0847 gave us the indication to predict the protein responsible for aminoglycoside antibiotic resistance.

  7. Crystal structures of human 108V and 108M catechol O-methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Rutherford, K.; Le Trong, I.; Stenkamp, R.E.; Parson, W.W. (UWASH)

    2008-08-01

    Catechol O-methyltransferase (COMT) plays important roles in the metabolism of catecholamine neurotransmitters and catechol estrogens. The development of COMT inhibitors for use in the treatment of Parkinson's disease has been aided by crystallographic structures of the rat enzyme. However, the human and rat proteins have significantly different substrate specificities. Additionally, human COMT contains a common valine-methionine polymorphism at position 108. The methionine protein is less stable than the valine polymorph, resulting in decreased enzyme activity and protein levels in vivo. Here we describe the crystal structures of the 108V and 108M variants of the soluble form of human COMT bound with S-adenosylmethionine (SAM) and a substrate analog, 3,5-dinitrocatechol. The polymorphic residue 108 is located in the {alpha}5-{beta}3 loop, buried in a hydrophobic pocket {approx}16 {angstrom} from the SAM-binding site. The 108V and 108M structures are very similar overall [RMSD of C{sup {alpha}} atoms between two structures (C{sup {alpha}} RMSD) = 0.2 {angstrom}], and the active-site residues are superposable, in accord with the observation that SAM stabilizes 108M COMT. However, the methionine side chain is packed more tightly within the polymorphic site and, consequently, interacts more closely with residues A22 ({alpha}2) and R78 ({alpha}4) than does valine. These interactions of the larger methionine result in a 0.7-{angstrom} displacement in the backbone structure near residue 108, which propagates along {alpha}1 and {alpha}5 toward the SAM-binding site. Although the overall secondary structures of the human and rat proteins are very similar (C{sup {alpha}} RMSD = 0.4 {angstrom}), several nonconserved residues are present in the SAM-(I89M, I91M, C95Y) and catechol- (C173V, R201M, E202K) binding sites. The human protein also contains three additional solvent-exposed cysteine residues (C95, C173, C188) that may contribute to intermolecular disulfide bond

  8. Evolution of the key alkaloid enzyme putrescine N-methyltransferase from spermidine synthase

    Science.gov (United States)

    Junker, Anne; Fischer, Juliane; Sichhart, Yvonne; Brandt, Wolfgang; Dräger, Birgit

    2013-01-01

    Putrescine N-methyltransferases (PMTs) are the first specific enzymes of the biosynthesis of nicotine and tropane alkaloids. PMTs transfer a methyl group onto the diamine putrescine from S-adenosyl-l-methionine (SAM) as coenzyme. PMT proteins have presumably evolved from spermidine synthases (SPDSs), which are ubiquitous enzymes of polyamine metabolism. SPDSs use decarboxylated SAM as coenzyme to transfer an aminopropyl group onto putrescine. In an attempt to identify possible and necessary steps in the evolution of PMT from SPDS, homology based modeling of Datura stramonium SPDS1 and PMT was employed to gain deeper insight in the preferred binding positions and conformations of the substrate and the alternative coenzymes. Based on predictions of amino acids responsible for the change of enzyme specificities, sites of mutagenesis were derived. PMT activity was generated in D. stramonium SPDS1 after few amino acid exchanges. Concordantly, Arabidopsis thaliana SPDS1 was mutated and yielded enzymes with both, PMT and SPDS activities. Kinetic parameters were measured for enzymatic characterization. The switch from aminopropyl to methyl transfer depends on conformational changes of the methionine part of the coenzyme in the binding cavity of the enzyme. The rapid generation of PMT activity in SPDS proteins and the wide-spread occurrence of putative products of N-methylputrescine suggest that PMT activity is present frequently in the plant kingdom. PMID:23908659

  9. Evolution of the key alkaloid enzyme putrescine N-methyltransferase from spermidine synthase.

    Directory of Open Access Journals (Sweden)

    Anne eJunker

    2013-07-01

    Full Text Available Putrescine N-methyltransferases (PMTs are the first specific enzymes of the biosynthesis of nicotine and tropane alkaloids. PMTs transfer a methyl group onto the diamine putrescine from S-adenosyl-L-methionine (SAM as coenzyme. PMT proteins have presumably evolved from spermidine synthases (SPDSs, which are ubiquitous enzymes of polyamine metabolism. SPDS use decarboxylated SAM as coenzyme to transfer an aminopropyl group onto putrescine. In an attempt to identify possible and necessary steps in the evolution of PMT from SPDS, homology based modeling of Datura stramonium SPDS1 and PMT was employed to gain deeper insight in the preferred binding positions and conformations of the substrate and the alternative coenzymes. Based on predictions of amino acids responsible for the change of enzyme specificities, sites of mutagenesis were derived. PMT activity was generated in Datura stramonium SPDS1 after few amino acid exchanges. Concordantly, Arabidopsis thaliana SPDS1 was mutated and yielded enzymes with both, PMT and SPDS activities. Kinetic parameters were measured for enzymatic characterization. The switch from aminopropyl to methyl transfer depends on conformational changes of the methionine part of the coenzyme in the binding cavity of the enzyme. The rapid generation of PMT activity in SPDS proteins and the wide-spread occurrence of putative products of N-methylputrescine suggest that PMT activity is present frequently in the plant kingdom.

  10. Chemical Probes of Histone Lysine Methyltransferases

    Science.gov (United States)

    2015-01-01

    Growing evidence suggests that histone methyltransferases (HMTs, also known as protein methyltransferases (PMTs)) play an important role in diverse biological processes and human diseases by regulating gene expression and the chromatin state. Therefore, HMTs have been increasingly recognized by the biomedical community as a class of potential therapeutic targets. High quality chemical probes of HMTs, as tools for deciphering their physiological functions and roles in human diseases and testing therapeutic hypotheses, are critical for advancing this promising field. In this review, we focus on the discovery, characterization, and biological applications of chemical probes for HMTs. PMID:25423077

  11. Elp3 and RlmN: A tale of two mitochondrial tail-anchored radical SAM enzymes in Toxoplasma gondii.

    Science.gov (United States)

    Padgett, Leah R; Lentini, Jenna M; Holmes, Michael J; Stilger, Krista L; Fu, Dragony; Sullivan, William J

    2018-01-01

    Radical S-adenosylmethionine (rSAM) enzymes use a 5'-deoxyadensyl 5'-radical to methylate a wide array of diverse substrates including proteins, lipids and nucleic acids. One such enzyme, Elongator protein-3 (TgElp3), is an essential protein in Toxoplasma gondii, a protozoan parasite that can cause life-threatening opportunistic disease. Unlike Elp3 homologues which are present in all domains of life, TgElp3 localizes to the outer mitochondrial membrane (OMM) via a tail-anchored trafficking mechanism in Toxoplasma. Intriguingly, we identified a second tail-anchored rSAM domain containing protein (TgRlmN) that also localizes to the OMM. The transmembrane domain (TMD) on Toxoplasma Elp3 and RlmN homologues is required for OMM localization and has not been seen beyond the chromalveolates. Both TgElp3 and TgRlmN contain the canonical rSAM amino acid sequence motif (CxxxCxxC) necessary to form the 4Fe-4S cluster required for tRNA modifications. In E. coli, RlmN is responsible for the 2-methlyadenosine (m2A) synthesis at purine 37 in tRNA while in S. cerevisiae, Elp3 is necessary for the formation of 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U) at the wobble tRNA position. To investigate why these two rSAM enzymes localize to the mitochondrion in Toxoplasma, and whether or not TgRlmN and TgElp3 possess tRNA methyltransferase activity, a series of mutational and biochemical studies were performed. Overexpression of either TgElp3 or TgRlmN resulted in a significant parasite replication defect, but overexpression was tolerated if either the TMD or rSAM domain was mutated. Furthermore, we show the first evidence that Toxoplasma tRNAGlu contains the mcm5s2U modification, which is the putative downstream product generated by TgElp3 activity.

  12. Nicotinamide N-methyltransferase knockdown protects against diet-induced obesity

    Science.gov (United States)

    Kraus, Daniel; Yang, Qin; Kong, Dong; Banks, Alexander S.; Zhang, Lin; Rodgers, Joseph T.; Pirinen, Eija; Pulinilkunnil, Thomas C.; Gong, Fengying; Wang, Ya-chin; Cen, Yana; Sauve, Anthony A.; Asara, John M.; Peroni, Odile D.; Monia, Brett P.; Bhanot, Sanjay; Alhonen, Leena; Puigserver, Pere; Kahn, Barbara B.

    2014-01-01

    In obesity and type 2 diabetes, Glut4 glucose transporter expression is decreased selectively in adipocytes1. Adipose-specific knockout or overexpression of Glut4 alters systemic insulin sensitivity2. Here we show, using DNA array analyses, that nicotinamide N-methyltransferase (Nnmt) is the most strongly reciprocally regulated gene when comparing gene expression in white adipose tissue (WAT) from adipose-specific Glut4-knockout or adipose-specific Glut4-overexpressing mice with their respective controls. NNMT methylates nicotinamide (vitamin B3) using S-adenosylmethionine (SAM) as a methyl donor3,4. Nicotinamide is a precursor of NAD+, an important cofactor linking cellular redox states with energy metabolism5. SAM provides propylamine for polyamine biosynthesis and donates a methyl group for histone methylation6. Polyamine flux including synthesis, catabolism and excretion, is controlled by the rate-limiting enzymes ornithine decarboxylase (ODC) and spermidine–spermine N1-acetyltransferase (SSAT; encoded by Sat1) and by polyamine oxidase (PAO), and has a major role in energy metabolism7,8. We report that NNMT expression is increased in WAT and liver of obese and diabetic mice. Nnmt knockdown in WAT and liver protects against diet-induced obesity by augmenting cellular energy expenditure. NNMT inhibition increases adipose SAM and NAD+ levels and upregulates ODC and SSAT activity as well as expression, owing to the effects of NNMT on histone H3 lysine 4 methylation in adipose tissue. Direct evidence for increased polyamine flux resulting from NNMT inhibition includes elevated urinary excretion and adipocyte secretion of diacetylspermine, a product of polyamine metabolism. NNMT inhibition in adipocytes increases oxygen consumption in an ODC-, SSAT- and PAO-dependent manner. Thus, NNMT is a novel regulator of histone methylation, polyamine flux and NAD+-dependent SIRT1 signalling, and is a unique and attractive target for treating obesity and type 2 diabetes. PMID

  13. Spirit Pluralisme dalam Klenteng Sam Po Kong Semarang

    Directory of Open Access Journals (Sweden)

    Edi Nurwahyu Julianto

    2015-07-01

    Full Text Available Klenteng Sam Po Kong has very deep meaning as a symbol of multi cultural; multi ethnic and multi religious. Klenteng Sam Po Kong has a different function, not only used by people with background religious Tri Dharma (Budha, Tao and Konghuchu, but also used by Javanese ethnic with different religious backgrounds. Between ethnic China and Java, mutual respect and tolerance run beliefs and rituals of each. More over, Klenteng Sam Po Kong is a form of pluralism which reflected the fact Sino Javanese Muslim Culture is preserved to date both of sightings physical culture and system cultural in the form of religious rites performed by ethnic China and Java.

  14. Selective and membrane-permeable small molecule inhibitors of nicotinamide N-methyltransferase reverse high fat diet-induced obesity in mice.

    Science.gov (United States)

    Neelakantan, Harshini; Vance, Virginia; Wetzel, Michael D; Wang, Hua-Yu Leo; McHardy, Stanton F; Finnerty, Celeste C; Hommel, Jonathan D; Watowich, Stanley J

    2018-01-01

    There is a critical need for new mechanism-of-action drugs that reduce the burden of obesity and associated chronic metabolic comorbidities. A potentially novel target to treat obesity and type 2 diabetes is nicotinamide-N-methyltransferase (NNMT), a cytosolic enzyme with newly identified roles in cellular metabolism and energy homeostasis. To validate NNMT as an anti-obesity drug target, we investigated the permeability, selectivity, mechanistic, and physiological properties of a series of small molecule NNMT inhibitors. Membrane permeability of NNMT inhibitors was characterized using parallel artificial membrane permeability and Caco-2 cell assays. Selectivity was tested against structurally-related methyltransferases and nicotinamide adenine dinucleotide (NAD + ) salvage pathway enzymes. Effects of NNMT inhibitors on lipogenesis and intracellular levels of metabolites, including NNMT reaction product 1-methylnicotianamide (1-MNA) were evaluated in cultured adipocytes. Effects of a potent NNMT inhibitor on obesity measures and plasma lipid were assessed in diet-induced obese mice fed a high-fat diet. Methylquinolinium scaffolds with primary amine substitutions displayed high permeability from passive and active transport across membranes. Importantly, methylquinolinium analogues displayed high selectivity, not inhibiting related SAM-dependent methyltransferases or enzymes in the NAD + salvage pathway. NNMT inhibitors reduced intracellular 1-MNA, increased intracellular NAD + and S-(5'-adenosyl)-l-methionine (SAM), and suppressed lipogenesis in adipocytes. Treatment of diet-induced obese mice systemically with a potent NNMT inhibitor significantly reduced body weight and white adipose mass, decreased adipocyte size, and lowered plasma total cholesterol levels. Notably, administration of NNMT inhibitors did not impact total food intake nor produce any observable adverse effects. These results support development of small molecule NNMT inhibitors as therapeutics to

  15. Putative DNA modification methylase DR_C0020 of Deinococcus radiodurans is an atypical SAM dependent C-5 cytosine DNA methylase.

    Science.gov (United States)

    Patil, Nayana A; Basu, Bhakti; Deobagkar, Deepti D; Apte, Shree K; Deobagkar, Dileep N

    2017-03-01

    Control of cellular processes by epigenetic modification of cytosine in DNA is widespread among living organisms, but, is hitherto unknown in the extremely radioresistant microbe D. radiodurans. C-5 methyl cytosines (m(5)C) were detected by immuno-blotting with m(5)C-specific antibody. Site of cytosine methylation by DR_C0020 encoded protein was investigated by bisulfite sequencing. The DR_C0020 knockout mutant (Δdcm), constructed by site directed mutagenesis, was assessed for effect on growth, radiation resistance and proteome. Proteins were identified by mass spectrometry. Methylated cytosines were detected in the D. radiodurans genome. The DR_C0020 encoded protein (Dcm, NCBI accession: WP_034351354.1), whose amino acid sequence resembles m(4)C methylases, was shown to be the lone SAM-dependent C-5 cytosine methyltransferase. Purified Dcm protein was found to methylate CpN sequence with a preference for methylation of two consecutive cytosines. The Δdcm strain completely lost m(5)C modification from its genome, had no effect on growth but became radiation sensitive. The Δdcm cells exhibited minor alterations in the abundance of several proteins involved primarily in protein homeostasis, oxidative stress defense, metabolism, etc. DR_C0020 encoded SAM-dependent methyltransferase Dcm is solely responsible for C-5cytosine methylation at CpN sites in the genome of D. radiodurans and regulates protein homeostasis under normal growth conditions. The protein is an unusual case of an amino methyltransferase that has evolved to producing m(5)C. Although, dispensable under optimal growth conditions, the presence of m(5)C may be important for recognition of parent strand and, thus, could contribute to the extraordinary DNA repair in D. radiodurans. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Crystal structures of the SAM-III/S[subscript MK] riboswitch reveal the SAM-dependent translation inhibition mechanism

    Energy Technology Data Exchange (ETDEWEB)

    Lu, C.; Smith, A.M.; Fuchs, R.T.; Ding, F.; Rajashankar, K.; Henkin, T.M.; Ke, A. (Cornell); (OSU)

    2010-01-07

    Three distinct classes of S-adenosyl-L-methionine (SAM)-responsive riboswitches have been identified that regulate bacterial gene expression at the levels of transcription attenuation or translation inhibition. The SMK box (SAM-III) translational riboswitch has been identified in the SAM synthetase gene in members of the Lactobacillales. Here we report the 2.2-{angstrom} crystal structure of the Enterococcus faecalis SMK box riboswitch. The Y-shaped riboswitch organizes its conserved nucleotides around a three-way junction for SAM recognition. The Shine-Dalgarno sequence, which is sequestered by base-pairing with the anti-Shine-Dalgarno sequence in response to SAM binding, also directly participates in SAM recognition. The riboswitch makes extensive interactions with the adenosine and sulfonium moieties of SAM but does not appear to recognize the tail of the methionine moiety. We captured a structural snapshot of the SMK box riboswitch sampling the near-cognate ligand S-adenosyl-L-homocysteine (SAH) in which SAH was found to adopt an alternative conformation and fails to make several key interactions.

  17. Selective inhibitors of bacterial t-RNA-(N(1)G37) methyltransferase (TrmD) that demonstrate novel ordering of the lid domain.

    Science.gov (United States)

    Hill, Pamela J; Abibi, Ayome; Albert, Robert; Andrews, Beth; Gagnon, Moriah M; Gao, Ning; Grebe, Tyler; Hajec, Laurel I; Huang, Jian; Livchak, Stephania; Lahiri, Sushmita D; McKinney, David C; Thresher, Jason; Wang, Hongming; Olivier, Nelson; Buurman, Ed T

    2013-09-26

    The tRNA-(N(1)G37) methyltransferase (TrmD) is essential for growth and highly conserved in both Gram-positive and Gram-negative bacterial pathogens. Additionally, TrmD is very distinct from its human orthologue TRM5 and thus is a suitable target for the design of novel antibacterials. Screening of a collection of compound fragments using Haemophilus influenzae TrmD identified inhibitory, fused thieno-pyrimidones that were competitive with S-adenosylmethionine (SAM), the physiological methyl donor substrate. Guided by X-ray cocrystal structures, fragment 1 was elaborated into a nanomolar inhibitor of a broad range of Gram-negative TrmD isozymes. These compounds demonstrated no activity against representative human SAM utilizing enzymes, PRMT1 and SET7/9. This is the first report of selective, nanomolar inhibitors of TrmD with demonstrated ability to order the TrmD lid in the absence of tRNA.

  18. SAM Chlorine Observations at Gale Crater

    Science.gov (United States)

    Conrad, P. G.; Farley, K. A.; Vasconcelos, P. M.; Malespin, C.; Franz, H.; McAdam, A.; Sutter, B.; Stern, J. C.; Clark, B. C.; Atreya, S. K.; Mahaffy, P. R.; Martín-Torres, J.; Zorzano, M. P.

    2014-12-01

    The Sample Analysis at Mars investigation has detected Cl-bearing phases of various oxidation states in its thermally evolved gas measurements of both a wind drift deposit of fines and three different rock samples delivered as sieved drill powders to the instrument suite. In addition to HCl (Leshin et al, 2013; Ming et al, 2013) and chlorinated hydrocarbon detections (Glavin et al, 2013; Freissinet et al, in review), oxygen releases consistent with the decomposition of perchlorate salts are also observed. We have also measured chlorine isotope ratios of the four different solid samples, which yielded variable and more negative δ37Cl than typically observed in SNC meteorite analyses. We summarize our chlorine observations in the context of other gases observed in the SAM solid sample analyses, including water, sulfur- and nitrogen-bearing compounds, and REMS observations of Relative Humidity and Temperature, and compare with knowledge of martian chlorine obtained from the SNC meteorites. Finally, we examine the implications of surface/atmosphere Cl interactions and isotopic ratios for the rise and decline of habitable surface environments on Mars. This research was supported by the National Aeronautics and Space Administration (NASA) Mars Science Laboratory mission.

  19. Influence of thiopurine methyltransferase gene polymorphism on ...

    Indian Academy of Sciences (India)

    Azza A. G. Tantawy

    2017-11-28

    Nov 28, 2017 ... Journal of Genetics, Vol. 96, No. ... aim of this study was to determine the influence of TPMT gene polymorphism in Egyptian children with acute lymphoblastic leukaemia (ALL). ... Keywords. thiopurine methyltransferase gene polymorphism; acute lymphoblastic leukaemia; Egyptian children; thiopurine.

  20. RESEARCH ARTICLE The influence of thiopurine methyltransferase ...

    Indian Academy of Sciences (India)

    Navya

    2017-02-06

    Feb 6, 2017 ... The influence of thiopurine methyltransferase gene polymorphism on Egyptian children with acute lymphoblastic leukaemia ... influence response and toxicity of therapy in children with acute lymphoblastic leukaemia. (ALL). [1]The ..... Egyptian Journal of Medical Human Genetics. 12, 183–186. 22. Hamdy ...

  1. Sam D. Hamilton Noxubee National Wildlife Refuge: Comprehensive Conservation Plan

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — This Comprehensive Conservation Plan (CCP) was written to guide management on Sam D. Hamilton Noxubee NWR for the next 15 years. This plan outlines the Refuge vision...

  2. Recent Updates to the System Advisor Model (SAM)

    Energy Technology Data Exchange (ETDEWEB)

    DiOrio, Nicholas A [National Renewable Energy Laboratory (NREL), Golden, CO (United States)

    2018-02-14

    The System Advisor Model (SAM) is a mature suite of techno-economic models for many renewable energy technologies that can be downloaded for free as a desktop application or software development kit. SAM is used for system-level modeling, including generating performance pro the release of the code as an open source project on GitHub. Other additions that will be covered include the ability to download data directly into SAM from the National Solar Radiation Database (NSRDB) and up- dates to a user-interface macro that assists with PV system sizing. A brief update on SAM's battery model and its integration with the detailed photovoltaic model will also be discussed. Finally, an outline of planned work for the next year will be presented, including the addition of a bifacial model, support for multiple MPPT inputs for detailed inverter modeling, and the addition of a model for inverter thermal behavior.

  3. General Safety and Waste Management Related to SAM

    Science.gov (United States)

    The General Safety and Waste Management page offers section-specific safety and waste management details for chemicals, radiochemicals, pathogens, and biotoxins included in EPA's Selected Analytical Methods for Environmental Remediation and Recovery (SAM).

  4. Safety and Waste Management for SAM Chemistry Methods

    Science.gov (United States)

    The General Safety and Waste Management page offers section-specific safety and waste management details for the chemical analytes included in EPA's Selected Analytical Methods for Environmental Remediation and Recovery (SAM).

  5. Safety and Waste Management for SAM Pathogen Methods

    Science.gov (United States)

    The General Safety and Waste Management page offers section-specific safety and waste management details for the pathogens included in EPA's Selected Analytical Methods for Environmental Remediation and Recovery (SAM).

  6. Safety and Waste Management for SAM Radiochemical Methods

    Science.gov (United States)

    The General Safety and Waste Management page offers section-specific safety and waste management details for the radiochemical analytes included in EPA's Selected Analytical Methods for Environmental Remediation and Recovery (SAM).

  7. Technical Manual for the SAM Physical Trough Model

    Energy Technology Data Exchange (ETDEWEB)

    Wagner, M. J.; Gilman, P.

    2011-06-01

    NREL, in conjunction with Sandia National Lab and the U.S Department of Energy, developed the System Advisor Model (SAM) analysis tool for renewable energy system performance and economic analysis. This paper documents the technical background and engineering formulation for one of SAM's two parabolic trough system models in SAM. The Physical Trough model calculates performance relationships based on physical first principles where possible, allowing the modeler to predict electricity production for a wider range of component geometries than is possible in the Empirical Trough model. This document describes the major parabolic trough plant subsystems in detail including the solar field, power block, thermal storage, piping, auxiliary heating, and control systems. This model makes use of both existing subsystem performance modeling approaches, and new approaches developed specifically for SAM.

  8. Kuula. Kuu artist Sam Sparro. Kuu plaat / Mart Juur

    Index Scriptorium Estoniae

    Juur, Mart, 1964-

    2008-01-01

    Artistist Sam Sparrost. Heliplaatidest: "Sex And The City" Original Motion Picture Soundtrack, "Maestro: Blue Note Trip", Melvin/Soo/Remmel/Julm "Geografix", Alanis Morrisette "Flavors Of Entanglement", Guillemots "Red"

  9. Sam Wagstaffi unustatud kired / Ahto Külvet

    Index Scriptorium Estoniae

    Külvet, Ahto

    2008-01-01

    Dokumentaalfilm "Black, White & Gray: Sam Wagstaff and Robert Mapplethorpe" : autor ja režissöör James Crump : Ameerika Ühendriigid 2007. Filmi näidati filminädala "Art in America" raames Tallinnas

  10. The role of the methyltransferase domain of bifunctional restriction enzyme RM.BpuSI in cleavage activity.

    Directory of Open Access Journals (Sweden)

    Arthur Sarrade-Loucheur

    Full Text Available Restriction enzyme (REase RM.BpuSI can be described as a Type IIS/C/G REase for its cleavage site outside of the recognition sequence (Type IIS, bifunctional polypeptide possessing both methyltransferase (MTase and endonuclease activities (Type IIC and endonuclease activity stimulated by S-adenosyl-L-methionine (SAM (Type IIG. The stimulatory effect of SAM on cleavage activity presents a major paradox: a co-factor of the MTase activity that renders the substrate unsusceptible to cleavage enhances the cleavage activity. Here we show that the RM.BpuSI MTase activity modifies both cleavage substrate and product only when they are unmethylated. The MTase activity is, however, much lower than that of M1.BpuSI and is thought not to be the major MTase for host DNA protection. SAM and sinefungin (SIN increase the Vmax of the RM.BpuSI cleavage activity with a proportional change in Km, suggesting the presence of an energetically more favorable pathway is taken. We further showed that RM.BpuSI undergoes substantial conformational changes in the presence of Ca(2+, SIN, cleavage substrate and/or product. Distinct conformers are inferred as the pre-cleavage/cleavage state (in the presence of Ca(2+, substrate or both and MTase state (in the presence of SIN and substrate, SIN and product or product alone. Interestingly, RM.BpuSI adopts a unique conformation when only SIN is present. This SIN-bound state is inferred as a branch point for cleavage and MTase activity and an intermediate to an energetically favorable pathway for cleavage, probably through increasing the binding affinity of the substrate to the enzyme under cleavage conditions. Mutation of a SAM-binding residue resulted in altered conformational changes in the presence of substrate or Ca(2+ and eliminated cleavage activity. The present study underscores the role of the MTase domain as facilitator of efficient cleavage activity for RM.BpuSI.

  11. DNA Methyltransferase Activity Assays: Advances and Challenges

    OpenAIRE

    Poh, Wan Jun; Wee, Cayden Pang Pee; Gao,Zhiqiang

    2016-01-01

    DNA methyltransferases (MTases), a family of enzymes that catalyse the methylation of DNA, have a profound effect on gene regulation. A large body of evidence has indicated that DNA MTase is potentially a predictive biomarker closely associated with genetic disorders and genetic diseases like cancer. Given the attention bestowed onto DNA MTases in molecular biology and medicine, highly sensitive detection of DNA MTase activity is essential in determining gene regulation, epigenetic modificati...

  12. Nicotinamide -Methyltransferase in Health and Cancer

    Directory of Open Access Journals (Sweden)

    David B Ramsden

    2017-06-01

    Full Text Available Over the past decade, the roles of nicotinamide N -methyltransferase and its product 1-methyl nicotinamide have emerged from playing merely minor roles in phase 2 xenobiotic metabolism as actors in some of the most important scenes of human life. In this review, the structures of the gene, messenger RNA, and protein are discussed, together with the role of the enzyme in many of the common cancers that afflict people today.

  13. Ensuring GRID resource availability with the SAM framework in LHCb

    Science.gov (United States)

    Closier, J.; Paterson, S.; Santinelli, R.

    2008-07-01

    The LHCb experiment has chosen to use the SAM framework (Service Availability Monitoring Environment from EGEE-II) [1] make extensive tests of the LHCb environment at all the accessible grid resources. The availability and the proper definition of the local Computing and Storage Elements, user interfaces as well as the WLCG software environment are checked. The SAM framework is also used to pre-install the LHCb applications in the shared software area provided by each site. The deployment of the LHCb applications is based on a python tool developed inside the experiment. It is used for software management including incremental installation of interdependent packages and clean package removal. After the application software is installed a validation test of the whole MC chain is run. According to the results of the experiment specific SAM tests, the sites are (re)integrated into the LHCb production system managed by DIRAC [2]. The possibility of automated dynamic site certification using the SAM test suite is explored. This paper will describe the various ways of the LHCb use of the SAM framework. Practical experience in the recent production runs, current limitations and future developments will be presented.

  14. Flaviviral methyltransferase/RNA interaction: structural basis for enzyme inhibition.

    Science.gov (United States)

    Milani, Mario; Mastrangelo, Eloise; Bollati, Michela; Selisko, Barbara; Decroly, Etienne; Bouvet, Mickaël; Canard, Bruno; Bolognesi, Martino

    2009-07-01

    Flaviviruses are the causative agents of severe diseases such as Dengue or Yellow fever. The replicative machinery used by the virus is based on few enzymes including a methyltransferase, located in the N-terminal domain of the NS5 protein. Flaviviral methyltransferases are involved in the last two steps of the mRNA capping process, transferring a methyl group from S-adenosyl-L-methionine onto the N7 position of the cap guanine (guanine-N7 methyltransferase) and the ribose 2'O position of the first nucleotide following the cap guanine (nucleoside-2'O methyltransferase). The RNA capping process is crucial for mRNA stability, protein synthesis and virus replication. Such an essential function makes methyltransferases attractive targets for the design of antiviral drugs. In this context, starting from the crystal structure of Wesselsbron flavivirus methyltransferase, we elaborated a mechanistic model describing protein/RNA interaction during N7 methyl transfer. Next we used an in silico docking procedure to identify commercially available compounds that would display high affinity for the methyltransferase active site. The best candidates selected were tested in vitro to assay their effective inhibition on 2'O and N7 methyltransferase activities on Wesselsbron and Dengue virus (Dv) methyltransferases. The results of such combined computational and experimental screening approach led to the identification of a high-potency inhibitor.

  15. Native interface of the SAM domain polymer of TEL

    Directory of Open Access Journals (Sweden)

    Bowie James U

    2002-08-01

    Full Text Available Abstract Background TEL is a transcriptional repressor containing a SAM domain that forms a helical polymer. In a number of hematologic malignancies, chromosomal translocations lead to aberrant fusions of TEL-SAM to a variety of other proteins, including many tyrosine kinases. TEL-SAM polymerization results in constitutive activation of the tyrosine kinase domains to which it becomes fused, leading to cell transformation. Thus, inhibitors of TEL-SAM self-association could abrogate transformation in these cells. In previous work, we determined the structure of a mutant TEL-SAM polymer bearing a Val to Glu substitution in center of the subunit interface. It remained unclear how much the mutation affected the architecture of the polymer, however. Results Here we determine the structure of the native polymer interface. To accomplish this goal, we introduced mutations that block polymer extension, producing a heterodimer with a wild-type interface. We find that the structure of the wild-type polymer interface is quite similar to the mutant structure determined previously. With the structure of the native interface, it is possible to evaluate the potential for developing therapeutic inhibitors of the interaction. We find that the interacting surfaces of the protein are relatively flat, containing no obvious pockets for the design of small molecule inhibitors. Conclusion Our results confirm the architecture of the TEL-SAM polymer proposed previously based on a mutant structure. The fact that the interface contains no obvious potential binding pockets suggests that it may be difficult to find small molecule inhibitors to treat malignancies in this way.

  16. Technoeconomic Modeling of Battery Energy Storage in SAM

    Energy Technology Data Exchange (ETDEWEB)

    DiOrio, Nicholas [National Renewable Energy Lab. (NREL), Golden, CO (United States); Dobos, Aron [National Renewable Energy Lab. (NREL), Golden, CO (United States); Janzou, Steven [National Renewable Energy Lab. (NREL), Golden, CO (United States); Nelson, Austin [National Renewable Energy Lab. (NREL), Golden, CO (United States); Lundstrom, Blake [National Renewable Energy Lab. (NREL), Golden, CO (United States)

    2015-09-01

    Detailed comprehensive lead-acid and lithium-ion battery models have been integrated with photovoltaic models in an effort to allow System Advisor Model (SAM) to offer the ability to predict the performance and economic benefit of behind the meter storage. In a system with storage, excess PV energy can be saved until later in the day when PV production has fallen, or until times of peak demand when it is more valuable. Complex dispatch strategies can be developed to leverage storage to reduce energy consumption or power demand based on the utility rate structure. This document describes the details of the battery performance and economic models in SAM.

  17. Obituary: Sam Roweis (1972-2010)

    Science.gov (United States)

    Hogg, David

    2011-12-01

    Computer scientist and statistical astronomer Sam Roweis took his own life in New York City on 2010 January 12. He was a brilliant and accomplished researcher in the field of machine learning, and a strong advocate for the use of computational statistics for automating discovery and scientific data analysis. He made several important contributions to astronomy and was working on adaptive astronomical data analysis at the time of his death. Roweis obtained his PhD in 1999 from the California Institute of Technology, where he worked on a remarkable range of subjects, including DNA computing, modeling of dynamical systems, signal processing, and speech recognition. During this time he unified and clarified some of the most important data analysis techniques, including Principal Component Analysis, Hidden Markov Models, and Expectation Maximization. His work was aimed at making data analysis and modeling faster, but also better justified scientifically. The last years of his PhD were spent in Princeton NJ, where he came in contact with a young generation of cosmologists thinking about microwave background and large-scale structure data. In a postdoc at University College London, Roweis co-created the Locally Linear Embedding (LLE) algorithm; a simple but flexible technique for mapping a large data set onto a low-dimensional manifold. The LLE paper obtained more than 2700 citations in 9 years, launched a new sub-field of machine learning known as "manifold learning," and inspired work in data visualization, search, and applied mathematics. In 2001, Roweis took a faculty job at the University of Toronto Computer Science Department. He continued working on data analysis methods that have probabilistic interpretation and therefore scientific applicability, but at the same time have good performance on large data sets. He was awarded a Sloan Fellowship, a Canada Research Chair, and a fellowship of the Canadian Institute for Advanced Research, among other honors and awards

  18. A Mononuclear Iron-Dependent Methyltransferase Catalyzes Initial Steps in Assembly of the Apratoxin A Polyketide Starter Unit.

    Science.gov (United States)

    Skiba, Meredith A; Sikkema, Andrew P; Moss, Nathan A; Tran, Collin L; Sturgis, Rebecca M; Gerwick, Lena; Gerwick, William H; Sherman, David H; Smith, Janet L

    2017-11-02

    Natural product biosynthetic pathways contain a plethora of enzymatic tools to carry out difficult biosynthetic transformations. Here we discover an unusual mononuclear iron-dependent methyltransferase that acts in the initiation steps of apratoxin A biosynthesis (AprA MT1). Iron-replete AprA MT1 catalyzes one or two methyl transfer reactions on the substrate malonyl-ACP (acyl carrier protein), whereas Co, Mn and Ni support only a single methyl transfer. MT1 homologs exist within the "GNAT" (GCN5-related N-acetyltransferase) loading modules of several modular biosynthetic pathways with propionyl, isobutyryl or pivaloyl starter units. GNAT domains are thought to catalyze decarboxylation of malonyl-CoA and acetyl transfer to a carrier protein. In AprA the GNAT domain lacks both decarboxylation and acyl transfer activity, and the MT1 acts on the malonyl α-carbon, which is more reactive than the acetyl methyl. A crystal structure of the AprA MT1-GNAT didomain with bound Mn2+, malonate and the methyl donor S-adenosylmethionine (SAM) reveals that the malonyl substrate is a bidentate metal ligand, indicating that the metal acts as a Lewis acid to promote methylation of the malonyl α-carbon. The GNAT domain is truncated relative to functional homologs. These results afford an expanded understanding of MT1-GNAT structure and activity, and permit the functional annotation of homologous GNAT loading modules both with and without methyltransferases, additionally revealing their rapid evolutionary adaptation in different biosynthetic contexts.

  19. Cross-Reactive Sensor Array for Metal Ion Sensing Based on Fluorescent SAMs

    NARCIS (Netherlands)

    Basabe Desmonts, M.L.; van der Baan, Frederieke; Zimmerman, R.S.; Reinhoudt, David; Crego Calama, Mercedes

    2007-01-01

    Fluorescent self assembled monolayers (SAMs) on glass were previouslydeveloped in our group as new sensing materials for metal ions. These fluorescent SAMs arecomprised by fluorophores and small molecules sequentially deposited on a monolayer onglass. The preorganization provided by the surface

  20. ISO19770-1:2012 SAM process guidance

    CERN Document Server

    Canavan, Rory

    2012-01-01

    The pocket guide offers a concise summary of the principles of software asset management as conveyed by ISO 19770-1: 2012, and provides advice and guidance on how to kick-start your own SAM programme - something the Standard alone doesn't offer.

  1. Sam Karlin and multi-locus population genetics.

    Science.gov (United States)

    Feldman, Marcus W

    2009-06-01

    Between 1967 and 1982, Sam Karlin made fundamental contributions to many areas of deterministic population genetic theory. This remembrance focuses on his work in multi-locus population genetics, primarily on the interaction between genotypic selection and the rate of recombination.

  2. Social Activity Method (SAM): A Fractal Language for Mathematics

    Science.gov (United States)

    Dowling, Paul

    2013-01-01

    In this paper I shall present and develop my organisational language, "social activity method" (SAM), and illustrate some of its applications. I shall introduce a new scheme for "modes of recontextualisation" that enables the analysis of the ways in which one activity--which might be school mathematics or social research or any…

  3. 77 FR 50493 - Sam Rayburn Dam Project Power Rate

    Science.gov (United States)

    2012-08-21

    .... Procedures for Public Participation in Power and Transmission Rate Adjustments and Extensions of the Power... transmission facilities, which consist of 1,380 miles of high-voltage transmission lines, 25 substations, and 46 microwave and VHF radio sites. Costs associated ] with the Sam Rayburn and Robert D. Willis Dams...

  4. Binding of the Methyl DonorS-Adenosyl-l-Methionine to Middle East Respiratory Syndrome Coronavirus 2'-O-Methyltransferase nsp16 Promotes Recruitment of the Allosteric Activator nsp10.

    Science.gov (United States)

    Aouadi, Wahiba; Blanjoie, Alexandre; Vasseur, Jean-Jacques; Debart, Françoise; Canard, Bruno; Decroly, Etienne

    2017-03-01

    The Middle East respiratory syndrome coronavirus (MERS-CoV) nonstructural protein 16 (nsp16) is an S -adenosyl-l-methionine (SAM)-dependent 2'- O -methyltransferase (2'-O-MTase) that is thought to methylate the ribose 2'-OH of the first transcribed nucleotide (N 1 ) of viral RNA cap structures. This 2'-O-MTase activity is regulated by nsp10. The 2'- O methylation prevents virus detection by cell innate immunity mechanisms and viral translation inhibition by the interferon-stimulated IFIT-1 protein. To unravel the regulation of nsp10/nsp16 2'-O-MTase activity, we used purified MERS-CoV nsp16 and nsp10. First, we showed that nsp16 recruited N7-methylated capped RNA and SAM. The SAM binding promotes the assembly of the enzymatically active nsp10/nsp16 complex that converted 7m GpppG (cap-0) into 7m GpppG 2'Om (cap-1) RNA by 2'-OH methylation of N 1 in a SAM-dependent manner. The subsequent release of SAH speeds up nsp10/nsp16 dissociation that stimulates the reaction turnover. Alanine mutagenesis and RNA binding assays allowed the identification of the nsp16 residues involved in RNA recognition forming the RNA binding groove (K46, K170, E203, D133, R38, Y47, and Y181) and the cap-0 binding site (Y30, Y132, and H174). Finally, we found that nsp10/nsp16 2'-O-MTase activity is sensitive to known MTase inhibitors, such as sinefungin and cap analogues. This characterization of the MERS-CoV 2'-O-MTase is a preliminary step toward the development of molecules to inhibit cap 2'-O methylation and to restore the host antiviral response. IMPORTANCE MERS-CoV codes for a cap 2'- O -methyltransferase that converts cap-0 into cap-1 structure in order to prevent virus detection by cell innate immunity mechanisms. We report the biochemical properties of MERS-CoV 2'O-methyltransferase, which is stimulated by nsp10 acting as an allosteric activator of the nsp16 2'- O -methyltransferase possibly through enhanced RNA binding affinity. In addition, we show that SAM promotes the formation

  5. Social accounting matrices. The development and application of SAMs at the local level

    OpenAIRE

    van Leeuwen, E.S.; Nijkamp, P.

    2009-01-01

    This contribution aims to highlight the importance of Social Accounting Matrices (SAMs) for the study of regional-economic interactions. After a conceptual review of SAMs, the attention is focused on the empirical meaning of SAMs for economic impact assessment. The potential of SAMs is illustrated by an extensive pedagogical treatment of this tool on the basis of several town-hinterland interactions in 5 different European countries.

  6. A cfr-Like Gene from Clostridium difficile Confers Multiple Antibiotic Resistance by the Same Mechanism as the cfr Gene

    DEFF Research Database (Denmark)

    Hansen, Lykke H; Vester, Birte

    2015-01-01

    The Cfr RNA methyltransferase causes multiple resistances to peptidyl transferase inhibitors by methylation of A2503 23S rRNA. Many cfr-like gene sequences in the databases code for unknown functions. This study confirms that a Cfr-like protein from a Peptoclostridium difficile (formerly Clostrid...... Clostridium difficile) strain does function as a Cfr protein. The enzyme is expressed in Escherichia coli and shows elevated MICs for five classes of antibiotics. A primer extension stop indicates a modification at A2503 in 23S rRNA.......The Cfr RNA methyltransferase causes multiple resistances to peptidyl transferase inhibitors by methylation of A2503 23S rRNA. Many cfr-like gene sequences in the databases code for unknown functions. This study confirms that a Cfr-like protein from a Peptoclostridium difficile (formerly...

  7. A cfr-Like Gene from Clostridium difficile Confers Multiple Antibiotic Resistance by the Same Mechanism as the cfr Gene

    DEFF Research Database (Denmark)

    Hansen, Lykke H; Vester, Birte

    2015-01-01

    The Cfr RNA methyltransferase causes multiple resistances to peptidyl transferase inhibitors by methylation of A2503 23S rRNA. Many cfr-like gene sequences in the databases code for unknown functions. This study confirms that a Cfr-like protein from a Peptoclostridium difficile (formerly Clostrid......The Cfr RNA methyltransferase causes multiple resistances to peptidyl transferase inhibitors by methylation of A2503 23S rRNA. Many cfr-like gene sequences in the databases code for unknown functions. This study confirms that a Cfr-like protein from a Peptoclostridium difficile (formerly...... Clostridium difficile) strain does function as a Cfr protein. The enzyme is expressed in Escherichia coli and shows elevated MICs for five classes of antibiotics. A primer extension stop indicates a modification at A2503 in 23S rRNA....

  8. Adsorption kinetic investigations of phthalocyanine derivatives self assembled monolayers (SAMs) on gold: Temperature influence on the SAM formation process and quality

    OpenAIRE

    Liličenko, Nadežda

    2015-01-01

    The ordered nano-structured surfaces, like self-assembled monolayers (SAMs) are of a great scientific interest, due to the low cost, simplicity, and versatility of this method. SAMs found numerous of applications in molecular electronics, biochemistry and optical devices. Phthalocyanine (Pc) complexes are of particular interest for the SAM preparation. These molecules exhibit fascinating physical properties and are chemically and thermally stable. Moreover their complex stru...

  9. Recurrent horizontal transfer of arsenite methyltransferase genes facilitated adaptation of life to arsenic.

    Science.gov (United States)

    Chen, Song-Can; Sun, Guo-Xin; Rosen, Barry P; Zhang, Si-Yu; Deng, Ye; Zhu, Bo-Kai; Rensing, Christopher; Zhu, Yong-Guan

    2017-08-10

    The toxic metalloid arsenic has been environmentally ubiquitous since life first arose nearly four billion years ago and presents a challenge for the survival of all living organisms. Its bioavailability has varied dramatically over the history of life on Earth. As life spread, biogeochemical and climate changes cyclically increased and decreased bioavailable arsenic. To elucidate the history of arsenic adaptation across the tree of life, we reconstructed the phylogeny of the arsM gene that encodes the As(III) S-adenosylmethionine (SAM) methyltransferase. Our results suggest that life successfully moved into arsenic-rich environments in the late Archean Eon and Proterozoic Eon, respectively, by the spread of arsM genes. The arsM genes of bacterial origin have been transferred to other kingdoms of life on at least six occasions, and the resulting domesticated arsM genes promoted adaptation to environmental arsenic. These results allow us to peer into the history of arsenic adaptation of life on our planet and imply that dissemination of genes encoding diverse adaptive functions to toxic chemicals permit adaptation to changes in concentrations of environmental toxins over evolutionary history.

  10. Isolation of promoter for N-methyltransferase gene associated with caffeine biosynthesis in Coffea canephora.

    Science.gov (United States)

    Satyanarayana, K V; Kumar, Vinod; Chandrashekar, A; Ravishankar, G A

    2005-09-22

    N-Methyltransferases (NMTs) catalyze the three SAM dependent sequential methylation of xanthosine, producing caffeine in Coffea species. In the present work, a PCR based genome walking method was adopted to isolate and clone the promoter for the NMT gene. Inspection of the promoter sequence revealed the presence of several motifs important for the regulation of the gene expression. The whole fragment was fused to the beta-glucuronidase (gus) reporter gene and used in Agrobacterium tumefaciens mediated transformation of Nicotiana tabacum. GUS assays proved that the isolated promoter was able to direct the expression of the reporter gene in transgenic tobacco. Based on the promoter sequence, primer was designed and the genomic fragment comprising the promoter and its corresponding gene was amplified and cloned. Sequencing of one of the genomic clones revealed the presence of four exons and three introns in NMT gene. The differences in the restriction pattern among the genomic clones were studied using PCR-RFLP. This is the first report of cloning of the promoter for a gene involved in caffeine biosynthetic pathway and it opens up the possibility of studying the molecular mechanisms that regulate the production of caffeine.

  11. Automated One-Loop Calculations with GoSam

    CERN Document Server

    Cullen, Gavin; Heinrich, Gudrun; Luisoni, Gionata; Mastrolia, Pierpaolo; Ossola, Giovanni; Reiter, Thomas; Tramontano, Francesco

    2012-01-01

    We present the program package GoSam which is designed for the automated calculation of one-loop amplitudes for multi-particle processes in renormalisable quantum field theories. The amplitudes, which are generated in terms of Feynman diagrams, can be reduced using either D-dimensional integrand-level decomposition or tensor reduction. GoSam can be used to calculate one-loop QCD and/or electroweak corrections to Standard Model processes and offers the flexibility to link model files for theories Beyond the Standard Model. A standard interface to programs calculating real radiation is also implemented. We demonstrate the flexibility of the program by presenting examples of processes with up to six external legs attached to the loop.

  12. Technical Manual for the SAM Biomass Power Generation Model

    Energy Technology Data Exchange (ETDEWEB)

    Jorgenson, J.; Gilman, P.; Dobos, A.

    2011-09-01

    This technical manual provides context for the implementation of the biomass electric power generation performance model in the National Renewable Energy Laboratory's (NREL's) System Advisor Model (SAM). Additionally, the report details the engineering and scientific principles behind the underlying calculations in the model. The framework established in this manual is designed to give users a complete understanding of behind-the-scenes calculations and the results generated.

  13. Genome sequencing and annotation of Stenotrophomonas sp. SAM8.

    Science.gov (United States)

    Selim, Samy; Hassan, Sherif; Hagagy, Nashwa

    2015-12-01

    We report draft genome sequence of Stenotrophomonas sp. strain SAM8, isolated from environmental water. The draft genome size is 3,665,538 bp with a G + C content of 67.2% and contains 6 rRNA sequence (single copies of 5S, 16S & 23S rRNA). The genome sequence can be accessed at DDBJ/EMBL/GenBank under the accession no. LDAV00000000.

  14. Simulated SAM A-scans on multilayer MEMS components

    DEFF Research Database (Denmark)

    Janting, Jakob; Petersen, Dirch Hjorth; Greisen, Christoffer

    2002-01-01

    A spreadsheet program for simulation of Scanning Acoustic Microscopy (SAM) A-scans on multilayer structures has been developed. Using this program, structure variations in samples can be analysed better. Further samples can be prepared to get optimal signal for enhanced failure and materials anal....... The calculation of N, the program interface, and simulated A-scans on MEMS test structures for a pressure sensor are presented. (C) 2002 Elsevier Science Ltd. All rights reserved....

  15. Genome sequencing and annotation of Stenotrophomonas sp. SAM8

    Directory of Open Access Journals (Sweden)

    Samy Selim

    2015-12-01

    Full Text Available We report draft genome sequence of Stenotrophomonas sp. strain SAM8, isolated from environmental water. The draft genome size is 3,665,538 bp with a G + C content of 67.2% and contains 6 rRNA sequence (single copies of 5S, 16S & 23S rRNA. The genome sequence can be accessed at DDBJ/EMBL/GenBank under the accession no. LDAV00000000.

  16. CE-SAM: a conversational interface for ISR mission support

    Science.gov (United States)

    Pizzocaro, Diego; Parizas, Christos; Preece, Alun; Braines, Dave; Mott, David; Bakdash, Jonathan Z.

    2013-05-01

    There is considerable interest in natural language conversational interfaces. These allow for complex user interactions with systems, such as fulfilling information requirements in dynamic environments, without requiring extensive training or a technical background (e.g. in formal query languages or schemas). To leverage the advantages of conversational interactions we propose CE-SAM (Controlled English Sensor Assignment to Missions), a system that guides users through refining and satisfying their information needs in the context of Intelligence, Surveillance, and Reconnaissance (ISR) operations. The rapidly-increasing availability of sensing assets and other information sources poses substantial challenges to effective ISR resource management. In a coalition context, the problem is even more complex, because assets may be "owned" by different partners. We show how CE-SAM allows a user to refine and relate their ISR information needs to pre-existing concepts in an ISR knowledge base, via conversational interaction implemented on a tablet device. The knowledge base is represented using Controlled English (CE) - a form of controlled natural language that is both human-readable and machine processable (i.e. can be used to implement automated reasoning). Users interact with the CE-SAM conversational interface using natural language, which the system converts to CE for feeding-back to the user for confirmation (e.g. to reduce misunderstanding). We show that this process not only allows users to access the assets that can support their mission needs, but also assists them in extending the CE knowledge base with new concepts.

  17. Nicotinamide N-Methyltransferase Suppression Participates in Nickel-Induced Histone H3 Lysine9 Dimethylation in BEAS-2B Cells

    Directory of Open Access Journals (Sweden)

    Qian Li

    2017-04-01

    Full Text Available Background: Nickel compounds are well-established human carcinogens with weak mutagenic activity. Histone methylation has been proposed to play an important role in nickel-induced carcinogenesis. Nicotinamide N-methyltransferase (NNMT decreases histone methylation in several cancer cells by altering the cellular ratio of S-adenosylmethionine (SAM to S-adenosylhomocysteine (SAH. However, the role of NNMT in nickel-induced histone methylation remains unclear. Methods: BEAS-2B cells were exposed to different concentrations of nickel chloride (NiCl2 for 72 h or 200 μM NiCl2 for different time periods. Histone H3 on lysine 9 (H3K9 mono-, di-, and trimethylation and NNMT protein levels were measured by western blot analysis. Expressions of NNMT mRNA and the H3k9me2-associated genes, mitogen-activated protein kinase 3 (MAP2K3 and dickkopf1 (DKK1, were determined by qPCR analysis. The cellular ratio of nicotinamide adenine dinucleotide (NAD+ to reduced NAD (NADH and SAM/SAH ratio were determined. Results: Exposure of BEAS-2B cells to nickel increased H3K9 dimethylation (H3K9me2, suppressed the expressions of H3K9me2-associated genes (MAP2K3 and DKK1, and induced NNMT repression at both the protein and mRNA levels. Furthermore, over-expression of NNMT inhibited nickel-induced H3K9me2 and altered the cellular SAM/SAH ratio. Additionally, the NADH oxidant phenazine methosulfate (PMS not only reversed the nickel-induced reduction in NAD+/NADH but also inhibited the increase in H3K9me2. Conclusions: These findings indicate that the repression of NNMT may underlie nickel-induced H3K9 dimethylation by altering the cellular SAM/SAH ratio.

  18. Nicotinamide N-Methyltransferase Suppression Participates in Nickel-Induced Histone H3 Lysine9 Dimethylation in BEAS-2B Cells.

    Science.gov (United States)

    Li, Qian; He, Min-Di; Mao, Lin; Wang, Xue; Jiang, Yu-Lin; Li, Min; Lu, Yong-Hui; Yu, Zheng-Ping; Zhou, Zhou

    2017-01-01

    Nickel compounds are well-established human carcinogens with weak mutagenic activity. Histone methylation has been proposed to play an important role in nickel-induced carcinogenesis. Nicotinamide N-methyltransferase (NNMT) decreases histone methylation in several cancer cells by altering the cellular ratio of S-adenosylmethionine (SAM) to S-adenosylhomocysteine (SAH). However, the role of NNMT in nickel-induced histone methylation remains unclear. BEAS-2B cells were exposed to different concentrations of nickel chloride (NiCl2) for 72 h or 200 μM NiCl2 for different time periods. Histone H3 on lysine 9 (H3K9) mono-, di-, and trimethylation and NNMT protein levels were measured by western blot analysis. Expressions of NNMT mRNA and the H3k9me2-associated genes, mitogen-activated protein kinase 3 (MAP2K3) and dickkopf1 (DKK1), were determined by qPCR analysis. The cellular ratio of nicotinamide adenine dinucleotide (NAD+) to reduced NAD (NADH) and SAM/SAH ratio were determined. Exposure of BEAS-2B cells to nickel increased H3K9 dimethylation (H3K9me2), suppressed the expressions of H3K9me2-associated genes (MAP2K3 and DKK1), and induced NNMT repression at both the protein and mRNA levels. Furthermore, over-expression of NNMT inhibited nickel-induced H3K9me2 and altered the cellular SAM/SAH ratio. Additionally, the NADH oxidant phenazine methosulfate (PMS) not only reversed the nickel-induced reduction in NAD+/NADH but also inhibited the increase in H3K9me2. These findings indicate that the repression of NNMT may underlie nickel-induced H3K9 dimethylation by altering the cellular SAM/SAH ratio. © 2017 The Author(s)Published by S. Karger AG, Basel.

  19. Monolignol 4-O-methyltransferases and uses thereof

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Chang-Jun; Bhuiya, Mohammad-Wadud; Zhang, Kewei

    2014-11-18

    Modified (iso)eugenol 4-O-methyltransferase enzymes having novel capacity for methylation of monolignols and reduction of lignin polymerization in plant cell wall are disclosed. Sequences encoding the modified enzymes are disclosed.

  20. The SAMS: Smartphone Addiction Management System and verification.

    Science.gov (United States)

    Lee, Heyoung; Ahn, Heejune; Choi, Samwook; Choi, Wanbok

    2014-01-01

    While the popularity of smartphones has given enormous convenience to our lives, their pathological use has created a new mental health concern among the community. Hence, intensive research is being conducted on the etiology and treatment of the condition. However, the traditional clinical approach based surveys and interviews has serious limitations: health professionals cannot perform continual assessment and intervention for the affected group and the subjectivity of assessment is questionable. To cope with these limitations, a comprehensive ICT (Information and Communications Technology) system called SAMS (Smartphone Addiction Management System) is developed for objective assessment and intervention. The SAMS system consists of an Android smartphone application and a web application server. The SAMS client monitors the user's application usage together with GPS location and Internet access location, and transmits the data to the SAMS server. The SAMS server stores the usage data and performs key statistical data analysis and usage intervention according to the clinicians' decision. To verify the reliability and efficacy of the developed system, a comparison study with survey-based screening with the K-SAS (Korean Smartphone Addiction Scale) as well as self-field trials is performed. The comparison study is done using usage data from 14 users who are 19 to 50 year old adults that left at least 1 week usage logs and completed the survey questionnaires. The field trial fully verified the accuracy of the time, location, and Internet access information in the usage measurement and the reliability of the system operation over more than 2 weeks. The comparison study showed that daily use count has a strong correlation with K-SAS scores, whereas daily use times do not strongly correlate for potentially addicted users. The correlation coefficients of count and times with total K-SAS score are CC = 0.62 and CC =0.07, respectively, and the t-test analysis for the

  1. Ablation of the Sam68 RNA binding protein protects mice from age-related bone loss.

    Directory of Open Access Journals (Sweden)

    Stéphane Richard

    2005-12-01

    Full Text Available The Src substrate associated in mitosis of 68 kDa (Sam68 is a KH-type RNA binding protein that has been shown to regulate several aspects of RNA metabolism; however, its physiologic role has remained elusive. Herein we report the generation of Sam68-null mice by homologous recombination. Aged Sam68-/- mice preserved their bone mass, in sharp contrast with 12-month-old wild-type littermates in which bone mass was decreased up to approximately 75%. In fact, the bone volume of the 12-month-old Sam68-/- mice was virtually indistinguishable from that of 4-month-old wild-type or Sam68-/- mice. Sam68-/- bone marrow stromal cells had a differentiation advantage for the osteogenic pathway. Moreover, the knockdown of Sam68 using short hairpin RNA in the embryonic mesenchymal multipotential progenitor C3H10T1/2 cells resulted in more pronounced expression of the mature osteoblast marker osteocalcin when differentiation was induced with bone morphogenetic protein-2. Cultures of mouse embryo fibroblasts generated from Sam68+/+ and Sam68-/- littermates were induced to differentiate into adipocytes with culture medium containing pioglitazone and the Sam68-/- mouse embryo fibroblasts shown to have impaired adipocyte differentiation. Furthermore, in vivo it was shown that sections of bone from 12-month-old Sam68-/- mice had few marrow adipocytes compared with their age-matched wild-type littermate controls, which exhibited fatty bone marrow. Our findings identify endogenous Sam68 as a positive regulator of adipocyte differentiation and a negative regulator of osteoblast differentiation, which is consistent with Sam68 being a modulator of bone marrow mesenchymal cell differentiation, and hence bone metabolism, in aged mice.

  2. Congressmember Sam Farr: Five Decades of Public Service

    OpenAIRE

    Reti, Irene H.; Farr, Sam

    2017-01-01

    Congressmember Sam Farr (born July 4, 1941) represented California’s Central Coast in the United States House of Representatives for twenty-three years until his retirement from office in 2016.  Farr also served six years as a member of the Monterey County Board of Supervisors and twelve years in the California State Assembly. This oral history, a transcript of twenty-five hours of interviews conducted by Irene Reti, director of the UCSC Library’s Regional History Project, during the period i...

  3. DNA Methyltransferase Activity Assays: Advances and Challenges.

    Science.gov (United States)

    Poh, Wan Jun; Wee, Cayden Pang Pee; Gao, Zhiqiang

    2016-01-01

    DNA methyltransferases (MTases), a family of enzymes that catalyse the methylation of DNA, have a profound effect on gene regulation. A large body of evidence has indicated that DNA MTase is potentially a predictive biomarker closely associated with genetic disorders and genetic diseases like cancer. Given the attention bestowed onto DNA MTases in molecular biology and medicine, highly sensitive detection of DNA MTase activity is essential in determining gene regulation, epigenetic modification, clinical diagnosis and therapeutics. Conventional techniques such as isotope labelling are effective, but they often require laborious sample preparation, isotope labelling, sophisticated equipment and large amounts of DNA, rendering them unsuitable for uses at point-of-care. Simple, portable, highly sensitive and low-cost assays are urgently needed for DNA MTase activity screening. In most recent technological advances, many alternative DNA MTase activity assays such as fluorescent, electrochemical, colorimetric and chemiluminescent assays have been proposed. In addition, many of them are coupled with nanomaterials and/or enzymes to significantly enhance their sensitivity. Herein we review the progress in the development of DNA MTase activity assays with an emphasis on assay mechanism and performance with some discussion on challenges and perspectives. It is hoped that this article will provide a broad coverage of DNA MTase activity assays and their latest developments and open new perspectives toward the development of DNA MTase activity assays with much improved performance for uses in molecular biology and clinical practice.

  4. Biochemical Characterization and Structural Basis of Reactivity and Regioselectivity Differences between Burkholderia thailandensis and Burkholderia glumae 1,6-Didesmethyltoxoflavin N-Methyltransferase.

    Science.gov (United States)

    Fenwick, Michael K; Almabruk, Khaled H; Ealick, Steven E; Begley, Tadhg P; Philmus, Benjamin

    2017-08-01

    Burkholderia glumae converts the guanine base of guanosine triphosphate into an azapteridine and methylates both the pyrimidine and triazine rings to make toxoflavin. Strains of Burkholderia thailandensis and Burkholderia pseudomallei have a gene cluster encoding seven putative biosynthetic enzymes that resembles the toxoflavin gene cluster. Four of the enzymes are similar in sequence to BgToxBCDE, which have been proposed to make 1,6-didesmethyltoxoflavin (1,6-DDMT). One of the remaining enzymes, BthII1283 in B. thailandensis E264, is a predicted S-adenosylmethionine (SAM)-dependent N-methyltransferase that shows a low level of sequence identity to BgToxA, which sequentially methylates N6 and N1 of 1,6-DDMT to form toxoflavin. Here we show that, unlike BgToxA, BthII1283 catalyzes a single methyl transfer to N1 of 1,6-DDMT in vitro. In addition, we investigated the differences in reactivity and regioselectivity by determining crystal structures of BthII1283 with bound S-adenosylhomocysteine (SAH) or 1,6-DDMT and SAH. BthII1283 contains a class I methyltransferase fold and three unique extensions used for 1,6-DDMT recognition. The active site structure suggests that 1,6-DDMT is bound in a reduced form. The plane of the azapteridine ring system is orthogonal to its orientation in BgToxA. In BthII1283, the modeled SAM methyl group is directed toward the p orbital of N1, whereas in BgToxA, it is first directed toward an sp(2) orbital of N6 and then toward an sp(2) orbital of N1 after planar rotation of the azapteridine ring system. Furthermore, in BthII1283, N1 is hydrogen bonded to a histidine residue whereas BgToxA does not supply an obvious basic residue for either N6 or N1 methylation.

  5. A novel tumor suppressor function of glycine N-methyltransferase is independent of its catalytic activity but requires nuclear localization.

    Directory of Open Access Journals (Sweden)

    Suchandra DebRoy

    Full Text Available Glycine N-methyltransferase (GNMT, an abundant cytosolic enzyme, catalyzes the transfer of a methyl group from S-adenosylmethionine (SAM to glycine generating S-adenosylhomocysteine and sarcosine (N-methylglycine. This reaction is regulated by 5-methyltetrahydrofolate, which inhibits the enzyme catalysis. In the present study, we observed that GNMT is strongly down regulated in human cancers and is undetectable in cancer cell lines while the transient expression of the protein in cancer cells induces apoptosis and results in the activation of ERK1/2 as an early pro-survival response. The antiproliferative effect of GNMT can be partially reversed by treatment with the pan-caspase inhibitor zVAD-fmk but not by supplementation with high folate or SAM. GNMT exerts the suppressor effect primarily in cells originated from malignant tumors: transformed cell line of non-cancer origin, HEK293, was insensitive to GNMT. Of note, high levels of GNMT, detected in regenerating liver and in NIH3T3 mouse fibroblasts, do not produce cytotoxic effects. Importantly, GNMT, a predominantly cytoplasmic protein, was translocated into nuclei upon transfection of cancer cells. The presence of GNMT in the nuclei was also observed in normal human tissues by immunohistochemical staining. We further demonstrated that the induction of apoptosis is associated with the GNMT nuclear localization but is independent of its catalytic activity or folate binding. GNMT targeted to nuclei, through the fusion with nuclear localization signal, still exerts strong antiproliferative effects while its restriction to cytoplasm, through the fusion with nuclear export signal, prevents these effects (in each case the protein was excluded from cytosol or nuclei, respectively. Overall, our study indicates that GNMT has a secondary function, as a regulator of cellular proliferation, which is independent of its catalytic role.

  6. Molecular cloning, characterization and expression analysis of the protein arginine N-methyltransferase 1 gene (As-PRMT1) from Artemia sinica.

    Science.gov (United States)

    Jiang, Xue; Yao, Feng; Li, Xuejie; Jia, Baolin; Zhong, Guangying; Zhang, Jianfeng; Zou, Xiangyang; Hou, Lin

    2015-07-01

    Protein arginine N-methyltransferase 1 (PRMT1) is an important epigenetic regulation factor in eukaryotic genomes. PRMT1 is involved in histone arginine loci methylation modification, changes in eukaryotic genomes' chromatin structure, and gene expression regulation. In the present paper, the full-length 1201-bp cDNA sequence of the PRMT1 homolog of Artemia sinica (As-PRMT1) was cloned for the first time. The putative As-PRMT1 protein comprises 346 amino acids with a SAM domain and a PRMT5 domain. Multiple sequence alignments revealed that the putative sequence of As-PRMT1 protein was relatively conserved across species, especially in the SAM domain. As-PRMT1 is widely expressed during embryo development of A. sinica. This is followed by a dramatic upregulation after diapause termination and then downregulation from the nauplius stage. Furthermore, As-PRMT1 transcripts are highly upregulated under conditions of high salinity and low temperature stress. These findings suggested that As-PRMT1 is a stress-related factor that might promote or inhibit the expression of certain genes, play a critical role in embryonic development and in resistance to low temperature and high salinity stress. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. The History of the Austin College Building and Old Main at Sam Houston State University

    Science.gov (United States)

    Singer, Erin; Shields, Samantha

    2017-01-01

    Austin Hall and Old Main serve as the heart of what is now Sam Houston State University. The buildings' rich histories help one to understand how Sam Houston State University and its proud teacher education heritage came to be. To begin with Austin Hall's story, the University's original building has a unique and interesting tale that journeys…

  8. S.A.M., the Italian Martian Simulation Chamber

    Science.gov (United States)

    Galletta, G.; Ferri, F.; Fanti, G.; D'Alessandro, M.; Bertoloni, G.; Pavarin, D.; Bettanini, C.; Cozza, P.; Pretto, P.; Bianchini, G.; Debei, S.

    2006-12-01

    The Martian Environment Simulator (SAM “Simulatore di Ambiente Marziano”) is a interdisciplinary project of Astrobiology done at University of Padua. The research is aimed to the study of the survival of the microorganisms exposed to the “extreme” planetary environment. The facility has been designed in order to simulate Mars’ environmental conditions in terms of atmospheric pressure, temperature cycles and UV radiation dose. The bacterial cells, contained into dedicated capsules, will be exposed to thermal cycles simulating diurnal and seasonal Martian cycles. The metabolism of the different biological samples will be analysed at different phases of the experiment, to study their survival and eventual activity of protein synthesis (mortality, mutations and capability of DNA reparing). We describe the experimental facility and provide the perspectives of the biological experiments we will perform in order to provide hints on the possibility of life on Mars either autochthonous or imported from Earth.

  9. Cooperation between Magnesium and Metabolite Controls Collapse of the SAM-I Riboswitch.

    Science.gov (United States)

    Roy, Susmita; Onuchic, José N; Sanbonmatsu, Karissa Y

    2017-07-25

    The S-adenosylmethionine (SAM)-I riboswitch is a noncoding RNA that regulates the transcription termination process in response to metabolite (SAM) binding. The aptamer portion of the riboswitch may adopt an open or closed state depending on the presence of metabolite. Although the transition between the open and closed states is critical for the switching process, its atomistic details are not well understood. Using atomistic simulations, we calculate the effect of SAM and magnesium ions on the folding free energy landscape of the SAM-I riboswitch. These molecular simulation results are consistent with our previous wetlab experiments and aid in interpreting the SHAPE probing measurements. Here, molecular dynamics simulations explicitly identify target RNA motifs sensitive to magnesium ions and SAM. In the simulations, we observe that, whereas the metabolite mostly stabilizes the P1 and P3 helices, magnesium serves an important role in stabilizing a pseudoknot interaction between the P2 and P4 helices, even at high metabolite concentrations. The pseudoknot stabilization by magnesium, in combination with P1 stabilization by SAM, explains the requirement of both SAM and magnesium to form the fully collapsed metabolite-bound closed state of the SAM-I riboswitch. In the absence of SAM, frequent open-to-closed conformational transitions of the pseudoknot occur, akin to breathing. These pseudoknot fluctuations disrupt the binding site by facilitating fluctuations in the 5'-end of helix P1. Magnesium biases the landscape toward a collapsed state (preorganization) by coordinating pseudoknot and 5'-P1 fluctuations. The cooperation between SAM and magnesium in stabilizing important tertiary interactions elucidates their functional significance in transcription regulation. Published by Elsevier Inc.

  10. Systematic biochemical characterization of the SAM domains in Eph receptor family from Mus Musculus.

    Science.gov (United States)

    Wang, Yue; Li, Qingxia; Zheng, Yunhua; Li, Gang; Liu, Wei

    2016-05-13

    The Eph receptor family is the largest subfamily of receptor tyrosine kinases and well-known for their pivotal roles in axon guidance, synaptogenesis, artery/venous differentiation and tumorigenesis, etc. Activation of the Eph receptor needs multimerization of the receptors. The intracellular C-terminal SAM domain of Eph receptor was reported to mediate self-association of Eph receptors via the homo SAM-SAM interaction. In this study, we systematically expressed and purified the SAM domain proteins of all fourteen Eph receptors of Mus musculus in Escherichia coli. The FPLC (fast protein liquid chromatography) results showed the recombinant SAM domains were highly homogeneous. Using CD (circular dichroism) spectrometry, we found that the secondary structure of all the SAM domains was typically alpha helical folded and remarkably similar. The thermo-stability tests showed that they were quite stable in solution. SEC-MALS (size exclusion chromatography coupled with multiple angle light scattering) results illustrated 200 μM Eph SAM domains behaved as good monomers in the size-exclusion chromatography. More importantly, DLS (dynamic light scattering) results revealed the overwhelming majority of SAM domains was not multimerized in solution either at 200 μM or 2000 μM protein concentration, which indicating the SAM domain alone was not sufficient to mediate the polymerization of Eph receptor. In summary, our studies provided the systematic biochemical characterizations of the Eph receptor SAM domains and implied their roles in Eph receptor mediated signaling pathways. Copyright © 2016 Elsevier Inc. All rights reserved.

  11. Teleconnection stationarity, variability and trends of the Southern Annular Mode (SAM) during the last millennium

    Science.gov (United States)

    Dätwyler, Christoph; Neukom, Raphael; Abram, Nerilie J.; Gallant, Ailie J. E.; Grosjean, Martin; Jacques-Coper, Martín; Karoly, David J.; Villalba, Ricardo

    2017-11-01

    The Southern Annular Mode (SAM) is the leading mode of atmospheric interannual variability in the Southern Hemisphere (SH) extra-tropics. Here, we assess the stationarity of SAM spatial correlations with instrumental and paleoclimate proxy data for the past millennium. The instrumental period shows that temporal non-stationarities in SAM teleconnections are not consistent across the SH land areas. This suggests that the influence of the SAM index is modulated by regional effects. However, within key-regions with good proxy data coverage (South America, Tasmania, New Zealand), teleconnections are mostly stationary over the instrumental period. Using different stationarity criteria for proxy record selection, we provide new austral summer and annual mean SAM index reconstructions over the last millennium. Our summer SAM reconstructions are very robust to changes in proxy record selection and the selection of the calibration period, particularly on the multi-decadal timescale. In contrast, the weaker performance and lower agreement in the annual mean SAM reconstructions point towards changing teleconnection patterns that may be particularly important outside the summer months. Our results clearly portend that the temporal stationarity of the proxy-climate relationships should be taken into account in the design of comprehensive regional and hemispherical climate reconstructions. The summer SAM reconstructions show no significant relationship to solar, greenhouse gas and volcanic forcing, with the exception of an extremely strong negative anomaly following the AD 1257 Samalas eruption. Furthermore, reconstructed pre-industrial summer SAM trends are very similar to trends obtained by model control simulations. We find that recent trends in the summer SAM lie outside the 5-95% range of pre-industrial natural variability.

  12. Social activity method (SAM): A fractal language for mathematics

    Science.gov (United States)

    Dowling, Paul

    2013-09-01

    In this paper I shall present and develop my organisational language, social activity method (SAM), and illustrate some of its applications. I shall introduce a new scheme for modes of recontextualisation that enables the analysis of the ways in which one activity - which might be school mathematics or social research or any empirically observed regularity of practice - recontextualises the practice of another and I shall also present, deploy, and develop an existing scheme - domains of action - in an analysis of school mathematics examination papers and in the structuring of what I refer to as the esoteric domain. This domain is here conceived as a hybrid domain of, first, linguistic and extralinguistic resources that are unambiguously mathematical in terms of both expression and content and, second, pedagogic theory - often tacit - that enables the mathematical gaze onto other practices and so recontextualises them. A second and more general theme that runs through the paper is the claim that there is nothing that is beyond semiosis, that there is nothing to which we have direct access, unmediated by interpretation. This state of affairs has implications for mathematics education. Specifically, insofar as an individual's mathematical semiotic system is under continuous development - the curriculum never being graspable all at once - understanding - as a stable semiotic moment - of any aspect or object of mathematics is always localised to the individual and is at best transient, and the sequencing of such moments may well also be more individualised than consistent in some correspondence with the sequencing of the curriculum. This being the case, a concentration on understanding as a goal may well serve to inhibit the pragmatic acquisition and deployment of mathematical technologies, which should be the principal aim of mathematics teaching and learning. The paper is primarily concerned with mathematics education. SAM, however, is a language that is available for

  13. Economic Analysis Case Studies of Battery Energy Storage with SAM

    Energy Technology Data Exchange (ETDEWEB)

    DiOrio, Nicholas [National Renewable Energy Lab. (NREL), Golden, CO (United States); Dobos, Aron [National Renewable Energy Lab. (NREL), Golden, CO (United States); Janzou, Steven [National Renewable Energy Lab. (NREL), Golden, CO (United States)

    2015-11-01

    Interest in energy storage has continued to increase as states like California have introduced mandates and subsidies to spur adoption. This energy storage includes customer sited behind-the-meter storage coupled with photovoltaics (PV). This paper presents case study results from California and Tennessee, which were performed to assess the economic benefit of customer-installed systems. Different dispatch strategies, including manual scheduling and automated peak-shaving were explored to determine ideal ways to use the storage system to increase the system value and mitigate demand charges. Incentives, complex electric tariffs, and site specific load and PV data were used to perform detailed analysis. The analysis was performed using the free, publically available System Advisor Model (SAM) tool. We find that installation of photovoltaics with a lithium-ion battery system priced at $300/kWh in Los Angeles under a high demand charge utility rate structure and dispatched using perfect day-ahead forecasting yields a positive net-present value, while all other scenarios cost the customer more than the savings accrued. Different dispatch strategies, including manual scheduling and automated peak-shaving were explored to determine ideal ways to use the storage system to increase the system value and mitigate demand charges. Incentives, complex electric tariffs, and site specific load and PV data were used to perform detailed analysis. The analysis was performed using the free, publically available System Advisor Model (SAM) tool. We find that installation of photovoltaics with a lithium-ion battery system priced at $300/kWh in Los Angeles under a high demand charge utility rate structure and dispatched using perfect day-ahead forecasting yields a positive net-present value, while all other scenarios cost the customer more than the savings accrued.

  14. Isoforms of purified methyltransferase from human blood platelets ...

    African Journals Online (AJOL)

    A membrane-bound protein with N-methyltransferase activity, associated with phospholipid metabolism, has been isolated from purified human blood platelet plasma membranes. The activity of this enzyme has been detected in crude platelet preparations. However, the nature and properties of this enzyme and its ...

  15. Cloning and characterization of a γ-tocopherol methyltransferase ...

    African Journals Online (AJOL)

    ... reading frame of 1041 nucleotides encoding a protein of 39 kD polypeptide. A BoTMT whole cell system was developed for the production of α-tocopherol through the expression of BoTMT in Escherichia coli BL21 (DE3) strain. Keywords: γ-Tocopherol methyltransferase, Chinese cabbage, Perilla frustescens, tocopherol, ...

  16. Label-free electrochemical detection of human methyltransferase from tumors.

    Science.gov (United States)

    Furst, Ariel L; Muren, Natalie B; Hill, Michael G; Barton, Jacqueline K

    2014-10-21

    The role of abnormal DNA methyltransferase activity in the development and progression of cancer is an essential and rapidly growing area of research, both for improved diagnosis and treatment. However, current technologies for the assessment of methyltransferase activity, particularly from crude tumor samples, limit this work because they rely on radioactivity or fluorescence and require bulky instrumentation. Here, we report an electrochemical platform that overcomes these limitations for the label-free detection of human DNA(cytosine-5)-methyltransferase1 (DNMT1) methyltransferase activity, enabling measurements from crude cultured colorectal cancer cell lysates (HCT116) and biopsied tumor tissues. Our multiplexed detection system involving patterning and detection from a secondary electrode array combines low-density DNA monolayer patterning and electrocatalytically amplified DNA charge transport chemistry to measure selectively and sensitively DNMT1 activity within these complex and congested cellular samples. Based on differences in DNMT1 activity measured with this assay, we distinguish colorectal tumor tissue from healthy adjacent tissue, illustrating the effectiveness of this two-electrode platform for clinical applications.

  17. Linking electrostatic effects and protein motions in enzymatic catalysis. A theoretical analysis of catechol o-methyltransferase.

    Science.gov (United States)

    García-Meseguer, Rafael; Zinovjev, Kirill; Roca, Maite; Ruiz-Pernía, Javier J; Tuñón, Iñaki

    2015-01-22

    The role of protein motions in enzymatic catalysis is the subject of a hot scientific debate. We here propose the use of an explicit solvent coordinate to analyze the impact of environmental motions during the reaction process. The example analyzed here is the reaction catalyzed by catechol O-methyltransferase, a methyl transfer reaction from S-adenosylmethionine (SAM) to the nucleophilic oxygen atom of catecholate. This reaction proceeds from a charged reactant to a neutral product, and then a large electrostatic coupling with the environment could be expected. By means of a two-dimensional free energy surface, we show that a large fraction of the environmental motions needed to attain the transition state happens during the first stages of the reaction because most of the environmental motions are slower than changes in the substrate. The incorporation of the solvent coordinate in the definition of the transition state improves the transmission coefficient and the committor histogram in solution, while the changes are much less significant in the enzyme. The equilibrium solvation approach seems then to work better in the enzyme than in aqueous solution because the enzyme provides a preorganized environment where the reaction takes place.

  18. Understanding the Catalytic Mechanism of Xanthosine Methyltransferase in Caffeine Biosynthesis from QM/MM Molecular Dynamics and Free Energy Simulations.

    Science.gov (United States)

    Qian, Ping; Guo, Hao-Bo; Yue, Yufei; Wang, Liang; Yang, Xiaohan; Guo, Hong

    2016-09-26

    S-Adenosyl-l-methionine (SAM) dependent xanthosine methyltransferase (XMT) is the key enzyme that catalyzes the first methyl transfer in the caffeine biosynthesis pathway to produce the intermediate 7-methylxanthosine (7mXR). Although XMT has been a subject of extensive discussions, the catalytic mechanism and nature of the substrate involved in the catalysis are still unclear. In this paper, quantum mechanical/molecular mechanical (QM/MM) molecular dynamics (MD) and free energy (potential of mean force or PMF) simulations are undertaken to determine the catalytic mechanism of the XMT-catalyzed reaction. Both xanthosine and its monoanionic form with N3 deprotonated are used as the substrates for the methylation. It is found that while the methyl group can be transferred to the monoanionic form of xanthosine with a reasonable free energy barrier (about 17 kcal/mol), that is not the case for the neutral xanthosine. The results suggest that the substrate for the first methylation step in the caffeine biosynthesis pathway is likely to be the monoanionic form of xanthosine rather than the neutral form as widely adopted. This conclusion is supported by the pKa value on N3 of xanthosine both measured in aqueous phase and calculated in the enzymatic environment. The structural and dynamics information from both the X-ray structure and MD simulations is also consistent with the monoanionic xanthosine scenario. The implications of this conclusion for caffeine biosynthesis are discussed.

  19. Sam68 exerts separable effects on cell cycle progression and apoptosis

    Directory of Open Access Journals (Sweden)

    Resnick Ross J

    2004-01-01

    Full Text Available Abstract Background The RNA-binding protein Sam68 has been implicated in a number of cellular processes, including transcription, RNA splicing and export, translation, signal transduction, cell cycle progression and replication of the human immunodeficiency virus and poliovirus. However, the precise impact it has on essential cellular functions remains largely obscure. Results In this report we show that conditional overexpression of Sam68 in fibroblasts results in both cell cycle arrest and apoptosis. Arrest in G1 phase of the cell cycle is associated with decreased levels of cyclins D1 and E RNA and protein, resulting in dramatically reduced Rb phosphorylation. Interestingly, cell cycle arrest does not require the specific RNA binding ability of Sam68. In marked contrast, induction of apoptosis by Sam68 absolutely requires a fully-functional RNA binding domain. Moreover, the anti-cancer agent trichostatin A potentiates Sam68-driven apoptosis. Conclusions For the first time we have shown that Sam68, an RNA binding protein with multiple apparent functions, exerts functionally separable effects on cell proliferation and survival, dependent on its ability to bind specifically to RNA. These findings shed new light on the ability of signal transducing RNA binding proteins to influence essential cell function. Moreover, the ability of a class of anti-cancer therapeutics to modulate its ability to promote apoptosis suggests that Sam68 status may impact some cancer treatments.

  20. Automation of One-Loop Calculations with GoSam: Present Status and Future Outlook

    CERN Document Server

    Cullen, Gavin; Heinrich, Gudrun; Luisoni, Gionata; Mastrolia, Pierpaolo; Ossola, Giovanni; Reiter, Thomas; Tramontano, Francesco

    2011-01-01

    In this presentation, we describe the GoSam (Golem/Samurai) framework for the automated computation of multi-particle scattering amplitudes at the one-loop level. The amplitudes are generated analytically in terms of Feynman diagrams, and can be evaluated using either D-dimensional integrand reduction or tensor decomposition. GoSam can be used to compute one-loop corrections to Standard Model (QCD and EW) processes, and it is ready to link generic model files for theories Beyond SM. We show the main features of GoSam through its application to several examples of different complexity.

  1. Molecular identification of carnosine N-methyltransferase as chicken histamine N-methyltransferase-like protein (hnmt-like.

    Directory of Open Access Journals (Sweden)

    Jakub Drozak

    Full Text Available Anserine (beta-alanyl-N(Pi-methyl-L-histidine, a naturally occurring derivative of carnosine (beta-alanyl-L-histidine, is an abundant constituent of skeletal muscles and brain of many vertebrates. Although it has long been proposed to serve as a proton buffer, radicals scavenger and transglycating agent, its physiological function remains obscure. The formation of anserine is catalyzed by carnosine N-methyltransferase which exhibits unknown molecular identity. In the present investigation, we have purified carnosine N-methyltransferase from chicken pectoral muscle about 640-fold until three major polypeptides of about 23, 26 and 37 kDa coeluting with the enzyme were identified in the preparation. Mass spectrometry analysis of these polypeptides resulted in an identification of histamine N-methyltransferase-like (HNMT-like protein as the only meaningful candidate. Analysis of GenBank database records indicated that the hnmt-like gene might be a paralogue of histamine N-methyltransferase gene, while comparison of their protein sequences suggested that HNMT-like protein might have acquired a new activity. Chicken HNMT-like protein was expressed in COS-7 cells, purified to homogeneity, and shown to catalyze the formation of anserine as confirmed by both chromatographic and mass spectrometry analysis. Both specificity and kinetic studies carried out on the native and recombinant enzyme were in agreement with published data. Particularly, several compounds structurally related to carnosine, including histamine and L-histidine, were tested as potential substrates for the enzyme, and carnosine was the only methyl group acceptor. The identification of the gene encoding carnosine N-methyltransferase might be beneficial for estimation of the biological functions of anserine.

  2. 1 Protein Methyltransferases: Their Distribution Among the Five Structural Classes of AdoMet-Dependent Methyltransferases.

    Science.gov (United States)

    Schubert, Heidi L; Blumenthal, Robert M; Cheng, Xiaodong

    2006-01-01

    S-adenosyl-l-methionine (AdoMet) dependent methyltransferases (MTases) are involved in biosynthesis, signal transduction, protein repair, chromatin regulation, and gene silencing. Five different structural folds (designated I through V) have been described that bind AdoMet and catalyze methyltransfer to diverse substrates, although the great majority of known MTases have the Class I fold. Even within a particular MTase class the amino-acid sequence similarity can be as low as 10%. Thus, the structural and catalytic requirements for methyltransfer from AdoMet appear to be remarkably flexible. MTases that act on protein substrates have been found to date among three of the five structural classes (I, the classical fold; III, the corrin MTase fold; and V, the SET fold). "There are many paths to the top of the mountain, but the view is always the same."-Chinese proverb The Columbia World of Quotations, New York, Columbia University Press, 1996. Copyright © 2006 Elsevier Inc. All rights reserved.

  3. Must, valge + hall : Sam Wagstaffi ja Robert Mapplethorpe'i portree / Madis Palm

    Index Scriptorium Estoniae

    Palm, Madis

    2008-01-01

    Dokumentaalfilm "Black, White & Gray: Sam Wagstaff and Robert Mapplethorpe" : autor ja režissöör James Crump : Ameerika Ühendriigid 2007. Filmi näidati filminädala "Art in America" raames Tallinnas

  4. Smoking-Attributable Mortality, Morbidity, and Economic Costs (SAMMEC) - Smoking-Attributable Mortality (SAM)

    Data.gov (United States)

    U.S. Department of Health & Human Services — 2005-2009. SAMMEC - Smoking-Attributable Mortality, Morbidity, and Economic Costs. Smoking-attributable mortality (SAM) is the number of deaths caused by cigarette...

  5. Stability of binary SAMs formed by omega-acid and alcohol functionalized thiol mixtures.

    Science.gov (United States)

    Tielens, Frederik; Humblot, Vincent; Pradier, Claire-Marie; Calatayud, Monica; Illas, Francesc

    2009-09-01

    A simple model is presented to describe the mixing process of acid- and alcohol-terminated thiol self-assembled monolayers (SAMs) on Au(111). It was found that the low-concentration acid-terminated SAMs are enriched in acid thiol compared with the original solution from which the SAM is made. Due to the relatively strong interaction between acid and alcohol head groups, homogeneously mixed SAMs showing alcohol/acid pairs are preferred when the acid-terminated thiol fraction in solution is below or equal to 50%. This particular behavior affecting the mixing process is explained using atomistic first-principle thermodynamics. The calculated phase diagram has been discussed and interpreted with the help of XPS.

  6. Bringing a Chemical Laboratory Named Sam to Mars on the 2011 Curiosity Rover

    Science.gov (United States)

    Mahaffy, P. R.; Bleacher, L.; Jones, A.; Atreya, S. K.; Manning, H. L.; Cabane, M.; Webster, C. R.; Sam Team

    2010-12-01

    Introduction: An important goal of upcoming missions to Mars is to understand if life could have developed there. The task of the Sample Analysis at Mars (SAM) suite of instruments [1] and the other Curiosity investigations [2] is to move us steadily toward that goal with an assessment of the habitability of our neighboring planet through a series of chemical and geological measurements. SAM is designed to search for organic compounds and inorganic volatiles and measure isotope ratios. Other instruments on Curiosity will provide elemental analysis and identify minerals. SAM will analyze both atmospheric samples and gases evolved from powdered rocks that may have formed billions of years ago with Curiosity providing access to interesting sites scouted by orbiting cameras and spectrometers. SAM Instrument Suite: SAM’s instruments are a Quadrupole Mass Spectrometer (QMS), a 6-column Gas Chromatograph (GC), and a 2-channel Tunable Laser Spectrometer (TLS). SAM can identify organic compounds in Mars rocks to sub-ppb sensitivity and secure precise isotope ratios for C, H, and O in carbon dioxide and water and measure trace levels of methane and its carbon 13 isotope. The SAM gas processing system consists of valves, heaters, pressure sensors, gas scrubbers and getters, traps, and gas tanks used for calibration or combustion experiments [2]. A variety of calibrant compounds interior and exterior to SAM will allow the science and engineering teams to assess SAM’s performance. SAM has been calibrated and tested in a Mars-like environment. Keeping Educators and the Public Informed: The Education and Public Outreach (EPO) goals of the SAM team are to make this complex chemical laboratory and its data widely available to educators, students, and the public. Formal education activities include developing templates for professional development workshops for educators to teach them about SAM and Curiosity, incorporating data into Mars Student Data Teams, and writing articles

  7. Identification of trans-acting factors regulating SamDC expression in Oryza sativa

    Energy Technology Data Exchange (ETDEWEB)

    Basu, Supratim, E-mail: supratim_genetics@yahoo.co.in [Department of Crop Soil and Environmental Sciences, University of Arkansas, Fayetteville, AR 72701 (United States); Division of Plant Biology, Bose Institute, Kolkata (India); Roychoudhury, Aryadeep [Post Graduate Department of Biotechnology, St. Xavier' s College (Autonomous), 30, Mother Teresa Sarani, Kolkata - 700016, West Bengal (India); Sengupta, Dibyendu N. [Division of Plant Biology, Bose Institute, Kolkata (India)

    2014-03-07

    Highlights: • Identification of cis elements responsible for SamDC expression by in silico analysis. • qPCR analysis of SamDC expression to abiotic and biotic stress treatments. • Detection of SamDC regulators using identified cis-elements as probe by EMSA. • Southwestern Blot analysis to predict the size of the trans-acting factors. - Abstract: Abiotic stress affects the growth and productivity of crop plants; to cope with the adverse environmental conditions, plants have developed efficient defense machinery comprising of antioxidants like phenolics and flavonoids, and osmolytes like polyamines. SamDC is a key enzyme in the polyamine biosynthesis pathway in plants. In our present communication we have done in silico analysis of the promoter region of SamDC to look for the presence of different cis-regulatory elements contributing to its expression. Based on the presence of different cis-regulatory elements we completed comparative analysis of SamDC gene expression in rice lamina of IR-29 and Nonabokra by qPCR in response to the abiotic stress treatments of salinity, drought, cold and the biotic stress treatments of ABA and light. Additionally, to explore the role of the cis-regulatory elements in regulating the expression of SamDC gene in plants we comparatively analyzed the binding of rice nuclear proteins prepared from IR-29 and Nonabokra undergoing various stress treatments. The intensity of the complex formed was low and inducible in IR-29 in contrast to Nonabokra. Southwestern blot analysis helped in predicting the size of the trans-acting factors binding to these cis-elements. To our knowledge this is the first report on the comprehensive analysis of SamDC gene expression in rice and identification of the trans-acting factors regulating its expression.

  8. The Combustion Experiment on the Sample Analysis at Mars (SAM) Instrument Suite on the Curiosity Rover

    Science.gov (United States)

    Stern, J. C.; Malespin, C. A.; Eigenbrode, J. L.; Graham, H. V.; Archer, P. D., Jr.; Brunner, A. E.; Freissinet, C.; Franz, H. B.; Fuentes, J.; Glavin, D. P.; hide

    2014-01-01

    The combustion experiment on the Sample Analysis at Mars (SAM) suite on Curiosity will heat a sample of Mars regolith in the presence of oxygen and measure composition of the evolved gases using quadrupole mass spectrometry (QMS) and tunable laser spectrometry (TLS). QMS will enable detection of combustion products such as CO, CO2, NO, and other oxidized species, while TLS will enable precise measurements of the abundance and carbon isotopic composition (delta(sup 13)C) of the evolved CO2 and hydrogen isotopic composition (deltaD) of H2O. SAM will perform a two-step combustion to isolate combustible materials below approx.550 C and above approx.550 C. The combustion experiment on SAM, if properly designed and executed, has the potential to answer multiple questions regarding the origins of volatiles seen thus far in SAM evolved gas analysis (EGA) on Mars. Constraints imposed by SAM and MSL time and power resources, as well as SAM consumables (oxygen gas), will limit the number of SAM combustion experiments, so it is imperative to design an experiment targeting the most pressing science questions. Low temperature combustion experiments will primarily target the quantification of carbon (and nitrogen) contributed by SAM wet chemistry reagants MTBSTFA (N-Methyl-N-tert-butyldimethylsilyltrifluoroacetamide) and DMF (Dimethylformamide), which have been identified in the background of blank and sample runs and may adsorb to the sample while the cup is in the Sample Manipulation System (SMS). In addition, differences between the sample and "blank" may yield information regarding abundance and delta(sup 13)C of bulk (both organic and inorganic) martian carbon. High temperature combustion experiments primarily aim to detect refractory organic matter, if present in Cumberland fines, as well as address the question of quantification and deltaD value of water evolution associated with hydroxyl hydrogen in clay minerals.

  9. Benchmark Simulation of Natural Circulation Cooling System with Salt Working Fluid Using SAM

    Energy Technology Data Exchange (ETDEWEB)

    Ahmed, K. K.; Scarlat, R. O.; Hu, R.

    2017-09-03

    Liquid salt-cooled reactors, such as the Fluoride Salt-Cooled High-Temperature Reactor (FHR), offer passive decay heat removal through natural circulation using Direct Reactor Auxiliary Cooling System (DRACS) loops. The behavior of such systems should be well-understood through performance analysis. The advanced system thermal-hydraulics tool System Analysis Module (SAM) from Argonne National Laboratory has been selected for this purpose. The work presented here is part of a larger study in which SAM modeling capabilities are being enhanced for the system analyses of FHR or Molten Salt Reactors (MSR). Liquid salt thermophysical properties have been implemented in SAM, as well as properties of Dowtherm A, which is used as a simulant fluid for scaled experiments, for future code validation studies. Additional physics modules to represent phenomena specific to salt-cooled reactors, such as freezing of coolant, are being implemented in SAM. This study presents a useful first benchmark for the applicability of SAM to liquid salt-cooled reactors: it provides steady-state and transient comparisons for a salt reactor system. A RELAP5-3D model of the Mark-1 Pebble-Bed FHR (Mk1 PB-FHR), and in particular its DRACS loop for emergency heat removal, provides steady state and transient results for flow rates and temperatures in the system that are used here for code-to-code comparison with SAM. The transient studied is a loss of forced circulation with SCRAM event. To the knowledge of the authors, this is the first application of SAM to FHR or any other molten salt reactors. While building these models in SAM, any gaps in the code’s capability to simulate such systems are identified and addressed immediately, or listed as future improvements to the code.

  10. Pengaruh Kompetensi Pegawai terhadap Peningkatan Kualitas Pelayanan Publik di Kantor Pusat Universitas Sam Ratulangi Ma

    OpenAIRE

    Tulusan, Femmy M; RARES, Joyce J; Lumi, Djefry

    2015-01-01

    Sam Ratulangi University as one of the major universities are very concerned will be given public service employees, to the competence of the employees is a problem that is considered by the leadership, because without a good employee competence given the public service will not be maximized. The research methodology used is that while the survey is descriptive analytical method. The target population in this study is that there are employees in the Central Office and the University of Sam Ra...

  11. Fabrication and Surface Properties of Composite Films of SAM/Pt/ZnO/SiO 2

    KAUST Repository

    Yao, Ke Xin

    2008-12-16

    Through synthetic architecture and functionalization with self-assembled monolayers (SAMs), complex nanocomposite films of SAM/Pt/ZnO/SiO2 have been facilely prepared in this work. The nanostructured films are highly uniform and porous, showing a wide range of tunable wettabilities from superhydrophilicity to superhydrophobicity (water contact angles: 0° to 170°). Our approach offers synthetic flexibility in controlling film architecture, surface topography, coating texture, crystallite size, and chemical composition of modifiers (e.g., SAMs derived from alkanethiols). For example, wettability properties of the nanocomposite films can be finely tuned with both inorganic phase (i.e., ZnO/SiO2 and Pt/ZnO/SiO2) and organic phase (i.e., SAMs on Pt/ZnO/SiO2). Due to the presence of catalytic components Pt/ZnO within the nanocomposites, surface reactions of the organic modifiers can further take place at room temperature and elevated temperatures, which provides a means for SAM formation and elimination. Because the Pt/ZnO forms an excellent pair of metal-semiconductors for photocatalysis, the anchored SAMs can also be modified or depleted by UV irradiation (i.e., the films possess self-cleaning ability). Potential applications of these nanocomposite films have been addressed. Our durability tests also confirm that the films are thermally stable and structurally robust in modification- regeneration cycles. © 2008 American Chemical Society.

  12. Plant isoflavone and isoflavanone O-methyltransferase genes

    Science.gov (United States)

    Broeckling, Bettina E.; Liu, Chang-Jun; Dixon, Richard A.

    2014-08-19

    The invention provides enzymes that encode O-methyltransferases (OMTs) from Medicago truncatula that allow modification to plant (iso)flavonoid biosynthetic pathways. In certain aspects of the invention, the genes encoding these enzymes are provided. The invention therefore allows the modification of plants for isoflavonoid content. Transgenic plants comprising such enzymes are also provided, as well as methods for improving disease resistance in plants. Methods for producing food and nutraceuticals, and the resulting compositions, are also provided.

  13. Histone H3 lysine 4 methyltransferase KMT2D.

    Science.gov (United States)

    Froimchuk, Eugene; Jang, Younghoon; Ge, Kai

    2017-09-05

    Histone-lysine N-methyltransferase 2D (KMT2D), also known as MLL4 and MLL2 in humans and Mll4 in mice, belongs to a family of mammalian histone H3 lysine 4 (H3K4) methyltransferases. It is a large protein over 5500 amino acids in size and is partially functionally redundant with KMT2C. KMT2D is widely expressed in adult tissues and is essential for early embryonic development. The C-terminal SET domain is responsible for its H3K4 methyltransferase activity and is necessary for maintaining KMT2D protein stability in cells. KMT2D associates with WRAD (WDR5, RbBP5, ASH2L, and DPY30), NCOA6, PTIP, PA1, and H3K27 demethylase UTX in one protein complex. It acts as a scaffold protein within the complex and is responsible for maintaining the stability of UTX. KMT2D is a major mammalian H3K4 mono-methyltransferase and co-localizes with lineage determining transcription factors on transcriptional enhancers. It is required for the binding of histone H3K27 acetyltransferases CBP and p300 on enhancers, enhancer activation and cell-type specific gene expression during differentiation. KMT2D plays critical roles in regulating development, differentiation, metabolism, and tumor suppression. It is frequently mutated in developmental diseases, such as Kabuki syndrome and congenital heart disease, and various forms of cancer. Further understanding of the mechanism through which KMT2D regulates gene expression will reveal why KMT2D mutations are so harmful and may help generate novel therapeutic approaches. Published by Elsevier B.V.

  14. Structural characterization of the mitomycin 7-O-methyltransferase.

    Science.gov (United States)

    Singh, Shanteri; Chang, Aram; Goff, Randal D; Bingman, Craig A; Grüschow, Sabine; Sherman, David H; Phillips, George N; Thorson, Jon S

    2011-07-01

    Mitomycins are quinone-containing antibiotics, widely used as antitumor drugs in chemotherapy. Mitomycin-7-O-methyltransferase (MmcR), a key tailoring enzyme involved in the biosynthesis of mitomycin in Streptomyces lavendulae, catalyzes the 7-O-methylation of both C9β- and C9α-configured 7-hydroxymitomycins. We have determined the crystal structures of the MmcR-S-adenosylhomocysteine (SAH) binary complex and MmcR-SAH-mitomycin A (MMA) ternary complex at resolutions of 1.9and 2.3 Å, respectively. The study revealed MmcR to adopt a common S-adenosyl-L-methionine-dependent O-methyltransferase fold and the presence of a structurally conserved active site general acid-base pair is consistent with a proton-assisted methyltransfer common to most methyltransferases. Given the importance of C7 alkylation to modulate mitomycin redox potential, this study may also present a template toward the future engineering of catalysts to generate uniquely bioactive mitomycins. Copyright © 2011 Wiley-Liss, Inc.

  15. Structural characterization of the mitomycin 7-O-methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Shanteri; Chang, Aram; Goff, Randal D.; Bingman, Craig A.; Grüschow, Sabine; Sherman, David H.; Phillips, Jr., George N.; Thorson, Jon S. (Michigan); (UW)

    2014-10-02

    Mitomycins are quinone-containing antibiotics, widely used as antitumor drugs in chemotherapy. Mitomycin-7-O-methyltransferase (MmcR), a key tailoring enzyme involved in the biosynthesis of mitomycin in Streptomyces lavendulae, catalyzes the 7-O-methylation of both C9{beta}- and C9{alpha}-configured 7-hydroxymitomycins. We have determined the crystal structures of the MmcR-S-adenosylhomocysteine (SAH) binary complex and MmcR-SAH-mitomycin A (MMA) ternary complex at resolutions of 1.9 and 2.3 {angstrom}, respectively. The study revealed MmcR to adopt a common S-adenosyl-L-methionine-dependent O-methyltransferase fold and the presence of a structurally conserved active site general acid-base pair is consistent with a proton-assisted methyltransfer common to most methyltransferases. Given the importance of C7 alkylation to modulate mitomycin redox potential, this study may also present a template toward the future engineering of catalysts to generate uniquely bioactive mitomycins.

  16. SAM-Like Evolved Gas Analyses of Phyllosilicate Minerals and Applications to SAM Analyses of the Sheepbed Mudstone, Gale Crater, Mars

    Science.gov (United States)

    McAdam, A. C.; Franz, H. B.; Mahaffy, P. R.; Eigenbrode, J. L.; Stern, J. C.; Brunner, B.; Sutter, B.; Archer, P. D.; Ming , D. W.; Morris, R. V.; hide

    2014-01-01

    While in Yellowknife Bay, the Mars Science Laboratory Curiosity rover collected two drilled samples, John Klein (hereafter "JK") and Cumberland ("CB"), from the Sheepbed mudstone, as well as a scooped sample from the Rocknest aeolian bedform ("RN"). These samples were sieved by Curiosity's sample processing system and then several subsamples of these materials were delivered to the Sample Analysis at Mars (SAM) instrument suite and the CheMin X-ray diffraction/X-ray fluorescence instrument. CheMin provided the first in situ X-ray diffraction-based evidence of clay minerals on Mars, which are likely trioctahedral smectites (e.g., Fe-saponite) and comprise 20 wt% of the mudstone samples [1]. SAM's evolved gas analysis (EGA) mass spectrometry analyses of JK and CB subsamples, as well as RN subsamples, detected H2O, CO2, O2, H2, SO2, H2S, HCl, NO, OCS, CS2 and other trace gases evolved during pyrolysis. The identity of evolved gases and temperature( s) of evolution can augment mineral detection by CheMin and place constraints on trace volatile-bearing phases present below the CheMin detection limit or those phases difficult to characterize with XRD (e.g., X-ray amorphous phases). Here we will focus on the SAM H2O data, in the context of CheMin analyses, and comparisons to laboratory SAM-like analyses of several phyllosilicate minerals including smectites.

  17. Drug delivery from therapeutic self-assembled monolayers (T-SAMs) on 316L stainless steel.

    Science.gov (United States)

    Mahapatro, Anil; Johnson, Dave M; Patel, Devang N; Feldman, Marc D; Ayon, Arturo A; Agrawal, C Mauli

    2008-01-01

    Delivery of therapeutic agents from self-assembled monolayers (SAMs) on 316L stainless steel (SS) has been demonstrated as a viable method to deliver drugs for localized coronary artery stent application. SAMs are highly-ordered, nano-sized molecular coatings, adding 1-10 nm thickness to a surface. Hydroxyl terminated alkanethiol SAMs of 11-mercapto-1-undecanol (-OH SAM) were formed on 316L SS with 48 hr immersion in ethanolic solutions. Attachment of ibuprofen (a model drug) to the functional SAMs was carried out in toluene for 5 hrs at 60 degrees C using Novozume-435 as a biocatalyst. SAM formation and subsequent attachment of ibuprofen was characterized collectively using X-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy (FTIR), and contact angle (CA) measure-ments. The quantitative in vitro release of ibuprofen into a "physiological" buffer solution was characterized using reverse phase HPLC. Drug release kinetics showed that 14.1 microg of ibuprofen eluted over a period of 35 days with 2.7microg being eluted in the first day and the remaining being eluted over a period of 35 days. The drug release kinetics showed an increase in ibuprofen elution that occurred during first 14 days (2.7microg in 1 day to 9.5 microg in 14 days), following which there was a decrease in the rate of elution. Thus, functional SAMs on 316L SS could be used as tethers for drug attachment and could serve as a drug delivery mechanism from stainless steel implants such as coronary artery stents.

  18. Regulation of Catechol-O-Methyltransferase Expression in Human Myometrial Cells

    Science.gov (United States)

    Wentz, Melissa J.; Jamaluddin, Mohammad; Garfield, Robert E.; Al-Hendy, Ayman

    2014-01-01

    OBJECTIVE The catechol-O-methyltransferase enzyme catalyzes the methylation of the catechol estrogens, 2- or 4-hydroxyestrogen, to 2- or 4-methoxyestrogen. Both the hydroxy estrogens and methoxy estrogens were shown to modulate the effects of estrogen. Because catechol-O-methyltransferase activity controls levels of these metabolites, it may help regulate the cellular estrogenic milieu. In this study, we examined the regulation of catechol-O-methyltransferase expression in human myometrial cells. METHODS Catechol-O-methyltransferase expression was assessed by reverse transcription–polymerase chain reaction, Western blot, and luciferase assays in human myometrial cells after treatment with estrogen or progesterone. Catechol-O-methyltransferase expression was measured in cells after treatment with tumor necrosis factor alpha (TNFα) alone or with lactacystin, a protea-some inhibitor. Luciferase assays were also conducted using human myometrial cells containing an estrogen response element–luciferase reporter gene to measure levels of estrogen-mediated transactivation after treatment with estrogen and increasing concentrations of 2-hydroxestrogen. RESULTS Catechol-O-methyltransferase expression was down-regulated by progesterone or estrogen. Tumor necrosis factor alpha upregulated catechol-O-methyltransferase expression, whereas cotreatment with lactacystin attenuated this response, suggesting that TNFα activated nuclear factor kappa B to induce catechol-O-methyltransferase expression. Increased concentrations of 2-hydroxyestrogen attenuated estrogen-mediated transcription in the myometrial cells. CONCLUSION Catechol-O-methyltransferase expression may be regulated in the myometrium to control the local action of estrogen. Low levels of catechol-O-methyltransferase in the myometrium would result in an accumulation of 2-hydroxyestrogen and may antagonize the local effect of estrogen. High levels of catechol-O-methyltransferase in the myometrium would result in

  19. Systematic analysis of O-methyltransferase gene family and identification of potential members involved in the formation of O-methylated flavonoids in Citrus.

    Science.gov (United States)

    Liu, Xiaogang; Luo, Yan; Wu, Hongkun; Xi, Wanpeng; Yu, Jie; Zhang, Qiuyun; Zhou, Zhiqin

    2016-01-10

    The O-methylation of various secondary metabolites is mainly catalyzed by S-adenosyl-l-methionine (SAM)-dependent O-methyltransferase (OMT) proteins that are encoded by the O-methyltransferase gene family. Citrus fruits are a rich source of O-methylated flavonoids that have a broad spectrum of biological activities, including anti-inflammatory, anticarcinogenic, and antiatherogenic properties. However, little is known about this gene family and its members that are involved in the O-methylation of flavonoids and their regulation in Citrus. In this study, 58 OMT genes were identified from the entire Citrus sinensis genome and compared with those from 3 other representative dicot plants. A comprehensive analysis was performed, including functional/substrate predictions, identification of chromosomal locations, phylogenetic relationships, gene structures, and conserved motifs. Distribution mapping revealed that the 58 OMT genes were unevenly distributed on the 9 citrus chromosomes. Phylogenetic analysis of 164 OMT proteins from C.sinensis, Arabidopsis thaliana, Populus trichocarpa, and Vitis vinifera showed that these proteins were categorized into group I (COMT subfamily) and group II (CCoAOMT subfamily), which were further divided into 10 and 2 subgroups, respectively. Finally, digital gene expression and quantitative real-time polymerase chain reaction analyses revealed that citrus OMT genes had distinct temporal and spatial expression patterns in different tissues and developmental stages. Interestingly, 18 and 11 of the 27 genes predicted to be involved in O-methylation of flavonoids had higher expression in the peel and pulp during fruit development, respectively. The citrus OMT gene family identified in this study might help in the selection of appropriate candidate genes and facilitate functional studies in Citrus. Copyright © 2015 Elsevier B.V. All rights reserved.

  20. Celiac Disease in Children with Severe Acute Malnutrition (SAM): A Hospital Based Study.

    Science.gov (United States)

    Beniwal, Neetu; Ameta, Gaurav; Chahar, Chandra Kumar

    2017-05-01

    To evaluate the prevalence and clinical features of Celiac disease among children with severe acute malnutrition (SAM). This prospective observational study was conducted in PBM Children Hospital, Bikaner from July 2012 through December 2013. All consecutively admitted children with SAM were recruited. All subjects were screened for Celiac disease by serological test for IgA-anti tissue Transglutaminase (IgA tTG) antibodies. All seropositive children underwent upper gastrointestinal endoscopy for small bowel biopsy for the confirmation. Clinical features of patients with and without celiac disease were compared. The sero-prevalence (IgA tTg positivity) of Celiac disease was found to be 15.38% while prevalence of biopsy confirmed Celiac disease was 14.42% among SAM children. Abdominal distension, diarrhea, anorexia, constipation, pain in abdomen, vitamin deficiencies, edema, clubbing and mouth ulcers were more common in patients of Celiac disease compared to patients without Celiac disease but the difference was statistically significant only for abdominal distension and pain abdomen. There is a high prevalence of Celiac disease in SAM. Screening for Celiac disease (especially in presence of pain abdomen and abdominal distension) should be an essential part of work-up in all children with SAM.

  1. The relative influence of ENSO and SAM on Antarctic Peninsula climate

    Science.gov (United States)

    Clem, Kyle R.; Renwick, James A.; McGregor, James; Fogt, Ryan L.

    2016-08-01

    Recent warming of the Antarctic Peninsula during austral autumn, winter, and spring has been linked to sea surface temperature (SST) trends in the tropical Pacific and tropical Atlantic, while warming of the northeast Peninsula during summer has been linked to a strengthening of westerly winds traversing the Peninsula associated with a positive trend in the Southern Annular Mode (SAM). Here we demonstrate that circulation changes associated with the SAM dominate interannual temperature variability across the entire Antarctic Peninsula during both summer and autumn, while relationships with tropical Pacific SST variability associated with the El Niño-Southern Oscillation (ENSO) are strongest and statistically significant primarily during winter and spring only. We find the ENSO-Peninsula temperature relationship during autumn to be weak on interannual time scales and regional circulation anomalies associated with the SAM more important for interannual temperature variability across the Peninsula during autumn. Consistent with previous studies, western Peninsula temperatures during autumn, winter, and spring are closely tied to changes in the Amundsen Sea Low (ASL) and associated meridional wind anomalies. The interannual variability of ASL depth is most strongly correlated with the SAM index during autumn, while the ENSO relationship is strongest during winter and spring. Investigation of western and northeast Peninsula temperatures separately reveals that interannual variability of northeast Peninsula temperatures is primarily sensitive to zonal wind anomalies crossing the Peninsula and resultant leeside adiabatic warming rather than to meridional wind anomalies, which is closely tied to variability in the zonal portion of the SAM pattern.

  2. A case study of inaccuracy of advanced SAM application in the OPR1000

    Energy Technology Data Exchange (ETDEWEB)

    Roh, Kyung Ho; Yang, Sung Tae; Jung, Ji Eun [Korea Hydro and Nuclear Power Co., Daejeon (Korea, Republic of)

    2011-05-15

    KHNP has developed a constrained simulated annealing method to account for the weakness of the least square method when using the SAM calculation for applications related to the OPR1000. The SAM is generated and installed once at the beginning of each cycle. Thus, the accuracy of ASI simulation by CPC is wholly dependent on the accuracy of the SAM constants. As the depletion proceeds, the deviation of the CPC axial power shape increases. When noise is included, the measurement data frequently exceeds the limit (8%) of the CPC ASI deviation. Therefore, KHNP has developed a constrained simulated annealing method to filter the noise from the measurement data and give the SAM a physical meaning. These two objectives form the core of the developed method. The least square method makes it difficult to implement the algorithm for filtering the noise. Thus, a mathematical method that has been used and verified in a variety of fields was adopted for the simulated annealing . In this paper, the relationship between the SAM and CPC axial power shape deviation is analyzed, based on results showing that EOC CPC axial power shape deviation has approached the limit (8%) in the OPR1000 when the simulated annealing method was applied

  3. Epistatic and functional interactions of catechol-o-methyltransferase (COMT and AKT1 on neuregulin1-ErbB signaling in cell models.

    Directory of Open Access Journals (Sweden)

    Yoshitatsu Sei

    2010-05-01

    Full Text Available Neuregulin1 (NRG1-ErbB signaling has been implicated in the pathogenesis of cancer and schizophrenia. We have previously reported that NRG1-stimulated migration of B lymphoblasts is PI3K-AKT1dependent and impaired in patients with schizophrenia and significantly linked to the catechol-o-methyltransferase (COMT Val108/158Met functional polymorphism.We have now examined AKT1 activation in NRG1-stimulated B lymphoblasts and other cell models and explored a functional relationship between COMT and AKT1. NRG1-induced AKT1 phosphorylation was significantly diminished in Val carriers compared to Met carriers in both normal subjects and in patients. Further, there was a significant epistatic interaction between a putatively functional coding SNP in AKT1 (rs1130233 and COMT Val108/158Met genotype on AKT1 phosphorylation. NRG1 induced translocation of AKT1 to the plasma membrane also was impaired in Val carriers, while PIP(3 levels were not decreased. Interestingly, the level of COMT enzyme activity was inversely correlated with the cells' ability to synthesize phosphatidylserine (PS, a factor that attracts the pleckstrin homology domain (PHD of AKT1 to the cell membrane. Transfection of SH-SY5Y cells with a COMT Val construct increased COMT activity and significantly decreased PS levels as well as NRG1-induced AKT1 phosphorylation and migration. Administration of S-adenosylmethionine (SAM rescued all of these deficits. These data suggest that AKT1 function is influenced by COMT enzyme activity through competition with PS synthesis for SAM, which in turn dictates AKT1-dependent cellular responses to NRG1-mediated signaling.Our findings implicate genetic and functional interactions between COMT and AKT1 and may provide novel insights into pathogenesis of schizophrenia and other ErbB-associated human diseases such as cancer.

  4. The Order Bacillales Hosts Functional Homologs of the Worrisome cfr Antibiotic Resistance Gene

    DEFF Research Database (Denmark)

    Hansen, Lykke H.; Planellas, Mercè H.; Long, Katherine S.

    2012-01-01

    The cfr gene encodes the Cfr methyltransferase that methylates a single adenine in the peptidyl transferase region of bacterial ribosomes. The methylation provides resistance to several classes of antibiotics that include drugs of clinical and veterinary importance. This paper describes a first...... coli, and MICs for selected antibiotics indicate that the cfr-like genes confer resistance to PhLOPSa (phenicol, lincosamide, oxazolidinone, pleuromutilin, and streptogramin A) antibiotics in the same way as the cfr gene. In addition, modification at A2503 on 23S rRNA was confirmed by primer extension...

  5. Negative in vitro selection identifies the rRNA recognition motif for ErmE methyltransferase

    DEFF Research Database (Denmark)

    Nielsen, A K; Douthwaite, S; Vester, B

    1999-01-01

    Erm methyltransferases modify bacterial 23S ribosomal RNA at adenosine 2058 (A2058, Escherichia coli numbering) conferring resistance to macrolide, lincosamide, and streptogramin B (MLS) antibiotics. The motif that is recognized by Erm methyltransferases is contained within helix 73 of 23S r...

  6. Association of myasthenia gravis with polymorphisms in the gene of histamine N-methyltransferase

    DEFF Research Database (Denmark)

    Kellermayer, Blanka; Polgar, Noemi; Pal, Jozsef

    2013-01-01

    Histamine N-methyltransferase (HNMT) is the main metabolizing enzyme of histamine. Histamine modulates immune responses and plays a role in the pathogenesis of autoimmune disorders.......Histamine N-methyltransferase (HNMT) is the main metabolizing enzyme of histamine. Histamine modulates immune responses and plays a role in the pathogenesis of autoimmune disorders....

  7. The MarR family transcription factor Rv1404 coordinates adaptation of Mycobacterium tuberculosis to acid stress via controlled expression of Rv1405c, a virulence-associated methyltransferase.

    Science.gov (United States)

    Healy, Claire; Golby, Paul; MacHugh, David E; Gordon, Stephen V

    2016-03-01

    Coordinated regulation of gene expression is essential for pathogen adaptation in vivo. Understanding the control of these virulence circuits in the TB pathogen Mycobacterium tuberculosis is a key challenge if we are to increase our basic understanding of how this organism establishes infection. In this study we focused on the transcriptional regulator Rv1404 that shows similarity to the MarR family of transcriptional repressors. Rv1404 derepresses a set of genes in vivo that have been implicated in virulence and may therefore allow adaptation of M. tuberculosis to the intracellular environment. We used a combination of ChIP-qPCR and Electromobility Band Shift Assays (EMSA) to show that Rv1404 coordinates gene expression in response to stresses such as low pH in M. tuberculosis. Two genes regulated by Rv1404, rv1403c and rv1405c, encode putative SAM-dependent methyltransferases. To elucidate gene function, M. tuberculosis rv1403c and rv1405c mutants were constructed. The mutants showed attenuated growth in response to in vitro stress conditions that mimic the intracellular milieu. Our data sheds new light on the function of a novel regulon controlled by Rv1404 that coordinates adaptation of M. tuberculosis to the in vivo environment and reveals the Rv1405c and Rv1403c methyltransferases as playing a role in this adaptive process. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Archease from Pyrococcus abyssi improves substrate specificity and solubility of a tRNA m5C methyltransferase

    DEFF Research Database (Denmark)

    Auxilien, Sylvie; El Khadali, Fatima; Rasmussen, Anette

    2007-01-01

    reading frame (PAB1947), which is shown here to encode a tRNA m(5)C methyltransferase. In vitro, the purified recombinant methyltransferase catalyzes m(5)C formation at several cytosines within tRNAs with preference for C49. The specificity of the methyltransferase is increased by the archease...

  9. Coordinate regulation of DNA methyltransferase expression during oogenesis

    Directory of Open Access Journals (Sweden)

    Bestor Timothy H

    2007-04-01

    Full Text Available Abstract Background Normal mammalian development requires the action of DNA methyltransferases (DNMTs for the establishment and maintenance of DNA methylation within repeat elements and imprinted genes. Here we report the expression dynamics of Dnmt3a and Dnmt3b, as well as a regulator of DNA methylation, Dnmt3L, in isolated female germ cells. Results Our results indicate that these enzymes are coordinately regulated and that their expression peaks during the stage of postnatal oocyte development when maternal methylation imprints are established. We find that Dnmt3a, Dnmt3b, Dnmt3L and Dnmt1o transcript accumulation is related to oocyte diameter. Furthermore, DNMT3L deficient 15 dpp oocytes have aberrantly methylated Snrpn, Peg3 and Igf2r DMRs, but normal IAP and LINE-1 methylation levels, thereby highlighting a male germ cell specific role for DNMT3L in the establishment of DNA methylation at repeat elements. Finally, real-time RT-PCR analysis indicates that the depletion of either DNMT3L or DNMT1o in growing oocytes results in the increased expression of the de novo methyltransferase Dnmt3b, suggesting a potential compensation mechanism by this enzyme for the loss of one of the other DNA methyltransferases. Conclusion Together these results provide a better understanding of the developmental regulation of Dnmt3a, Dnmt3b and Dnmt3L at the time of de novo methylation during oogenesis and demonstrate that the involvement of DNMT3L in retrotransposon silencing is restricted to the male germ line. This in turn suggests the existence of other factors in the oocyte that direct DNA methylation to transposons.

  10. A SABATH Methyltransferase from the moss Physcomitrella patens catalyzes

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Nan [ORNL; Ferrer, Jean-Luc [Universite Joseph Fourier, France; Moon, Hong S [Department of Plant Sciences, University of Tennessee; Kapteyn, Jeremy [Institute of Biological Chemistry, Washington State University; Zhuang, Xiaofeng [Department of Plant Sciences, University of Tennessee; Hasebe, Mitsuyasu [Laboratory of Evolutionary Biology, National Institute for Biology, 38 Nishigounaka; Stewart, Neal C. [Department of Plant Sciences, University of Tennessee; Gang, David R. [Institute of Biological Chemistry, Washington State University; Chen, Feng [University of Tennessee, Knoxville (UTK)

    2012-01-01

    Known SABATH methyltransferases, all of which were identified from seed plants, catalyze methylation of either the carboxyl group of a variety of low molecular weight metabolites or the nitrogen moiety of precursors of caffeine. In this study, the SABATH family from the bryophyte Physcomitrella patens was identified and characterized. Four SABATH-like sequences (PpSABATH1, PpSABATH2, PpSABATH3, and PpSABATH4) were identified from the P. patens genome. Only PpSABATH1 and PpSABATH2 showed expression in the leafy gametophyte of P. patens. Full-length cDNAs of PpSABATH1 and PpSABATH2 were cloned and expressed in soluble form in Escherichia coli. Recombinant PpSABATH1 and PpSABATH2 were tested for methyltransferase activity with a total of 75 compounds. While showing no activity with carboxylic acids or nitrogen-containing compounds, PpSABATH1 displayed methyltransferase activity with a number of thiols. PpSABATH2 did not show activity with any of the compounds tested. Among the thiols analyzed, PpSABATH1 showed the highest level of activity with thiobenzoic acid with an apparent Km value of 95.5 lM, which is comparable to those of known SABATHs. Using thiobenzoic acid as substrate, GC MS analysis indicated that the methylation catalyzed by PpSABATH1 is on the sulfur atom. The mechanism for S-methylation of thiols catalyzed by PpSABATH1 was partially revealed by homology-based structural modeling. The expression of PpSABATH1 was induced by the treatment of thiobenzoic acid. Further transgenic studies showed that tobacco plants overexpressing PpSABATH1 exhibited enhanced tolerance to thiobenzoic acid, suggesting that PpSABATH1 have a role in the detoxification of xenobiotic thiols.

  11. Chromosomal replication incompatibility in Dam methyltransferase deficient Escherichia coli cells

    DEFF Research Database (Denmark)

    Freiesleben, Ulrik Von

    1996-01-01

    Dam methyltransferase deficient Escherichia coli cells containing minichromosomes were constructed. Free plasmid DNA could not be detected in these cells and the minichromosomes were found to be integrated in multiple copies in the origin of replication (oriC) region of the host chromosome...... and functional oriC sequence. The seqA2 mutation was found to overcome the incompatibility phenotype by increasing the cellular oriC copy nnumber 3-fold thereby allowing minichromosomes to coexist with the chromosome. The replication pattern of a wild type strain with multiple integrated minichromosomes...

  12. Molecular junctions based on SAMs of cruciform oligo(phenylene ethynylene)s

    DEFF Research Database (Denmark)

    Wei, Zhongming; Li, Tao; Jennum, Karsten Stein

    2012-01-01

    Cruciform oligo(phenylene ethynylene)s (OPEs) with an extended tetrathiafulvalene (TTF) donor moiety (OPE5-TTF and OPE3-TTF) and their simple analogues (OPE5-S and OPE3) without conjugated substituents were used to form high quality self-assembled monolayers (SAMs) on ultra-flat gold substrates....... Molecular junctions based on these SAMs were investigated using conducting-probe atomic force microscopy (CP-AFM). The TTF substituent changes the molecular orbital energy levels and decreases the HOMO-LUMO energy gap, resulting in a nine-fold increase in conductance for both TTF cruciform OPEs compared...... to the unsubstituted analogues. The difference in electrical transport properties of the SAMs was reproduced by the theoretical transport calculations for the single molecules....

  13. Bringing a Chemical Laboratory Named Sam to Mars on the 2011 Curiosity Rover

    Science.gov (United States)

    Mahaffy, P. R.; Bleacher, L.; Jones, A.; Conrad, P. G.; Cabane, M.; Webster, C. R.; Atreya, S. A.; Manning, H.

    2010-01-01

    An important goal of upcoming missions to Mars is to understand if life could have developed there. The task of the Sample Analysis at Mars (SAM) suite of instruments [1] and the other Curiosity investigations [2] is to move us steadily toward that goal with an assessment of the habitability of our neighboring planet through a series of chemical and geological measurements. SAM is designed to search for organic compounds and inorganic volatiles and measure isotope ratios. Other instruments on Curiosity will provide elemental analysis and identify minerals. SAM will analyze both atmospheric samples and gases evolved from powdered rocks that may have formed billions of years ago with Curiosity providing access to interesting sites scouted by orbiting cameras and spectrometers.

  14. Detection of Organics at Mars: How Wet Chemistry Onboard SAM Helps

    Science.gov (United States)

    Buch, A.; Freissinet, Caroline; Szopa, C.; Glavin, D.; Coll, P.; Cabane, M.; Eigenbrode, J.; Navarro-Gonzalez, R.; Coscia, D.; Teinturier, S.; hide

    2013-01-01

    For the first time in the history of space exploration, a mission of interest to astrobiology could be able to analyze refractory organic compounds in the soil of Mars. Wet chemistry experiment allow organic components to be altered in such a way that improves there detection either by releasing the compounds from sample matricies or by changing the chemical structure to be amenable to analytical conditions. The latter is particular important when polar compounds are present. Sample Analysis at Mars (SAM), on the Curiosity rover of the Mars Science Laboratory mission, has onboard two wet chemistry experiments: derivatization and thermochemolysis. Here we report on the nature of the MTBSTFA derivatization experiment on SAM, the detection of MTBSTFA in initial SAM results, and the implications of this detection.

  15. Data handling with SAM and art at the NOνA experiment

    Science.gov (United States)

    Aurisano, A.; Backhouse, C.; Davies, G. S.; Illingworth, R.; Mayer, N.; Mengel, M.; Norman, A.; Rocco, D.; Zirnstein, J.

    2015-12-01

    During operations, NOvA produces between 5,000 and 7,000 raw files per day with peaks in excess of 12,000. These files must be processed in several stages to produce fully calibrated and reconstructed analysis files. In addition, many simulated neutrino interactions must be produced and processed through the same stages as data. To accommodate the large volume of data and Monte Carlo, production must be possible both on the Fermilab grid and on off-site farms, such as the ones accessible through the Open Science Grid. To handle the challenge of cataloging these files and to facilitate their off-line processing, we have adopted the SAM system developed at Fermilab. SAM indexes files according to metadata, keeps track of each file's physical locations, provides dataset management facilities, and facilitates data transfer to off-site grids. To integrate SAM with Fermilab's art software framework and the NOvA production workflow, we have developed methods to embed metadata into our configuration files, art files, and standalone ROOT files. A module in the art framework propagates the embedded information from configuration files into art files, and from input art files to output art files, allowing us to maintain a complete processing history within our files. Embedding metadata in configuration files also allows configuration files indexed in SAM to be used as inputs to Monte Carlo production jobs. Further, SAM keeps track of the input files used to create each output file. Parentage information enables the construction of self-draining datasets which have become the primary production paradigm used at NOvA. In this paper we will present an overview of SAM at NOvA and how it has transformed the file production framework used by the experiment.

  16. GoSam: A program for automated one-loop Calculations

    CERN Document Server

    Cullen, G; Heinrich, G; Luisoni, G; Mastrolia, P; Ossola, G; Reiter, T; Tramontano, F

    2012-01-01

    The program package GoSam is presented which aims at the automated calculation of one-loop amplitudes for multi-particle processes. The amplitudes are generated in terms of Feynman diagrams and can be reduced using either D-dimensional integrand-level decomposition or tensor reduction, or a combination of both. GoSam can be used to calculate one-loop corrections to both QCD and electroweak theory, and model files for theories Beyond the Standard Model can be linked as well. A standard interface to programs calculating real radiation is also included. The flexibility of the program is demonstrated by various examples.

  17. GoSam. A program for automated one-loop calculations

    Energy Technology Data Exchange (ETDEWEB)

    Cullen, G. [Deutsches Elektronen-Synchrotron (DESY), Zeuthen (Germany); Greiner, N.; Heinrich, G.; Reiter, T. [Max-Planck-Institut fuer Physik, Muenchen (Germany); Luisoni, G. [Durham Univ. (United Kingdom). Inst. for Particle Physics Phenomenology; Mastrolia, P. [Max-Planck-Institut fuer Physik, Muenchen (Germany); Padua Univ. (Italy). Dipt. di Fisica; Ossola, G. [City Univ. of New York, NY (United States). New York City College of Technology; Tramontano, F. [European Organization for Nuclear Research (CERN), Geneva (Switzerland)

    2011-11-15

    The program package GoSam is presented which aims at the automated calculation of one-loop amplitudes for multi-particle processes. The amplitudes are generated in terms of Feynman diagrams and can be reduced using either D-dimensional integrand-level decomposition or tensor reduction, or a combination of both. GoSam can be used to calculate one-loop corrections to both QCD and electroweak theory, and model files for theories Beyond the Standard Model can be linked as well. A standard interface to programs calculating real radiation is also included. The flexibility of the program is demonstrated by various examples. (orig.)

  18. The nucleation and growth of calcium oxalate monohydrate on self- assembled monolayers (SAMs)

    Energy Technology Data Exchange (ETDEWEB)

    Campbell, A.A.; Tarasevich, B.J.; Graff, G.L.; Fryxell, G.E.; Rieke, P.C.

    1992-05-01

    A physical chemical approach was used to study calcium oxalate monohydrate (COM) nucleation and growth on various organic interfaces. Self-assembling monolayers (SAMs), containing derivatized organic functional groups, were designed to mimic various amino acid residues present in both urine and stone matrix macromolecules. Derivatized surfaces include SAMs with terminal methyl, bromo, imidazole, and thiazolidine-carboxylic acid functional groups. Pronounced differences in COM deposition were observed for the various interfaces with the imidazole and thiazolidine surfaces having the greatest effect and the methyl and bromo groups having little or no nucleating potential.

  19. Rhodium deposition onto a 4-mercaptopyridine SAM on Au(1 1 1)

    Energy Technology Data Exchange (ETDEWEB)

    Manolova, M. [Institute of Electrochemistry, University of Ulm, 89069 Ulm (Germany); Kayser, M. [Institute of Electrochemistry, University of Ulm, 89069 Ulm (Germany); Kolb, D.M. [Institute of Electrochemistry, University of Ulm, 89069 Ulm (Germany)]. E-mail: dieter.kolb@uni-ulm.de; Boyen, H.-G. [Institute of Solid State Physics, University of Ulm, 89069 Ulm (Germany); Ziemann, P. [Institute of Solid State Physics, University of Ulm, 89069 Ulm (Germany); Mayer, D. [BASF Electronic Materials GmbH, 67056 Ludwigshafen (Germany); Wirth, A. [BASF Electronic Materials GmbH, 67056 Ludwigshafen (Germany)

    2007-02-10

    The application of a recently developed method for the deposition of Pd and Pt on top of a SAM, has been successfully extended to Rh, thus proving the versatility of the new concept. Experimental evidence from cyclic voltammetry, in situ STM and ex situ X-ray photoemission spectroscopy is presented for the deposition of monoatomic high rhodium islands onto a 4-mercaptopyridine self-assembled monolayer on a Au(1 1 1) electrode. By repetitive complexation of the Rh ions to the ring-nitrogen and reduction in a Rh-ion free solution, an almost completely covered SAM is obtained. The consequences of making contacts for molecular electronics are briefly discussed.

  20. Damage Simulation Analysis of Discrete Rods to SAM based on FEA

    Directory of Open Access Journals (Sweden)

    Wang Chengcheng

    2015-01-01

    Full Text Available At present ship to air missile is the main force of anti-missile. The warhead damage element’ impact and penetration effect are the main factors for damaging supersonic anti-ship missile (SAM. In this paper, the damage mechanism of the anti-ship missile is studied. Dynamic initial velocity of damage elements is modeled and simulated. The four important parts of SAM with typical material are modeled and simulated based on finite element analysis (FEA. It’s useful to the study of the further model of vulnerability analysis of whole anti-ship missiles.

  1. Changes in SAM2 expression affect lactic acid tolerance and lactic acid production in Saccharomyces cerevisiae.

    Science.gov (United States)

    Dato, Laura; Berterame, Nadia Maria; Ricci, Maria Antonietta; Paganoni, Paola; Palmieri, Luigi; Porro, Danilo; Branduardi, Paola

    2014-10-30

    The great interest in the production of highly pure lactic acid enantiomers comes from the application of polylactic acid (PLA) for the production of biodegradable plastics. Yeasts can be considered as alternative cell factories to lactic acid bacteria for lactic acid production, despite not being natural producers, since they can better tolerate acidic environments. We have previously described metabolically engineered Saccharomyces cerevisiae strains producing high amounts of L-lactic acid (>60 g/L) at low pH. The high product concentration represents the major limiting step of the process, mainly because of its toxic effects. Therefore, our goal was the identification of novel targets for strain improvement possibly involved in the yeast response to lactic acid stress. The enzyme S-adenosylmethionine (SAM) synthetase catalyses the only known reaction leading to the biosynthesis of SAM, an important cellular cofactor. SAM is involved in phospholipid biosynthesis and hence in membrane remodelling during acid stress. Since only the enzyme isoform 2 seems to be responsive to membrane related signals (e.g. myo-inositol), Sam2p was tagged with GFP to analyse its abundance and cellular localization under different stress conditions. Western blot analyses showed that lactic acid exposure correlates with an increase in protein levels. The SAM2 gene was then overexpressed and deleted in laboratory strains. Remarkably, in the BY4741 strain its deletion conferred higher resistance to lactic acid, while its overexpression was detrimental. Therefore, SAM2 was deleted in a strain previously engineered and evolved for industrial lactic acid production and tolerance, resulting in higher production. Here we demonstrated that the modulation of SAM2 can have different outcomes, from clear effects to no significant phenotypic responses, upon lactic acid stress in different genetic backgrounds, and that at least in one genetic background SAM2 deletion led to an industrially relevant

  2. A new application of SAM in the non-destructive inspection for SIM card

    Science.gov (United States)

    Cai, Bin; He, Yandong; Zhao, Yang; Chen, Yanning; Zhang, Haifeng; Zhao, Dongyan

    2017-07-01

    Scanning Acoustic Microscopy (SAM) is a typical inspection method in the semiconductor IC manufacturing industry. Because the die thickness is a key parameter for SIM card, a new method to measure the internal die thickness of SIM card is proposed with SAM’s reflective scanning mode. Using this method the internal die thickness of SIM card can be accurately measured without introducing any damages to SIM card. The thickness model and methodology based on the SAM signals have been established. The model was properly verified and calibrated by two real test cases.

  3. Reaction mechanism of isoflavone O-methyltransferase: A theoretical investigation

    Science.gov (United States)

    Cui, Feng-Chao; Pan, Xiao-Liang; Liu, Jing-Yao

    2011-01-01

    The methyl-transfer reaction mechanism catalyzed by Isoflavone O-methyl-transferase (IOMT) and the roles of several residues around the active site are investigated by employing density functional method. The calculations confirm that the proton transfer from daidzein to His257 occurs barrierlessly, and the methyl group is transferred from S-adenosyl-L-methionine (SAM) to phenolate ion in a single SN 2 step with a barrier of 17.0 kcal/mol, consistent with experimental value. Glu318 and Asp288 play important roles in lowering the reaction barrier.

  4. Mutagenesis of S-Adenosyl-l-Methionine-Binding Residues in Coronavirus nsp14 N7-Methyltransferase Demonstrates Differing Requirements for Genome Translation and Resistance to Innate Immunity.

    Science.gov (United States)

    Case, James Brett; Ashbrook, Alison W; Dermody, Terence S; Denison, Mark R

    2016-08-15

    Eukaryotic mRNAs possess a methylated 5'-guanosine cap that is required for RNA stability, efficient translation, and protection from cell-intrinsic defenses. Many viruses use 5' caps or other mechanisms to mimic a cap structure to limit detection of viral RNAs by intracellular innate sensors and to direct efficient translation of viral proteins. The coronavirus (CoV) nonstructural protein 14 (nsp14) is a multifunctional protein with N7-methyltransferase (N7-MTase) activity. The highly conserved S-adenosyl-l-methionine (SAM)-binding residues of the DxG motif are required for nsp14 N7-MTase activity in vitro However, the requirement for CoV N7-MTase activity and the importance of the SAM-binding residues during viral replication have not been determined. Here, we engineered mutations in murine hepatitis virus (MHV) nsp14 N7-MTase at residues D330 and G332 and determined the effects of these mutations on viral replication, sensitivity to mutagen, inhibition by type I interferon (IFN), and translation efficiency. Virus encoding a G332A substitution in nsp14 displayed delayed replication kinetics and decreased peak titers relative to wild-type (WT) MHV. In addition, replication of nsp14 G332A virus was diminished following treatment of cells with IFN-β, and nsp14 G332A genomes were translated less efficiently both in vitro and during viral infection. In contrast, substitution of alanine at MHV nsp14 D330 did not affect viral replication, sensitivity to mutagen, or inhibition by IFN-β compared to WT MHV. Our results demonstrate that the conserved MHV N7-MTase SAM-binding-site residues are not required for MHV viability and suggest that the determinants of CoV N7-MTase activity differ in vitro and during virus infection. Human coronaviruses, most notably severe acute respiratory syndrome (SARS)-CoV and Middle East respiratory syndrome (MERS)-CoV, cause severe and lethal human disease. Since specific antiviral therapies are not available for the treatment of human

  5. Structural biology of human H3K9 methyltransferases.

    Directory of Open Access Journals (Sweden)

    Hong Wu

    2010-01-01

    Full Text Available SET domain methyltransferases deposit methyl marks on specific histone tail lysine residues and play a major role in epigenetic regulation of gene transcription. We solved the structures of the catalytic domains of GLP, G9a, Suv39H2 and PRDM2, four of the eight known human H3K9 methyltransferases in their apo conformation or in complex with the methyl donating cofactor, and peptide substrates. We analyzed the structural determinants for methylation state specificity, and designed a G9a mutant able to tri-methylate H3K9. We show that the I-SET domain acts as a rigid docking platform, while induced-fit of the Post-SET domain is necessary to achieve a catalytically competent conformation. We also propose a model where long-range electrostatics bring enzyme and histone substrate together, while the presence of an arginine upstream of the target lysine is critical for binding and specificity.This article can also be viewed as an enhanced version in which the text of the article is integrated with interactive 3D representations and animated transitions. Please note that a web plugin is required to access this enhanced functionality. Instructions for the installation and use of the web plugin are available in Text S1.

  6. Crystal structure of arginine methyltransferase 6 from Trypanosoma brucei.

    Directory of Open Access Journals (Sweden)

    Chongyuan Wang

    Full Text Available Arginine methylation plays vital roles in the cellular functions of the protozoan Trypanosoma brucei. The T. brucei arginine methyltransferase 6 (TbPRMT6 is a type I arginine methyltransferase homologous to human PRMT6. In this study, we report the crystal structures of apo-TbPRMT6 and its complex with the reaction product S-adenosyl-homocysteine (SAH. The structure of apo-TbPRMT6 displays several features that are different from those of type I PRMTs that were structurally characterized previously, including four stretches of insertion, the absence of strand β15, and a distinct dimerization arm. The comparison of the apo-TbPRMT6 and SAH-TbPRMT6 structures revealed the fine rearrangements in the active site upon SAH binding. The isothermal titration calorimetry results demonstrated that SAH binding greatly increases the affinity of TbPRMT6 to a substrate peptide derived from bovine histone H4. The western blotting and mass spectrometry results revealed that TbPRMT6 methylates bovine histone H4 tail at arginine 3 but cannot methylate several T. brucei histone tails. In summary, our results highlight the structural differences between TbPRMT6 and other type I PRMTs and reveal that the active site rearrangement upon SAH binding is important for the substrate binding of TbPRMT6.

  7. Hypnotizability and Catechol-O-Methyltransferase (COMT polymorphysms in Italians

    Directory of Open Access Journals (Sweden)

    Silvano ePresciuttini

    2014-01-01

    Full Text Available Higher brain dopamine content depending on lower activity of Catechol-O-Methyltransferase (COMT in subjects with high hypnotisability scores (highs has been considered responsible for their attentional characteristics. However, the results of the previous genetic studies on association between hypnotisability and the Catechol-O-Methyltransferase (COMT single nucleotide polymorphism (SNP rs4680 (Val158Met were inconsistent. Here, we used a selective genotyping approach to re-evaluate the association between hypnotisability and COMT in the context of a two-SNP haplotype analysis, considering not only the Val158Met polymorphism, but also the closely located rs4818 SNP. An Italian sample of 53 highs, 49 low hypnotizable subjects (lows and 57 controls, were genotyped for a segment of 805 bp of the COMT gene, including Val158Met and the closely located rs4818 SNP. Our selective genotyping approach had 97.1% power to detect the previously reported strongest association at the significance level of 5%. We found no evidence of association at the SNP, haplotype and diplotype levels. Thus, our results challenge the dopamine-based theory of hypnosis and indirectly support recent neuropsychological and neurophysiological findings reporting the lack of any association between hypnotisability and focused attention abilities.

  8. Roles of the EZH2 histone methyltransferase in cancer epigenetics.

    Science.gov (United States)

    Simon, Jeffrey A; Lange, Carol A

    2008-12-01

    EZH2 is the catalytic subunit of Polycomb repressive complex 2 (PRC2), which is a highly conserved histone methyltransferase that targets lysine-27 of histone H3. This methylated H3-K27 chromatin mark is commonly associated with silencing of differentiation genes in organisms ranging from plants to flies to humans. Studies on human tumors show that EZH2 is frequently over-expressed in a wide variety of cancerous tissue types, including prostate and breast. Although the mechanistic contributions of EZH2 to cancer progression are not yet determined, functional links between EZH2-mediated histone methylation and DNA methylation suggest partnership with the gene silencing machinery implicated in tumor suppressor loss. Here we review the basic molecular biology of EZH2 and the findings that implicate EZH2 in different cancers. We also discuss EZH2 connections to other silencing enzymes, such as DNA methyltransferases and histone deacetylases, and we consider progress on deciphering mechanistic consequences of EZH2 overabundance and its potential roles in tumorigenesis. Finally, we review recent findings that link EZH2 roles in stem cells and cancer, and we consider prospects for integrating EZH2 blockade into strategies for developing epigenetic therapies.

  9. Analysis of the interactions between host factor Sam68 and viral elements during foot-and-mouth disease virus infection

    Science.gov (United States)

    The nuclear protein Src-associated protein of 68 kDa in mitosis (Sam68) is known to bind RNA and be involved in cellular processes triggered in response to environmental stresses, including virus infection. Interestingly, Sam68, is a multi-functional protein implicated in the life cycle of retroviru...

  10. "SAMs meet MEMS": surface modification with self-assembled monolayers for the dry-demolding of photoplastic MEMS/NEMS

    NARCIS (Netherlands)

    Kim, B.J.; Kim, G.M.; Liebau, M.; Huskens, Jurriaan; Reinhoudt, David; Brugger, J.P.

    2001-01-01

    In this contribution we demonstrate the use of self-assembled monolayers (SAMs) as anti-adhesion coating to assist the removal of photoplastic MEMS/NEMS with a patterned metal layer from the surface without wet chemical sacrificial layer etching, so-called 'dry-demolding'. The SAMs functionality

  11. cljam: a library for handling DNA sequence alignment/map (SAM) with parallel processing.

    Science.gov (United States)

    Takeuchi, Toshiki; Yamada, Atsuo; Aoki, Takashi; Nishimura, Kunihiro

    2016-01-01

    Next-generation sequencing can determine DNA bases and the results of sequence alignments are generally stored in files in the Sequence Alignment/Map (SAM) format and the compressed binary version (BAM) of it. SAMtools is a typical tool for dealing with files in the SAM/BAM format. SAMtools has various functions, including detection of variants, visualization of alignments, indexing, extraction of parts of the data and loci, and conversion of file formats. It is written in C and can execute fast. However, SAMtools requires an additional implementation to be used in parallel with, for example, OpenMP (Open Multi-Processing) libraries. For the accumulation of next-generation sequencing data, a simple parallelization program, which can support cloud and PC cluster environments, is required. We have developed cljam using the Clojure programming language, which simplifies parallel programming, to handle SAM/BAM data. Cljam can run in a Java runtime environment (e.g., Windows, Linux, Mac OS X) with Clojure. Cljam can process and analyze SAM/BAM files in parallel and at high speed. The execution time with cljam is almost the same as with SAMtools. The cljam code is written in Clojure and has fewer lines than other similar tools.

  12. Initial SAM Calibration Gas Experiments on Mars: Quadrupole Mass Spectrometer Results and Implications

    Science.gov (United States)

    Franz, Heather B.; Trainer, Melissa G.; Malespin, Charles A.; Mahaffy, Paul R.; Atreya, Sushil K.; Becker, Richard H,; Benna, Mehdi; Conrad, Pamela G.; Eigenbrode, Jennifer L.; Freissinet, Caroline; hide

    2017-01-01

    The Sample Analysis at Mars (SAM) instrument suite of the Mars Science Laboratory (MSL) Curiosity rover is equipped to analyze both martian atmospheric gases and volatiles released by pyrolysis of solid surface materials, with target measurements including chemical and isotopic composition (Mahaffy et al., 2012). To facilitate assessment of instrument performance and validation of results obtained on Mars, SAM houses a calibration cell containing CO2, Ar, N2, Xe, and several fluorinated hydrocarbon compounds (Franz et al., 2014; Mahaffy et al., 2012). This report describes the first two experiments utilizing this calibration cell on Mars and gives results from analysis of data acquired with the SAM Quadrupole Mass Spectrometer (QMS). These data support the accuracy of isotope ratios obtained with the QMS (Conrad et al., 2016; Mahaffy et al., 2013) and provide ground-truth for reassessment of analytical constants required for atmospheric measurements, which were reported in previous contributions (Franz et al., 2015, 2014). The most significant implication of the QMS data involves reinterpretation of pre-launch contamination previously believed to affect only CO abundance measurements (Franz et al., 2015) to affect N2 abundances, as well. The corresponding adjustment to the N2 calibration constant presented here brings the atmospheric volume mixing ratios for Ar and N2 retrieved by SAM into closer agreement with those reported by the Viking mission (Owen et al., 1977; Oyama and Berdahl, 1977).

  13. Initial SAM calibration gas experiments on Mars: Quadrupole mass spectrometer results and implications

    Science.gov (United States)

    Franz, Heather B.; Trainer, Melissa G.; Malespin, Charles A.; Mahaffy, Paul R.; Atreya, Sushil K.; Becker, Richard H.; Benna, Mehdi; Conrad, Pamela G.; Eigenbrode, Jennifer L.; Freissinet, Caroline; Manning, Heidi L. K.; Prats, Benito D.; Raaen, Eric; Wong, Michael H.

    2017-04-01

    The Sample Analysis at Mars (SAM) instrument suite of the Mars Science Laboratory (MSL) Curiosity rover is equipped to analyze both martian atmospheric gases and volatiles released by pyrolysis of solid surface materials, with target measurements including chemical and isotopic composition (Mahaffy et al., 2012). To facilitate assessment of instrument performance and validation of results obtained on Mars, SAM houses a calibration cell containing CO2, Ar, N2, Xe, and several fluorinated hydrocarbon compounds (Franz et al., 2014; Mahaffy et al., 2012). This report describes the first two experiments utilizing this calibration cell on Mars and gives results from analysis of data acquired with the SAM Quadrupole Mass Spectrometer (QMS). These data support the accuracy of isotope ratios obtained with the QMS (Conrad et al., 2016; Mahaffy et al., 2013) and provide ground-truth for reassessment of analytical constants required for atmospheric measurements, which were reported in previous contributions (Franz et al., 2015, 2014). The most significant implication of the QMS data involves reinterpretation of pre-launch contamination previously believed to affect only CO abundance measurements (Franz et al., 2015) to affect N2 abundances, as well. The corresponding adjustment to the N2 calibration constant presented here brings the atmospheric volume mixing ratios for Ar and N2 retrieved by SAM into closer agreement with those reported by the Viking mission (Owen et al., 1977; Oyama and Berdahl, 1977).

  14. How Safe is Sam Lloyd's Bet? The 15-Puzzle and Beyond

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 5; Issue 11. How Safe is Sam Lloyd's Bet? The 15-Puzzle and Beyond. Jyoti Ramakrishnan. Classroom Volume 5 Issue 11 November 2000 pp 80-89. Fulltext. Click here to view fulltext PDF. Permanent link:

  15. Case Studies Comparing System Advisor Model (SAM) Results to Real Performance Data: Preprint

    Energy Technology Data Exchange (ETDEWEB)

    Blair, N.; Dobos, A.; Sather, N.

    2012-06-01

    NREL has completed a series of detailed case studies comparing the simulations of the System Advisor Model (SAM) and measured performance data or published performance expectations. These case studies compare PV measured performance data with simulated performance data using appropriate weather data. The measured data sets were primarily taken from NREL onsite PV systems and weather monitoring stations.

  16. Constraints on the Mineralogy of Gale Crater Mudstones from MSL SAM Evolved Water

    Science.gov (United States)

    McAdam, A. C.; Sutter, B.; Franz, H. B.; Hogancamp, J. V. (Clark); Knudson, C. A.; Andrejkovicova, S.; Archer, P. D.; Eigenbrode, J. L.; Ming, D. W.; Mahaffy, P. R.

    2017-01-01

    The Sample Analysis at Mars (SAM) and Chemistry and Mineralogy (CheMin) instruments on the Mars Science Laboratory (MSL) have analysed more than 150 micron fines from 14 sites at Gale Crater. Here we focus on the mudstone samples. Two were drilled from sites John Klein (JK) and Cumberland (CB) in the Sheepbed mudstone. Six were drilled from Murray Formation mudstone: Confidence Hills (CH), Mojave (MJ), Telegraph Peak (TP), Buckskin (BK), Oudam (OU), Marimba (MB). SAM's evolved gas analysis mass spectrometry (EGA-MS) detected H2O, CO2, O2, H2, SO2, H2S, HCl, NO, and other trace gases, including organic fragments. The identity and evolution temperature of evolved gases can support CheMin mineral detection and place constraints on trace volatile-bearing phases or phases difficult to characterize with X-ray diffraction (e.g., amorphous phases). Here we will focus on SAM H2O data and comparisons to SAM-like analyses of key reference materials.

  17. Stealing the gold a celebration of the pioneering physics of Sam Edwards

    CERN Document Server

    Goldenfeld, Nigel; Sherrington, D C; Edwards, S F

    2004-01-01

    This title presents a survey of some of the most exciting topics in condensed matter physics today, from the perspective of the pioneering work of Sam Edwards. Original articles from leaders in the field highlight the historical development as well as new and emerging areas.

  18. Cross-Reactive Sensor Array for Metal Ion Sensing Based on Fluorescent SAMs

    Directory of Open Access Journals (Sweden)

    Mercedes Crego-Calama

    2007-09-01

    Full Text Available Fluorescent self assembled monolayers (SAMs on glass were previouslydeveloped in our group as new sensing materials for metal ions. These fluorescent SAMs arecomprised by fluorophores and small molecules sequentially deposited on a monolayer onglass. The preorganization provided by the surface avoids the need for complex receptordesign, allowing for a combinatorial approach to sensing systems based on small molecules.Now we show the fabrication of an effective microarray for the screening of metal ions andthe properties of the sensing SAMs. A collection of fluorescent sensing SAMs wasgenerated by combinatorial methods and immobilized on the glass surfaces of a custom-made 140 well microtiter-plate. The resulting libraries are easily measured and show variedresponses to a series cations such as Cu2+ , Co2+ , Pb2+ , Ca2+ and Zn2+ . These surfaces are notdesigned to complex selectively a unique analyte but rather they are intended to producefingerprint type responses to a range of analytes by less specific interactions. The unselectiveresponses of the library to the presence of different cations generate a characteristic patternfor each analyte, a “finger print” response.

  19. A Normalizing Approach to Cognitive Therapy for Intrusive Obsessional and Psychotic Phenomena: The Case of Sam

    Science.gov (United States)

    Morrison, Anthony P.

    2005-01-01

    The case of Sam is conceptualized using a normalizing cognitive approach, which assumes that the cultural acceptability of his appraisals distinguishes them as psychotic. The treatment approach that is based upon such a case formulation involves the evaluation of such distressing appraisals and the generation of alternative explanations.…

  20. Making Time for Instructional Leadership. Volume 1: The Evolution of the SAM Process

    Science.gov (United States)

    Goldring, Ellen; Grissom, Jason A.; Neumerski, Christine M.; Murphy, Joseph; Blissett, Richard; Porter, Andy

    2015-01-01

    This three-volume report describes the "SAM (School Administration Manager) process," an approach that about 700 schools around the nation are using to direct more of principals' time and effort to improve teaching and learning in classrooms. Research has shown that a principal's instructional leadership is second only to teaching among…

  1. Molten Salt Power Tower Cost Model for the System Advisor Model (SAM)

    Energy Technology Data Exchange (ETDEWEB)

    Turchi, C. S.; Heath, G. A.

    2013-02-01

    This report describes a component-based cost model developed for molten-salt power tower solar power plants. The cost model was developed by the National Renewable Energy Laboratory (NREL), using data from several prior studies, including a contracted analysis from WorleyParsons Group, which is included herein as an Appendix. The WorleyParsons' analysis also estimated material composition and mass for the plant to facilitate a life cycle analysis of the molten salt power tower technology. Details of the life cycle assessment have been published elsewhere. The cost model provides a reference plant that interfaces with NREL's System Advisor Model or SAM. The reference plant assumes a nominal 100-MWe (net) power tower running with a nitrate salt heat transfer fluid (HTF). Thermal energy storage is provided by direct storage of the HTF in a two-tank system. The design assumes dry-cooling. The model includes a spreadsheet that interfaces with SAM via the Excel Exchange option in SAM. The spreadsheet allows users to estimate the costs of different-size plants and to take into account changes in commodity prices. This report and the accompanying Excel spreadsheet can be downloaded at https://sam.nrel.gov/cost.

  2. Evidence for Smectite Clays from MSL SAM Analyses of Mudstone at Yellowknife Bay, Gale Crater, Mars

    Science.gov (United States)

    McAdam, Amy; Franz, Heather; Mahaffy, Paul R.; Eigenbrode, Jennifer L.; Stern, Jennifer C.; Brunner, Anna; Archer, Paul Douglas; Ming, Douglas W.; Morris, Richard V.; Atreya, Sushil K.

    2013-01-01

    Drilled samples of mudstone from the Sheepbed unit at Yellowknife Bay were analyzed by MSL instruments including the Sample Analysis at Mars (SAM) and Chemistry and Mineralogy (CheMin) instruments in MSL's Analytical Laboratory. CheMin analyses revealed the first in situ X-ray diffraction based evidence of clay minerals on Mars, which are likely trioctahedral smectites (e.g., saponite) and comprise approx 20% of the mudstone sample (e.g., Bristow et al., this meeting). SAM analyses, which heated the mudstone samples to 1000 C and monitored volatiles evolved to perform in situ evolved gas analysis mass spectrometry (EGA-MS), resulted in a H2O trace exhibiting a wide evolution at temperatures clay mineral. Comparison to EGA-MS data collected under SAM-like conditions on a variety of clay mineral reference materials indicate that a trioctahedral smectite, such as saponite, is most consistent with the high temperature H2O evolution observed. There may also be SAM EGA-MS evidence for a small high temperature H2O evolution from scoop samples from the Yellowknife Bay Rocknest sand shadow bedform. As in the mudstone samples, this evolution may indicate the detection of smectite clays, and the idea that minor clays may be present in Rocknest materials that could be expected to be at least partially derived from local sources is reasonable. But, because smectite clays were not definitively observed in CheMin analyses of Rocknest materials, they must be present at much lower abundances than the approx 20% observed in the mudstone samples. This potential detection underscores the complementary nature of the MSL CheMin and SAM instruments for investigations of martian sample mineralogy. Information on the nature of Yellowknife Bay clay minerals may also be available from the detection of H2 evolved during SAM EGA-MS at high temperature. A likely source of at least some of this H2 is H2O evolved from the smectite clays at high temperature, and it is possible these evolutions can

  3. Mechanism elucidation of the radical SAM enzyme spore photoproduct lyase (SPL)

    Science.gov (United States)

    Li, Lei

    2011-01-01

    Spore photoproduct lyase (SPL) repairs a special thymine dimer 5-thyminyl-5,6-dihydrothymine, which is commonly called spore photoproduct or SP at the bacterial early germination phase. SP is the exclusive DNA photo-damage product in bacterial endospores; its generation and swift repair by SPL are responsible for the spores’ extremely high UV resistance. The early in vivo studies suggested that SPL utilizes a direct reversal strategy to repair the SP in the absence of light. The research in the past decade further established SPL as a radical SAM enzyme, which utilizes a tri-cysteine CXXXCXXC motif to harbor a [4Fe-4S] cluster. At the 1+ oxidation state, the cluster provides an electron to the S-adenosylmethionine (SAM), which binds to the cluster in a bidentate manner as the fourth and fifth ligands, to reductively cleave the C-S bond associated with the sulfonium ion in SAM, generating a reactive 5′-deoxyadenosyl (5′-dA) radical. This 5′-dA radical abstracts the proR hydrogen atom from the C6 carbon of SP to initiate the repair process; the resulting SP radical subsequently fragments to generate a putative thymine methyl radical, which accepts a back-donated H atom to yield the repaired TpT. SAM is suggested to be regenerated at the end of each catalytic cycle; and only a catalytic amount of SAM is needed in the SPL reaction. The H atom source for the back donation step is suggested to be a cysteine residue (C141 in B. subtilis SPL), and the H-atom transfer reaction leaves a thiyl radical behind on the protein. This thiyl radical thus must participate in the SAM regeneration process; however how the thiyl radical abstracts an H atom from the 5′-dA to regenerate SAM is unknown. This paper reviews and discusses the history and the latest progress in the mechanistic elucidation of SPL. Despite some recent breakthroughs, more questions are raised in the mechanistic understanding of this intriguing DNA repair enzyme. PMID:22197590

  4. Experimental and computational investigation of graphene/SAMs/n-Si Schottky diodes

    Science.gov (United States)

    Aydin, H.; Bacaksiz, C.; Yagmurcukardes, N.; Karakaya, C.; Mermer, O.; Can, M.; Senger, R. T.; Sahin, H.; Selamet, Y.

    2018-01-01

    We have investigated the effect of two different self-assembled monolayers (SAMs) on electrical characteristics of bilayer graphene (BLG)/n-Si Schottky diodes. Novel 4″bis(diphenylamino)-1, 1‧:3″-terphenyl-5‧ carboxylic acids (TPA) and 4,4-di-9H-carbazol-9-yl-1,1‧:3‧1‧-terphenyl-5‧ carboxylic acid (CAR) aromatic SAMs have been used to modify n-Si surfaces. Cyclic voltammetry (CV) and Kelvin probe force microscopy (KPFM) results have been evaluated to verify the modification of n-Si surface. The current-voltage (I-V) characteristics of bare and SAMs modified devices show rectification behaviour verifying a Schottky junction at the interface. The ideality factors (n) from ln(I)-V dependences were determined as 2.13, 1.96 and 2.07 for BLG/n-Si, BLG/TPA/n-Si and BLG/CAR/n-Si Schottky diodes, respectively. In addition, Schottky barrier height (SBH) and series resistance (Rs) of SAMs modified diodes were decreased compared to bare diode due to the formation of a compatible interface between graphene and Si as well as π-π interaction between aromatic SAMs and graphene. The CAR-based device exhibits better diode characteristic compared to the TPA-based device. Computational simulations show that the BLG/CAR system exhibits smaller energy-level-differences than the BLG/TPA, which supports the experimental findings of a lower Schottky barrier and series resistance in BLG/CAR diode.

  5. Molecular cloning, characterization and expression analysis of the SAMS gene during adventitious root development in IBA-induced tetraploid black locust.

    Directory of Open Access Journals (Sweden)

    Jine Quan

    Full Text Available S-Adenosylmethionine synthetase (SAMS catalyzes the synthesis of S-adenosylmethionine (SAM, a precursor for ethylene and polyamine biosynthesis. Here, we report the isolation of the 1498 bp full-length cDNA sequence encoding tetraploid black locust (Robinia pseudoacacia L. SAMS (TrbSAMS, which contains an open reading frame of 1179 bp encoding 392 amino acids. The amino acid sequence of TrbSAMS has more than 94% sequence identity to SAMSs from other plants, with a closer phylogenetic relationship to SAMSs from legumes than to SAMS from other plants. The TrbSAMS monomer consists of N-terminal, central, and C-terminal domains. Subcellular localization analysis revealed that the TrbSAMS protein localizes mainly to in the cell membrane and cytoplasm of onion epidermal cells and Arabidopsis mesophyll cell protoplasts. Indole-3-butyric acid (IBA-treated cuttings showed higher levels of TrbSAMS transcript than untreated control cuttings during root primordium and adventitious root formation. TrbSAMS and its downstream genes showed differential expression in shoots, leaves, bark, and roots, with the highest expression observed in bark. IBA-treated cuttings also showed higher SAMS activity than control cuttings during root primordium and adventitious root formation. These results indicate that TrbSAMS might play an important role in the regulation of IBA-induced adventitious root development in tetraploid black locust cuttings.

  6. Development of fluorescent methods for DNA methyltransferase assay

    Science.gov (United States)

    Li, Yueying; Zou, Xiaoran; Ma, Fei; Tang, Bo; Zhang, Chun-yang

    2017-03-01

    DNA methylation modified by DNA methyltransferase (MTase) plays an important role in regulating gene transcription, cell growth and proliferation. The aberrant DNA MTase activity may lead to a variety of human diseases including cancers. Therefore, accurate and sensitive detection of DNA MTase activity is crucial to biomedical research, clinical diagnostics and therapy. However, conventional DNA MTase assays often suffer from labor-intensive operations and time-consuming procedures. Alternatively, fluorescent methods have significant advantages of simplicity and high sensitivity, and have been widely applied for DNA MTase assay. In this review, we summarize the recent advances in the development of fluorescent methods for DNA MTase assay. These emerging methods include amplification-free and the amplification-assisted assays. Moreover, we discuss the challenges and future directions of this area.

  7. The role of histone methyltransferase EZH2 in myelodysplastic syndromes.

    Science.gov (United States)

    Xu, Feng; Li, Xiao

    2012-04-01

    Previous epigenetics research in myelodysplastic syndromes (MDS) mainly focused on the DNA methylation of tumor suppressor genes. Recent studies reported that around 6% of MDS patients have several EZH2 mutations including missense, frameshift and truncated mutations. Histone methyltransferase EZH2 plays a critical role in epigenetic regulation as a bridge between histone methylation/deacetylation and DNA methylation. EZH2 is frequently overexpressed and considered to be an oncogene in cancers; nevertheless, EZH2 is considered as a candidate tumor suppressor gene in MDS due to EZH2 mutations associated with poor survival. Many questions still need further discussion. Moreover, 3-deazaneplanocin can reduce EZH2 levels and H3K27 trimethylation, and synergistic effects are seen in combination with DNA demethylation agents or histone deacetylation inhibitors. All of the above give us more chances to improve epigenetic therapy in MDS. Therefore, the molecular mechanisms of EZH2 in tumorigenesis and the role of EZH2 in MDS are studied.

  8. Detection of a cfr(B) Variant in German Enterococcus faecium Clinical Isolates and the Impact on Linezolid Resistance in Enterococcus spp.

    Science.gov (United States)

    Bender, Jennifer K; Fleige, Carola; Klare, Ingo; Fiedler, Stefan; Mischnik, Alexander; Mutters, Nico T; Dingle, Kate E; Werner, Guido

    2016-01-01

    The National Reference Centre for Staphylococci and Enterococci in Germany has received an increasing number of clinical linezolid-resistant E. faecium isolates in recent years. Five isolates harbored a cfr(B) variant gene locus the product of which is capable of conferring linezolid resistance. The cfr(B)-like methyltransferase gene was also detected in Clostridium difficile. Antimicrobial susceptibility was determined for cfr(B)-positive and linezolid-resistant E. faecium isolates and two isogenic C. difficile strains. All strains were subjected to whole genome sequencing and analyzed with respect to mutations in the 23S rDNA, rplC, rplD and rplV genes and integration sites of the cfr(B) variant locus. To evaluate methyltransferase function, the cfr(B) variant of Enterococcus and Clostridium was expressed in both E. coli and Enterococcus spp. Ribosomal target site mutations were detected in E. faecium strains but absent in clostridia. Sequencing revealed 99.9% identity between cfr(B) of Enterococcus and cfr of Clostridium. The methyltransferase gene is encoded by transposon Tn6218 which was present in C. difficile Ox3196, truncated in some E. faecium and absent in C. difficile Ox3206. The latter finding explains the lack of linezolid and chloramphenicol resistance in C. difficile Ox3206 and demonstrates for the first time a direct correlation of elevated linezolid MICs in C. difficile upon cfr acquisition. Tn6218 insertion sites revealed novel target loci for integration, both within the bacterial chromosome and as an integral part of plasmids. Importantly, the very first plasmid-association of a cfr(B) variant was observed. Although we failed to measure cfr(B)-mediated resistance in transformed laboratory strains the occurrence of the multidrug resistance gene cfr on putatively highly mobile and/or extrachromosomal DNA in clinical isolates is worrisome with respect to dissemination of antibiotic resistances.

  9. Stability of phosphonic self assembled monolayers (SAMs) on cobalt chromium (Co-Cr) alloy under oxidative conditions

    Science.gov (United States)

    Bhure, Rahul; Abdel-Fattah, Tarek M.; Bonner, Carl; Hall, Felicia; Mahapatro, Anil

    2011-04-01

    Cobalt chromium (Co-Cr) alloys have been widely used in the biomedical arena for cardiovascular, orthopedic and dental applications. Surface modification of the alloy allows us to tailor the interfacial properties to address critical challenges of Co-Cr alloy in medical applications. Self assembled monolayers (SAMs) of Octadecylphosphonic acid (ODPA) have been used to form thin films on the oxide layer of the Co-Cr alloy surface by solution deposition technique. The SAMs formed were investigated for their stability to oxidative conditions of ambient laboratory environment over periods of 1, 3, 7 and 14 days. The samples were then characterized for their stability using X-ray Photoelectron Spectroscopy (XPS), Atomic Force Microscopy (AFM) and contact angle measurements. Detailed high energy XPS elemental scans confirmed the presence of the phosphonic monolayer after oxidative exposure which suggested that the SAMs were firmly attached to the oxide layer of Co-Cr alloy. AFM images gave topographical data of the surface and showed islands of SAMs on Co-Cr alloy surface, before and after SAM formation and also over the duration of the oxidative exposure. Contact angle measurements confirmed the hydrophobicity of the surface over 14 days. Thus the SAMs were found to be stable for the duration of the study. These SAMs could be subsequently tailored by modifying the terminal functional groups and could be used for various potential biomedical applications such as drug delivery, biocompatibility and tissue integration.

  10. Methyltransferase G9A regulates T cell differentiation during murine intestinal inflammation

    National Research Council Canada - National Science Library

    Antignano, Frann; Burrows, Kyle; Hughes, Michael R; Han, Jonathan M; Kron, Ken J; Penrod, Nadia M; Oudhoff, Menno J; Wang, Steven Kai Hao; Min, Paul H; Gold, Matthew J; Chenery, Alistair L; Braam, Mitchell J S; Fung, Thomas C; Rossi, Fabio M V; McNagny, Kelly M; Arrowsmith, Cheryl H; Lupien, Mathieu; Levings, Megan K; Zaph, Colby

    2014-01-01

    .... Using a murine T cell transfer model of colitis, we found that T cell-intrinsic expression of the histone lysine methyltransferase G9A was required for development of pathogenic T cells and intestinal inflammation...

  11. Molecular Basis for the Regulation of the H3K4 Methyltransferase Activity of PRDM9

    Directory of Open Access Journals (Sweden)

    Hong Wu

    2013-10-01

    Full Text Available PRDM9, a histone lysine methyltransferase, is a key determinant of the localization of meiotic recombination hot spots in humans and mice and the only vertebrate protein known to be involved in hybrid sterility. Here, we report the crystal structure of the PRDM9 methyltransferase domain in complex with a histone H3 peptide dimethylated on lysine 4 (H3K4me2 and S-adenosylhomocysteine (AdoHcy, which provides insights into the methyltransferase activity of PRDM proteins. We show that the genuine substrate of PRDM9 is histone H3 lysine 4 (H3K4 and that the enzyme possesses mono-, di-, and trimethylation activities. We also determined the crystal structure of PRDM9 in its autoinhibited state, which revealed a rearrangement of the substrate and cofactor binding sites by a concerted action of the pre-SET and post-SET domains, providing important insights into the regulatory mechanisms of histone lysine methyltransferase activity.

  12. An audit of thiopurine methyltransferase genotyping and phenotyping before intended azathioprine treatment for dermatological conditions

    DEFF Research Database (Denmark)

    Vestergaard, T; Bygum, A

    2009-01-01

    Summary Background. Determining thiopurine methyltransferase (TPMT) genotype and phenotype before azathioprine treatment predicts which patients are most likely to develop myelosuppression. Aim. To evaluate the course of azathioprine treatment in people with TPMT heterozygosity and whether this d...

  13. Evolved Gas Analyses of Sedimentary Materials in Gale Crater, Mars: Results of the Curiosity Rover's Sample Analysis at Mars (SAM) Instrument from Yellowknife Bay to the Stimson Formation

    Science.gov (United States)

    Sutter, B.; McAdam, A. C.; Rampe, E. B.; Ming, D. W.; Mahaffy, P. R.; Navarro-Gonzalez, R.; Stern, J. C.; Eigenbrode, J. L.; Archer, P. D.

    2016-01-01

    The Sample Analysis at Mars (SAM) instrument aboard the Mars Science Laboratory rover has analyzed 10 samples from Gale Crater. All SAM evolved gas analyses have yielded a multitude of volatiles (e.g, H2O, SO2, H2S, CO2, CO, NO, O2, HC1). The objectives of this work are to 1) Characterize the evolved H2O, SO2, CO2, and O2 gas traces of sediments analyzed by SAM through sol 1178, 2) Constrain sediment mineralogy/composition based on SAM evolved gas analysis (SAM-EGA), and 3) Discuss the implications of these results releative to understanding the geochemical history of Gale Crater.

  14. Presence of DNA methyltransferase activity and CpC methylation in Drosophila melanogaster.

    Science.gov (United States)

    Panikar, Chitra S; Rajpathak, Shriram N; Abhyankar, Varada; Deshmukh, Saniya; Deobagkar, Deepti D

    2015-12-01

    Drosophila melanogaster lacks DNMT1/DNMT3 based methylation machinery. Despite recent reports confirming the presence of low DNA methylation in Drosophila; little is known about the methyltransferase. Therefore, in this study, we have aimed to investigate the possible functioning of DNA methyltransferase in Drosophila. The 14 K oligo microarray slide was incubated with native cell extract from adult Drosophila to check the presence of the methyltransferase activity. After incubation under appropriate conditions, the methylated oligo sequences were identified by the binding of anti 5-methylcytosine monoclonal antibody. The antibody bound to the methylated oligos was detected using Cy3 labeled secondary antibody. Methylation sensitive restriction enzyme mediated PCR was used to assess the methylation at a few selected loci identified on the array. It could be seen that a few of the total oligos got methylated under the assay conditions. Analysis of methylated oligo sequences provides evidence for the presence of de novo methyltransferase activity and allows identification of its sequence specificity in adult Drosophila. With the help of methylation sensitive enzymes we could detect presence of CpC methylation in the selected genomic regions. This study reports presence of an active DNA methyltransferase in adult Drosophila, which exhibits sequence specificity confirmed by presence of asymmetric methylation at corresponding sites in the genomic DNA. It also provides an innovative approach to investigate methylation specificity of a native methyltransferase.

  15. Wide Range Vacuum Pumps for the SAM Instrument on the MSL Curiosity Rover

    Science.gov (United States)

    Sorensen, Paul; Kline-Schoder, Robert; Farley, Rodger

    2014-01-01

    Creare Incorporated and NASA Goddard Space Flight Center developed and space qualified two wide range pumps (WRPs) that were included in the Sample Analysis at Mars (SAM) instrument. This instrument was subsequently integrated into the Mars Science Laboratory (MSL) "Curiosity Rover," launched aboard an Atlas V rocket in 2011, and landed on August 6, 2012, in the Gale Crater on Mars. The pumps have now operated for more than 18 months in the Gale Crater and have been evacuating the key components of the SAM instrument: a quadrupole mass spectrometer, a tunable laser spectrometer, and six gas chromatograph columns. In this paper, we describe the main design challenges and the ways in which they were solved. This includes the custom design of a miniaturized, high-speed motor to drive the turbo drag pump rotor, analysis of rotor dynamics for super critical operation, and bearing/lubricant design/selection.

  16. Carbon tax effects on the poor: a SAM-based approach

    Science.gov (United States)

    Chapa, Joana; Ortega, Araceli

    2017-09-01

    A SAM-based price model for Mexico is developed in order to assess the effects of the carbon tax, which was part of the fiscal reform approved in 2014. The model is formulated based on a social accounting matrix (SAM) that distinguishes households by the official poverty condition and geographical area. The main results are that the sector that includes coke, refined petroleum and nuclear fuel shows the highest price increase due to the direct impact of the carbon tax; in addition, air transport and inland transport are the most affected sectors, in an indirect manner, because both employ inputs from the former sector. Also, it is found that welfare diminishes more in the rural strata than in the urban one. In the urban area, the carbon tax is regressive: the negative impact of carbon tax on family welfare is greater on the poorest families.

  17. Parabolic Trough Reference Plant for Cost Modeling with the Solar Advisor Model (SAM)

    Energy Technology Data Exchange (ETDEWEB)

    Turchi, C.

    2010-07-01

    This report describes a component-based cost model developed for parabolic trough solar power plants. The cost model was developed by the National Renewable Energy Laboratory (NREL), assisted by WorleyParsons Group Inc., for use with NREL's Solar Advisor Model (SAM). This report includes an overview and explanation of the model, two summary contract reports from WorleyParsons, and an Excel spreadsheet for use with SAM. The cost study uses a reference plant with a 100-MWe capacity and six hours of thermal energy storage. Wet-cooling and dry-cooling configurations are considered. The spreadsheet includes capital and operating cost by component to allow users to estimate the impact of changes in component costs.

  18. REPRESENTASI HOMOSEKSUALITAS DI YOUTUBE: (Studi Semiotika pada Video Pernikahan Sam Tsui

    Directory of Open Access Journals (Sweden)

    Lilis Rucirisyanti

    2017-12-01

    Full Text Available Abstract. Social media is instrumental in giving effect to nitizens, good effects or bad effect, then social media can be also represent a person. Diserve social media make it interesting for nitizens. One of social media is Youtube. Many a lot of video at there, strat from tips and trick videos, journey or vacation video, wedding video, and ect. Everyone can publish their video on Youtube. No exception of same sex enthusiast, in this study are homosexual or gay. One of is a wedding video Sam Tsui and Casey Braves. This research is a qualitative research and this research uses semiotcs analysus of Roland Barthes. By doing an analysis of video that have been published by Sam on Youtube, also do document search and literature. The author sees the existence of verbal and non vebal forms of representation from same sex merriage video of men and men.

  19. Sam68 promotes Schwann cell proliferation by enhancing the PI3K/Akt pathway and acts on regeneration after sciatic nerve crush

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Weijie, E-mail: 459586768@qq.com; Liu, Yuxi, E-mail: 924013616@qq.com; Wang, Youhua, E-mail: wyouhua1516@163.com

    2016-05-13

    Sam68 (Src-associated in mitosis of 68 kD), a KH domain RNA-binding protein, is not only important in signaling transduction cascades, but crucial in a variety of cellular processes. Sam68 is reported to be involved in the phospoinositide3-kinase (PI3K) and nuclear factor-kappa B (NF-κB) signaling pathways, and it is closely associated with cell proliferation, RNA metabolism, and tumor progression. However, we know little about the role of Sam68 during peripheral nervous system injury and regeneration. In this study, we investigated the expression of Sam68 and its biological significances in sciatic nerve crush. Interestingly, we found Sam68 had a co-localization with S100 (Schwann cell marker). Moreover, after crush, Sam68 had a spatiotemporal protein expression, which was in parallel with proliferation cell nuclear antigen (PCNA). In vitro, we also observed increased expression of Sam68 during the process of TNF-α-induced Schwann cell proliferation model. Besides, flow cytometry analyses, CCK-8, and EDU were all performed with the purpose of investigating the role of Sam68 in the regulation of Schwann cell proliferation. Even more importantly, we discovered that Sam68 could enhance the phosphorylation of Akt while LY294002 (a PI3K inhibitor) obviously reversed Sam68-induced cell proliferation. Finally, we detected the variance during regeneration progress through the rat walk footprint test. In summary, all these evidences demonstrated that Sam68 might participate in Schwann cell proliferation partially via PI3K/Akt pathway and also regulate regeneration after sciatic nerve crush. -- Highlights: •The dynamic changes and location of Sam68 after sciatic nerve crush. •Sam68 promoted Schwann cell proliferation via PI3K/Akt pathway. •Sam68 modulated functional recovery after sciatic nerve crush.

  20. Monovalent Cation Activation of the Radical SAM Enzyme Pyruvate Formate-Lyase Activating Enzyme

    OpenAIRE

    Shisler, Krista A.; Hutcheson, Rachel U.; Horitani, Masaki; Duschene, Kaitlin S.; Crain, Adam V.; Byer, Amanda S.; Shepard, Eric M.; Rasmussen, Ashley; Yang, Jian; Broderick, William E.; Vey, Jessica L.; Drennan, Catherine L.; Hoffman, Brian M.; Broderick, Joan B

    2017-01-01

    Pyruvate formate-lyase activating enzyme (PFL-AE) is a radical S-adenosyl-l-methionine (SAM) enzyme that installs a catalytically essential glycyl radical on pyruvate formate-lyase. We show that PFL-AE binds a catalytically essential monovalent cation at its active site, yet another parallel with B12 enzymes, and we characterize this cation site by a combination of structural, biochemical, and spectroscopic approaches. Refinement of the PFL-AE crystal structure reveals Na+ as the most likely ...

  1. Peran Pimpinan Dalam Meningkatkan Kualitas Pelayanan Di Perpustakaan Fakultas Teknik Universitas Sam Ratulangi Manado

    OpenAIRE

    Kojoh, Fricky F

    2014-01-01

    Ladership role in improving the quality of library services in university engineering faculty Sam Ratulangi. The role of leadership is crucial to a library, The low quality of service in the library, certainly caused by many factors. Fator factors include the low role of leadership, employee performance, interpersonal communication, commitment to employees, the duties and others. Research Objectives How to determine the role of leadership in improving the quality of services at the Library of...

  2. Parabolic Trough Collector Cost Update for the System Advisor Model (SAM)

    Energy Technology Data Exchange (ETDEWEB)

    Kurup, Parthiv [National Renewable Energy Lab. (NREL), Golden, CO (United States); Turchi, Craig S. [National Renewable Energy Lab. (NREL), Golden, CO (United States)

    2015-11-01

    This report updates the baseline cost for parabolic trough solar fields in the United States within NREL's System Advisor Model (SAM). SAM, available at no cost at https://sam.nrel.gov/, is a performance and financial model designed to facilitate decision making for people involved in the renewable energy industry. SAM is the primary tool used by NREL and the U.S. Department of Energy (DOE) for estimating the performance and cost of concentrating solar power (CSP) technologies and projects. The study performed a bottom-up build and cost estimate for two state-of-the-art parabolic trough designs -- the SkyTrough and the Ultimate Trough. The SkyTrough analysis estimated the potential installed cost for a solar field of 1500 SCAs as $170/m2 +/- $6/m2. The investigation found that SkyTrough installed costs were sensitive to factors such as raw aluminum alloy cost and production volume. For example, in the case of the SkyTrough, the installed cost would rise to nearly $210/m2 if the aluminum alloy cost was $1.70/lb instead of $1.03/lb. Accordingly, one must be aware of fluctuations in the relevant commodities markets to track system cost over time. The estimated installed cost for the Ultimate Trough was only slightly higher at $178/m2, which includes an assembly facility of $11.6 million amortized over the required production volume. Considering the size and overall cost of a 700 SCA Ultimate Trough solar field, two parallel production lines in a fully covered assembly facility, each with the specific torque box, module and mirror jigs, would be justified for a full CSP plant.

  3. Hubungan Penggunaan Laptop Dan Fungsi Penglihatan Mahasiswa Angkatan 2011 Fakultas Kedokteran Universitas Sam Ratulangi Manado

    OpenAIRE

    Ningsih, Sri S; Lintong, Fransiska; Rumampuk, Jimmy F

    2015-01-01

    : Nowadays, laptop is very trendy because it can be used anywhere. It is very useful in teaching and learning. This study aimed to determine the relation of the use of laptop and visual function. This was a descriptive study using questionnairre and survey. Respondents were students Faculty of Medicine, University of Sam Ratulangi batch 2011. Based on duration, time interval, occurence of complaints, and laptop lighting, the bivariate analysis showed a P value >0.05. Conclusion: There was no ...

  4. Efektivitas Kelompok Diskusi Tutorial Problem Based Learning di Fakultas Kedokteran Universitas Sam Ratulangi

    OpenAIRE

    Pioh, Virgin Enjel; Mewo, Yanti; Berhimpon, Siemona

    2016-01-01

    : The Faculty of Medicine, Sam Ratulangi University has applied the Problem Based Learning (PBL) method in which the tutorial group discussion is the center in this learning method. The purpose of PBL is to improve the students' problem solving skill which is essential for the students who are going to be professional medical practitioners in the future. Therefore, the effectiveness of the tutorial group discussion of PBL must be implemented in order to achieve the learning goalsThe cognitive...

  5. New solutions for large scale functional tests in the WLCG infrastructure with SAM/Nagios: the experiments experience

    CERN Document Server

    Andreeva, J; Di Girolamo, A; Kakkar, A; Litmaath, M; Magini, N; Negri, G; Ramachandran, S; Roiser, S; Saiz, P; Saiz Santos, M D; Sarkar, B; Schovancova, J; Sciabà, A; Wakankar, A

    2012-01-01

    Since several years the LHC experiments rely on the WLCG Service Availability Monitoring framework (SAM) to run functional tests on their distributed computing systems. The SAM tests have become an essential tool to measure the reliability of the Grid infrastructure and to ensure reliable computing operations, both for the sites and the experiments. Recently the old SAM framework was replaced with a completely new system based on Nagios and ActiveMQ to better support the transition to EGI and to its more distributed infrastructure support model and to implement several scalability and functionality enhancements. This required all LHC experiments and the WLCG support teams to migrate their tests, to acquire expertise on the new system, to validate the new availability and reliability computations and to adopt new visualisation tools. In this contribution we describe in detail the current state of the art of functional testing in WLCG: how the experiments use the new SAM/Nagios framework, the advanced functiona...

  6. SAMSVM: A tool for misalignment filtration of SAM-format sequences with support vector machine.

    Science.gov (United States)

    Yang, Jianfeng; Ding, Xiaofan; Sun, Xing; Tsang, Shui-Ying; Xue, Hong

    2015-12-01

    Sequence alignment/map (SAM) formatted sequences [Li H, Handsaker B, Wysoker A et al., Bioinformatics 25(16):2078-2079, 2009.] have taken on a main role in bioinformatics since the development of massive parallel sequencing. However, because misalignment of sequences poses a significant problem in analysis of sequencing data that could lead to false positives in variant calling, the exclusion of misaligned reads is a necessity in analysis. In this regard, the multiple features of SAM-formatted sequences can be treated as vectors in a multi-dimension space to allow the application of a support vector machine (SVM). Applying the LIBSVM tools developed by Chang and Lin [Chang C-C, Lin C-J, ACM Trans Intell Syst Technol 2:1-27, 2011.] as a simple interface for support vector classification, the SAMSVM package has been developed in this study to enable misalignment filtration of SAM-formatted sequences. Cross-validation between two simulated datasets processed with SAMSVM yielded accuracies that ranged from 0.89 to 0.97 with F-scores ranging from 0.77 to 0.94 in 14 groups characterized by different mutation rates from 0.001 to 0.1, indicating that the model built using SAMSVM was accurate in misalignment detection. Application of SAMSVM to actual sequencing data resulted in filtration of misaligned reads and correction of variant calling.

  7. Determinants of tRNA Recognition by the Radical SAM Enzyme RlmN.

    Directory of Open Access Journals (Sweden)

    Christina M Fitzsimmons

    Full Text Available RlmN, a bacterial radical SAM methylating enzyme, has the unusual ability to modify two distinct types of RNA: 23S rRNA and tRNA. In rRNA, RlmN installs a methyl group at the C2 position of A2503 of 23S rRNA, while in tRNA the modification occurs at nucleotide A37, immediately adjacent to the anticodon triplet. Intriguingly, only a subset of tRNAs that contain an adenosine at position 37 are substrates for RlmN, suggesting that the enzyme carefully probes the highly conserved tRNA fold and sequence features to identify its targets. Over the past several years, multiple studies have addressed rRNA modification by RlmN, while relatively few investigations have focused on the ability of this enzyme to modify tRNAs. In this study, we utilized in vitro transcribed tRNAs as model substrates to interrogate RNA recognition by RlmN. Using chimeras and point mutations, we probed how the structure and sequence of RNA influences methylation, identifying position 38 of tRNAs as a critical determinant of substrate recognition. We further demonstrate that, analogous to previous mechanistic studies with fragments of 23S rRNA, tRNA methylation requirements are consistent with radical SAM reactivity. Together, our findings provide detailed insight into tRNA recognition by a radical SAM methylating enzyme.

  8. Integration, Validation, and Application of a PV Snow Coverage Model in SAM

    Energy Technology Data Exchange (ETDEWEB)

    Ryberg, David [National Renewable Energy Lab. (NREL), Golden, CO (United States); Freeman, Janine [National Renewable Energy Lab. (NREL), Golden, CO (United States)

    2015-09-01

    Due to the increasing deployment of PV systems in snowy climates, there is significant interest in a method capable of estimating PV losses resulting from snow coverage that has been verified for a wide variety of system designs and locations. A scattering of independent snow coverage models have been developed over the last 15 years; however, there has been very little effort spent on verifying these models beyond the system design and location on which they were based. Moreover, none of the major PV modeling software products have incorporated any of these models into their workflow. In response to this deficiency, we have integrated the methodology of the snow model developed in the paper by Marion et al. [1] into the National Renewable Energy Laboratory's (NREL) System Advisor Model (SAM). In this work we describe how the snow model is implemented in SAM and discuss our demonstration of the model's effectiveness at reducing error in annual estimations for two PV arrays. Following this, we use this new functionality in conjunction with a long term historical dataset to estimate average snow losses across the United States for a typical PV system design. The open availability of the snow loss estimation capability in SAM to the PV modeling community, coupled with our results of the nation-wide study, will better equip the industry to accurately estimate PV energy production in areas affected by snowfall.

  9. Formation and characterization of low resistivity sub-100 nm copper films deposited by electroless on SAM

    Energy Technology Data Exchange (ETDEWEB)

    Asher, T. [Dept. of Physical Electronics, Faculty of Engineering, Tel-Aviv University, Ramat-Aviv 69978, Tel-Aviv (Israel)], E-mail: tamaras3@tau.ac.il; Inberg, A.; Glickman, E.; Fishelson, N.; Shacham-Diamand, Y. [Dept. of Physical Electronics, Faculty of Engineering, Tel-Aviv University, Ramat-Aviv 69978, Tel-Aviv (Israel)

    2009-10-30

    Thin Cu films of microelectronic quality and low electrical resistivity were created by electroless deposition (ELD) onto SiO{sub 2} surface modified first with self-assembled monolayer (SAM) of 3-aminopropyltrimethoxysilane (APTMS) and activated then by 5 nm gold nano-particles (AuNPs). The presence of highly oriented amino-terminated SAM was revealed by XPS and ToF-SIMS analyses. The Cu films were deposited in boron- and phosphorous-free tartrate/formaldehyde electrolyte. Controlling the deposition rate via the solution pH permitted a minimum value in resistivity {rho}. XPS depth profile revealed that diffusion of Cu into SiO{sub 2} modified by APTMS did not take place after annealing at 220 deg. C, 4 h. Moreover, annealing resulted in the drop of electrical resistivity to {rho} = 4 {+-} 0.4 {mu}{omega} cm for the films with the thickness of 35-100 nm. This value of {rho} is several times smaller than those reported in literature for sub-100 nm Cu films deposited by electroless on different SAMs. It is speculated that nano-scale porosity and corrugated structure observed by HRTEM and AFM in the ELD Cu films contribute to the resistivity. The obtained results demonstrate a viable route for formation of low resistivity, sub-100 nm Cu films on dielectrics for microelectronic application.

  10. Integration, Validation, and Application of a PV Snow Coverage Model in SAM

    Energy Technology Data Exchange (ETDEWEB)

    Freeman, Janine M. [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Ryberg, David Severin [National Renewable Energy Laboratory (NREL), Golden, CO (United States)

    2017-08-01

    Due to the increasing deployment of PV systems in snowy climates, there is significant interest in a method capable of estimating PV losses resulting from snow coverage that has been verified for a variety of system designs and locations. Many independent snow coverage models have been developed over the last 15 years; however, there has been very little effort verifying these models beyond the system designs and locations on which they were based. Moreover, major PV modeling software products have not yet incorporated any of these models into their workflows. In response to this deficiency, we have integrated the methodology of the snow model developed in the paper by Marion et al. (2013) into the National Renewable Energy Laboratory's (NREL) System Advisor Model (SAM). In this work, we describe how the snow model is implemented in SAM and we discuss our demonstration of the model's effectiveness at reducing error in annual estimations for three PV arrays. Next, we use this new functionality in conjunction with a long term historical data set to estimate average snow losses across the United States for two typical PV system designs. The open availability of the snow loss estimation capability in SAM to the PV modeling community, coupled with our results of the nationwide study, will better equip the industry to accurately estimate PV energy production in areas affected by snowfall.

  11. Microtribological and electrochemical corrosion behaviors of polydopamine coating on APTS-SAM modified Si substrate

    Energy Technology Data Exchange (ETDEWEB)

    Ou Junfei [State Key Laboratory of Solid Lubrication, Lanzhou Institute of Chemical Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Graduate University of Chinese Academy of Sciences, Beijing 100080 (China); Wang Jinqing, E-mail: jqwang@lzb.ac.cn [State Key Laboratory of Solid Lubrication, Lanzhou Institute of Chemical Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Liu Sheng [State Key Laboratory of Solid Lubrication, Lanzhou Institute of Chemical Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Graduate University of Chinese Academy of Sciences, Beijing 100080 (China); Zhou Jinfang [State Key Laboratory of Solid Lubrication, Lanzhou Institute of Chemical Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Ren Sili, E-mail: slren@lzb.ac.cn [State Key Laboratory of Solid Lubrication, Lanzhou Institute of Chemical Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Yang Shengrong [State Key Laboratory of Solid Lubrication, Lanzhou Institute of Chemical Physics, Chinese Academy of Sciences, Lanzhou 730000 (China)

    2009-11-15

    A polydopamine coating (coded as PDAc) was prepared successfully on a Si substrate through a two-step process. Briefly, to improve the adhesion of PDAc on the Si substrate, a self-assembled monolayer of 3-aminopropyl triethoxysilane (coded as APTS-SAM) was firstly generated on the bare Si wafer. Thereafter, the PDAc with different thickness was fabricated through the chemical adsorption and autopolymerization of the dopamine hydrochloride on the APTS-SAM coated Si substrate. The formation of PDAc on the APTS-SAM modified Si substrate was proved by the characterizations of contact angle measurement, attenuated total reflectance Fourier transform infrared (ATR-FTIR) analysis, and X-ray photoelectron spectroscope (XPS), etc. The ellipsometric thickness measurement and atomic force microscopy (AFM) image analysis showed that the PDAc became thicker and rougher with the deposition time prolongation. Microtribological study showed that the thickness and roughness of the PDAc played a significant role in the tribological properties. In comparison with the bare Si substrate, the PDAc with thinner thickness possessed lower friction and was anticipated to be used as protecting coating in the field of boundary lubrication. The electrochemical corrosion behaviors of the prepared PDAc were investigated using the electrochemical station and a low corrosion current density was revealed, implying that the PDAc had good anti-corrosion capability and might find potential applications in the field of corrosion resistance.

  12. Microtribological and electrochemical corrosion behaviors of polydopamine coating on APTS-SAM modified Si substrate

    Science.gov (United States)

    Ou, Junfei; Wang, Jinqing; Liu, Sheng; Zhou, Jinfang; Ren, Sili; Yang, Shengrong

    2009-11-01

    A polydopamine coating (coded as PDAc) was prepared successfully on a Si substrate through a two-step process. Briefly, to improve the adhesion of PDAc on the Si substrate, a self-assembled monolayer of 3-aminopropyl triethoxysilane (coded as APTS-SAM) was firstly generated on the bare Si wafer. Thereafter, the PDAc with different thickness was fabricated through the chemical adsorption and autopolymerization of the dopamine hydrochloride on the APTS-SAM coated Si substrate. The formation of PDAc on the APTS-SAM modified Si substrate was proved by the characterizations of contact angle measurement, attenuated total reflectance Fourier transform infrared (ATR-FTIR) analysis, and X-ray photoelectron spectroscope (XPS), etc. The ellipsometric thickness measurement and atomic force microscopy (AFM) image analysis showed that the PDAc became thicker and rougher with the deposition time prolongation. Microtribological study showed that the thickness and roughness of the PDAc played a significant role in the tribological properties. In comparison with the bare Si substrate, the PDAc with thinner thickness possessed lower friction and was anticipated to be used as protecting coating in the field of boundary lubrication. The electrochemical corrosion behaviors of the prepared PDAc were investigated using the electrochemical station and a low corrosion current density was revealed, implying that the PDAc had good anti-corrosion capability and might find potential applications in the field of corrosion resistance.

  13. Photoresponsive SAMs on gold fabricated from azobenzene-functionalised asparagusic acid derivatives.

    Science.gov (United States)

    Siemeling, Ulrich; Bruhn, Clemens; Bretthauer, Frauke; Borg, Marta; Träger, Frank; Vogel, Florian; Azzam, Waleed; Badin, Mihaela; Strunskus, Thomas; Wöll, Christof

    2009-10-28

    We have prepared a range of azobenzene derivatives equipped with an asparagusic acid-based 1,2-dithiolane headgroup suitable for chemisorption on solid gold substrates. The formation of self-assembled monolayers (SAMs) of the amide cyclo-S2C3H5-4-C(O)NH-p-C6H4-N=N-Ph (1) and the ester cyclo-S2C3H5-4-C(O)O-p-C6H4-N=N-Ph (2) on gold was monitored in situ and in real time by optical second harmonic generation (SHG). The structure and composition of these SAMs was investigated by a range of ex situ methods, viz. ellipsometry, X-ray photoelectron spectroscopy (XPS), near-edge X-ray absorption fine structure (NEXAFS) spectroscopy and Fourier transform infrared reflection absorption spectroscopy (FTIRRAS). Reversible, but moderate, photoswitchability was observed for these one-component SAMs by ellipsometry and dynamic contact angle measurements. Use of a second 1,2-dithiolane component for lateral dilution of the photoactive terminal groups resulted in a much more pronounced photoresponse.

  14. Mitofilin and CHCHD6 physically interact with Sam50 to sustain cristae structure.

    Science.gov (United States)

    Ding, Chengli; Wu, Zhifei; Huang, Lei; Wang, Yajie; Xue, Jie; Chen, Si; Deng, Zixin; Wang, Lianrong; Song, Zhiyin; Chen, Shi

    2015-11-04

    The inner mitochondrial membrane (IMM) invaginates to form cristae and the maintenance of cristae depends on the mitochondrial contact site (MICOS) complex. Mitofilin and CHCHD6, which physically interact, are two components of the MICOS. In this study, we performed immunoprecipitation experiments with Mitofilin and CHCHD6 antibodies and identified a complex containing Mitofilin, Sam50, and CHCHD 3 and 6. Using transcription activator-like effector nucleases (TALENs), we generated knockdown/knockout clones of Mitofilin and CHCHD6. Transmission electron microscopy (TEM) revealed that vesicle-like cristae morphology appeared in cell lines lacking Mitofilin, and mitochondria exhibited lower cristae density in CHCHD6-knockout cells. Immunoblot analysis showed that knockdown of Mitofilin, but not knockout of CHCHD6, affected their binding partners that control cristae morphology. We also demonstrated that Mitofilin and CHCHD6 directly interacted with Sam50. Additionally, we observed that Mitofilin-knockdown cells showed decreased mitochondrial membrane potential (ΔΨm) and intracellular ATP content, which were minimally affected in CHCHD6-knockout cells. Taken together, we conclude that the integrity of MICOS and its efficient interaction with Sam50 are indispensable for cristae organization, which is relevant to mitochondrial function.

  15. A magnesium-induced triplex pre-organizes the SAM-II riboswitch.

    Directory of Open Access Journals (Sweden)

    Susmita Roy

    2017-03-01

    Full Text Available Our 13C- and 1H-chemical exchange saturation transfer (CEST experiments previously revealed a dynamic exchange between partially closed and open conformations of the SAM-II riboswitch in the absence of ligand. Here, all-atom structure-based molecular simulations, with the electrostatic effects of Manning counter-ion condensation and explicit magnesium ions are employed to calculate the folding free energy landscape of the SAM-II riboswitch. We use this analysis to predict that magnesium ions remodel the landscape, shifting the equilibrium away from the extended, partially unfolded state towards a compact, pre-organized conformation that resembles the ligand-bound state. Our CEST and SAXS experiments, at different magnesium ion concentrations, quantitatively confirm our simulation results, demonstrating that magnesium ions induce collapse and pre-organization. Agreement between theory and experiment bolsters microscopic interpretation of our simulations, which shows that triplex formation between helix P2b and loop L1 is highly sensitive to magnesium and plays a key role in pre-organization. Pre-organization of the SAM-II riboswitch allows rapid detection of ligand with high selectivity, which is important for biological function.

  16. Regulation of Ack1 localization and activity by the amino-terminal SAM domain

    Directory of Open Access Journals (Sweden)

    Brown Deborah A

    2010-10-01

    Full Text Available Abstract Background The mechanisms that regulate the activity of the nonreceptor tyrosine kinase Ack1 (activated Cdc42-associated kinase are poorly understood. The amino-terminal region of Ack1 is predicted to contain a sterile alpha motif (SAM domain. SAM domains share a common fold and mediate protein-protein interactions in a wide variety of proteins. Here, we addressed the importance of the Ack1 SAM domain in kinase activity. Results We used immunofluorescence and Western blotting to show that Ack1 deletion mutants lacking the N-terminus displayed significantly reduced autophosphorylation in cells. A minimal construct comprising the N-terminus and kinase domain (NKD was autophosphorylated, while the kinase domain alone (KD was not. When expressed in mammalian cells, NKD localized to the plasma membrane, while KD showed a more diffuse cytosolic localization. Co-immunoprecipitation experiments showed a stronger interaction between full length Ack1 and NKD than between full length Ack1 and KD, indicating that the N-terminus was important for Ack1 dimerization. Increasing the local concentration of purified Ack1 kinase domain at the surface of lipid vesicles stimulated autophosphorylation and catalytic activity, consistent with a requirement for dimerization and trans-phosphorylation for activity. Conclusions Collectively, the data suggest that the N-terminus of Ack1 promotes membrane localization and dimerization to allow for autophosphorylation.

  17. Nuclear Protein Sam68 Interacts with the Enterovirus 71 Internal Ribosome Entry Site and Positively Regulates Viral Protein Translation.

    Science.gov (United States)

    Zhang, Hua; Song, Lei; Cong, Haolong; Tien, Po

    2015-10-01

    Enterovirus 71 (EV71) recruits various cellular factors to assist in the replication and translation of its genome. Identification of the host factors involved in the EV71 life cycle not only will enable a better understanding of the infection mechanism but also has the potential to be of use in the development of antiviral therapeutics. In this study, we demonstrated that the cellular factor 68-kDa Src-associated protein in mitosis (Sam68) acts as an internal ribosome entry site (IRES) trans-acting factor (ITAF) that binds specifically to the EV71 5' untranslated region (5'UTR). Interaction sites in both the viral IRES (stem-loops IV and V) and the heterogeneous nuclear ribonucleoprotein K homology (KH) domain of Sam68 protein were further mapped using an electrophoretic mobility shift assay (EMSA) and biotin RNA pulldown assay. More importantly, dual-luciferase (firefly) reporter analysis suggested that overexpression of Sam68 positively regulated IRES-dependent translation of virus proteins. In contrast, both IRES activity and viral protein translation significantly decreased in Sam68 knockdown cells compared with the negative-control cells treated with short hairpin RNA (shRNA). However, downregulation of Sam68 did not have a significant inhibitory effect on the accumulation of the EV71 genome. Moreover, Sam68 was redistributed from the nucleus to the cytoplasm and interacts with cellular factors, such as poly(rC)-binding protein 2 (PCBP2) and poly(A)-binding protein (PABP), during EV71 infection. The cytoplasmic relocalization of Sam68 in EV71-infected cells may be involved in the enhancement of EV71 IRES-mediated translation. Since Sam68 is known to be a RNA-binding protein, these results provide direct evidence that Sam68 is a novel ITAF that interacts with EV71 IRES and positively regulates viral protein translation. The nuclear protein Sam68 is found as an additional new host factor that interacts with the EV71 IRES during infection and could potentially

  18. Inhibition of HIV-1 gene expression by Sam68ΔC: multiple targets but a common mechanism?

    Directory of Open Access Journals (Sweden)

    Cochrane Alan

    2009-03-01

    Full Text Available Abstract Two recent publications have explored the mechanisms by which a mutant of the host protein Sam68 blocks HIV-1 structural protein synthesis and expands its activity to encompass Nef. Although the two studies propose different mechanisms for the responses observed, it is possible that a common activity is responsible. Understanding how this Sam68 mutant discriminates among the multiple viral mRNAs promises to reveal unique properties of HIV-1 RNA metabolism.

  19. Generation of induced pluripotent stem cells (iPSCs) stably expressing CRISPR-based synergistic activation mediator (SAM)

    DEFF Research Database (Denmark)

    Xiong, Kai; Zhou, Yan; Hyttel, Poul

    2016-01-01

    Human fibroblasts were engineered to express the CRISPR-based synergistic activation mediator (SAM) complex: dCas9-VP64 and MS2-P65-HSF1. Two induced pluripotent stem cells (iPSCs) clones expressing SAM were established by transducing these fibroblasts with lentivirus expressing OCT4, SOX2, KLF4...... a novel, useful tool to investigate genetic regulation of stem cell proliferation and differentiation through CRISPR-mediated activation of endogenous genes....

  20. Suicide assessment and management initiative (SAM): evaluating the implementation and uptake of suicide prevention activities in a healthcare setting.

    OpenAIRE

    Ishmael, Kiera Suzanne

    2010-01-01

    Suicide is one of the leading causes of death among Canadian men and women from adolescence to middle age and is strongly associated with mental illness. BC Mental Health and Addiction Services has developed Suicide Assessment and Management (SAM) Guidelines to identify safety risks within its client populations. Rigourous evaluation of the SAM Guidelines Initiative is essential to determine the impact of the intervention. This paper describes the literature review and logic model for the SA...

  1. Evidence for Perchlorates and the Origin of Chlorinated Hydrocarbons Detected by SAM at the Rocknest Aeolian Deposit in Gale Crater

    Science.gov (United States)

    Glavin, Daniel P.; Freissinet, Caroline; Miller, Kristen E.; Eigenbrode, Jennifer L.; Brunner, Anna E.; Buch, Arnaud; Sutter, Brad; Archer, P. Douglas, Jr.; Atreya, Sushil K.; Brinckerhoff, William B.; hide

    2013-01-01

    A single scoop of the Rocknest aeolian deposit was sieved (less than 150 micrometers), and four separate sample portions, each with a mass of approximately 50 mg, were delivered to individual cups inside the Sample Analysis at Mars (SAM) instrument by the Mars Science Laboratory rover's sample acquisition system. The samples were analyzed separately by the SAM pyrolysis evolved gas and gas chromatograph mass spectrometer analysis modes. Several chlorinated hydrocarbons including chloromethane, dichloromethane, trichloromethane, a chloromethylpropene, and chlorobenzene were identified by SAM above background levels with abundances of approximately 0.01 to 2.3 nmol. The evolution of the chloromethanes observed during pyrolysis is coincident with the increase in O2 released from the Rocknest sample and the decomposition of a product of N-methyl-N-(tert-butyldimethylsilyl)-trifluoroacetamide (MTBSTFA), a chemical whose vapors were released from a derivatization cup inside SAM. The best candidate for the oxychlorine compounds in Rocknest is a hydrated calcium perchlorate (Ca(ClO4)2·nH2O), based on the temperature release of O2 that correlates with the release of the chlorinated hydrocarbons measured by SAM, although other chlorine-bearing phases are being considered. Laboratory analog experiments suggest that the reaction of Martian chlorine from perchlorate decomposition with terrestrial organic carbon from MTBSTFA during pyrolysis can explain the presence of three chloromethanes and a chloromethylpropene detected by SAM. Chlorobenzene may be attributed to reactions of Martian chlorine released during pyrolysis with terrestrial benzene or toluene derived from 2,6-diphenylphenylene oxide (Tenax) on the SAM hydrocarbon trap. At this time we do not have definitive evidence to support a nonterrestrial carbon source for these chlorinated hydrocarbons, nor do we exclude the possibility that future SAM analyses will reveal the presence of organic compounds native to the

  2. Supplementary Material for: A new mode of SAM domain mediated oligomerization observed in the CASKIN2 neuronal scaffolding protein

    KAUST Repository

    Smirnova, Ekaterina

    2016-01-01

    Abstract Background CASKIN2 is a homolog of CASKIN1, a scaffolding protein that participates in a signaling network with CASK (calcium/calmodulin-dependent serine kinase). Despite a high level of homology between CASKIN2 and CASKIN1, CASKIN2 cannot bind CASK due to the absence of a CASK Interaction Domain and consequently, may have evolved undiscovered structural and functional distinctions. Results We demonstrate that the crystal structure of the Sterile Alpha Motif (SAM) domain tandem (SAM1-SAM2) oligomer from CASKIN2 is different than CASKIN1, with the minimal repeating unit being a dimer, rather than a monomer. Analytical ultracentrifugation sedimentation velocity methods revealed differences in monomer/dimer equilibria across a range of concentrations and ionic strengths for the wild type CASKIN2 SAM tandem and a structure-directed double mutant that could not oligomerize. Further distinguishing CASKIN2 from CASKIN1, EGFP-tagged SAM tandem proteins expressed in Neuro2a cells produced punctae that were distinct both in shape and size. Conclusions This study illustrates a new way in which neuronal SAM domains can assemble into large macromolecular assemblies that might concentrate and amplify synaptic responses.

  3. The Porter-Whitesides Discrepancy: Revisiting Odd-Even Effects in Wetting Properties of n-Alkanethiolate SAMs

    Directory of Open Access Journals (Sweden)

    Zhengjia Wang

    2015-12-01

    Full Text Available This review discusses the Porter-Whitesides discrepancy in wetting properties of n-alkanethiolate self-assembled monolayers (SAMs. About 25 years ago, Whitesides and coworker failed to observe any odd-even effect in wetting, however, Porter and his coworker did, albeit in select cases. Most previous studies agreed with Whitesides’ results, suggesting the absence of the odd-even effect in hydrophobicity of n-alkanethiolate SAMs. Recent reports have, however, found the odd-even effect in hydrophobicity of n-alkanethiolate SAMs on smooth substrates, indicating that hydrophobicity, and analogous interfacial properties, of n-alkanethiolate SAMs significantly depends on the properties of substrate. Unfortunately, the Whitesides and Porter papers do not report on the quality of the surfaces used. Based on recent work, we inferred that the original discrepancy between Whitesides and Porter can be attributed to the quality of the surface. Odd-even effect of SAMs in charge transport, capacitance, friction, and SAM structure are also discussed in this review to inform the general discussion. The discrepancy between Porter's group and Whitesides’ group could be due to surface roughness, morphology, oxidation, and adventitious contaminants.

  4. The Seismic Aftershock Monitoring System (SAMS) for OSI - Experiences from IFE14

    Science.gov (United States)

    Gestermann, Nicolai; Sick, Benjamin; Häge, Martin; Blake, Thomas; Labak, Peter; Joswig, Manfred

    2016-04-01

    An on-site inspection (OSI) is the third of four elements of the verification regime of the Comprehensive Nuclear-Test-Ban Treaty (CTBT). The sole purpose of an OSI is to confirm whether a nuclear weapon test explosion or any other nuclear explosion has been carried out in violation of the treaty and to gather any facts which might assist in identifying any possible violator. It thus constitutes the final verification measure under the CTBT if all other available measures are not able to confirm the nature of a suspicious event. The Provisional Technical Secretariat (PTS) carried out the Integrated Field Exercise 2014 (IFE14) in the Dead Sea Area of Jordan from 3 November to 9. December 2014. It was a fictitious OSI whose aim was to test the inspection capabilities in an integrated manner. The technologies allowed during an OSI are listed in the Treaty. The aim of the Seismic Aftershock Monitoring System (SAMS) is to detect and localize aftershocks of low magnitudes of the triggering event or collapses of underground cavities. The locations of these events are expected in the vicinity of a possible previous explosion and help to narrow down the search area within an inspection area (IA) of an OSI. The success of SAMS depends on the main elements, hardware, software, deployment strategy, the search logic and not least the effective use of personnel. All elements of SAMS were tested and improved during the Built-Up Exercises (BUE) which took place in Austria and Hungary. IFE14 provided more realistic climatic and hazardous terrain conditions with limited resources. Significant variations in topography of the IA of IFE14 in the mountainous Dead Sea Area of Jordan led to considerable challenges which were not expected from experiences encountered during BUE. The SAMS uses mini arrays with an aperture of about 100 meters and with a total of 4 elements. The station network deployed during IFE14 and results of the data analysis will be presented. Possible aftershocks of

  5. Two protein lysine methyltransferases methylate outer membrane protein B from Rickettsia.

    Science.gov (United States)

    Abeykoon, Amila H; Chao, Chien-Chung; Wang, Guanghui; Gucek, Marjan; Yang, David C H; Ching, Wei-Mei

    2012-12-01

    Rickettsia prowazekii, the etiologic agent of epidemic typhus, is a potential biological threat agent. Its outer membrane protein B (OmpB) is an immunodominant antigen and plays roles as protective envelope and as adhesins. The observation of the correlation between methylation of lysine residues in rickettsial OmpB and bacterial virulence has suggested the importance of an enzymatic system for the methylation of OmpB. However, no rickettsial lysine methyltransferase has been characterized. Bioinformatic analysis of genomic DNA sequences of Rickettsia identified putative lysine methyltransferases. The genes of the potential methyltransferases were synthesized, cloned, and expressed in Escherichia coli, and expressed proteins were purified by nickel-nitrilotriacetic acid (Ni-NTA) affinity chromatography. The methyltransferase activities of the purified proteins were analyzed by methyl incorporation of radioactively labeled S-adenosylmethionine into recombinant fragments of OmpB. Two putative recombinant methyltransferases (rRP789 and rRP027-028) methylated recombinant OmpB fragments. The specific activity of rRP789 is 10- to 30-fold higher than that of rRP027-028. Western blot analysis using specific antibodies against trimethyl lysine showed that both rRP789 and rRP027-028 catalyzed trimethylation of recombinant OmpB fragments. Liquid chromatography-tandem mass spectrometry (LC/MS-MS) analysis showed that rRP789 catalyzed mono-, di-, and trimethylation of lysine, while rRP027-028 catalyzed exclusively trimethylation. To our knowledge, rRP789 and rRP027-028 are the first biochemically characterized lysine methyltransferases of outer membrane proteins from Gram-negative bacteria. The production and characterization of rickettsial lysine methyltransferases provide new tools to investigate the mechanism of methylation of OmpB, effects of methylation on the structure and function of OmpB, and development of methylated OmpB-based diagnostic assays and vaccine candidates.

  6. DNA methyltransferase inhibitor CDA-II inhibits myogenic differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Zirong [State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-Sen University, Guangzhou 510060 (China); Department of Molecular Genetics and Microbiology, Shands Cancer Center, University of Florida, Gainesville, FL 32610 (United States); Jin, Guorong [State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-Sen University, Guangzhou 510060 (China); Lin, Shuibin [Department of Molecular Genetics and Microbiology, Shands Cancer Center, University of Florida, Gainesville, FL 32610 (United States); Lin, Xiumei [Department of Hematology, Guangzhou First Municipal People' s Hospital, Guangzhou 510180 (China); Gu, Yumei [Department of Molecular Genetics and Microbiology, Shands Cancer Center, University of Florida, Gainesville, FL 32610 (United States); Zhu, Yujuan; Hu, Chengbin; Zhang, Qingjiong [State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-Sen University, Guangzhou 510060 (China); Wu, Lizi [Department of Molecular Genetics and Microbiology, Shands Cancer Center, University of Florida, Gainesville, FL 32610 (United States); Shen, Huangxuan, E-mail: shenhx@mail.sysu.edu.cn [State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-Sen University, Guangzhou 510060 (China)

    2012-06-08

    Highlights: Black-Right-Pointing-Pointer CDA-II inhibits myogenic differentiation in a dose-dependent manner. Black-Right-Pointing-Pointer CDA-II repressed expression of muscle transcription factors and structural proteins. Black-Right-Pointing-Pointer CDA-II inhibited proliferation and migration of C2C12 myoblasts. -- Abstract: CDA-II (cell differentiation agent II), isolated from healthy human urine, is a DNA methyltransferase inhibitor. Previous studies indicated that CDA-II played important roles in the regulation of cell growth and certain differentiation processes. However, it has not been determined whether CDA-II affects skeletal myogenesis. In this study, we investigated effects of CDA-II treatment on skeletal muscle progenitor cell differentiation, migration and proliferation. We found that CDA-II blocked differentiation of murine myoblasts C2C12 in a dose-dependent manner. CDA-II repressed expression of muscle transcription factors, such as Myogenin and Mef2c, and structural proteins, such as myosin heavy chain (Myh3), light chain (Mylpf) and MCK. Moreover, CDA-II inhibited C1C12 cell migration and proliferation. Thus, our data provide the first evidence that CDA-II inhibits growth and differentiation of muscle progenitor cells, suggesting that the use of CDA-II might affect skeletal muscle functions.

  7. Identification and functional characterization of lysine methyltransferases of Entamoeba histolytica.

    Science.gov (United States)

    Borbolla-Vázquez, Jessica; Orozco, Esther; Medina-Gómez, Christian; Martínez-Higuera, Aarón; Javier-Reyna, Rosario; Chávez, Bibiana; Betanzos, Abigail; Rodríguez, Mario A

    2016-07-01

    Lysine methylation of histones, a posttranslational modification catalyzed by lysine methyltransferases (HKMTs), plays an important role in the epigenetic regulation of transcription. Lysine methylation of non-histone proteins also impacts the biological function of proteins. Previously it has been shown that lysine methylation of histones of Entamoeba histolytica, the protozoan parasite that infects 50 million people worldwide each year and causing up to 100,000 deaths annually, is implicated in the epigenetic machinery of this microorganism. However, the identification and characterization of HKMTs in this parasite had not yet been determined. In this work we identified four HKMTs in E. histolytica (EhHKMT1 to EhHKMT4) that are expressed by trophozoites. Enzymatic assays indicated that all of them are able to transfer methyl groups to commercial histones. EhHKMT1, EhHKMT2 and EhHKMT4 were detected in nucleus and cytoplasm of trophozoites. In addition EhHKMT2 and EhHKMT4 were located in vesicles containing ingested cells during phagocytosis, and they co-immunoprecipitated with EhADH, a protein involved in the phagocytosis of this parasite. Results suggest that E. histolytica uses its HKMTs to regulate transcription by epigenetic mechanisms, and at least two of them could also be implicated in methylation of proteins that participate in phagocytosis. © 2016 John Wiley & Sons Ltd.

  8. Cell and molecular biology of DNA methyltransferase 1.

    Science.gov (United States)

    Mohan, K Naga; Chaillet, J Richard

    2013-01-01

    The DNA cytosine methyltransferase 1 (DNMT1) is a ubiquitous nuclear enzyme that catalyzes the well-established reaction of placing methyl groups on the unmethylated cytosines in methyl-CpG:CpG base pairs in the hemimethylated DNA formed by methylated parent and unmethylated daughter strands. This activity regenerates fully methylated methyl-CpG:methyl-CpG pairs. Despite the straightforward nature of its catalytic activity, detailed biochemical, genetic, and developmental studies revealed intricate details of the central regulatory role of DNMT1 in governing the epigenetic makeup of the nuclear genome. DNMT1 mediates demethylation and also participates in seemingly wide cellular functions unrelated to maintenance DNA methylation. This review brings together mechanistic details of maintenance methylation by DNMT1, its regulation at transcriptional and posttranscriptional levels, and the seemingly unexpected functions of DNMT1 in the context of DNA methylation which is central to epigenetic changes that occur during development and the process of cell differentiation. © 2013 Elsevier Inc. All rights reserved.

  9. Theoretical insights into catalytic mechanism of protein arginine methyltransferase 1.

    Directory of Open Access Journals (Sweden)

    Ruihan Zhang

    Full Text Available Protein arginine methyltransferase 1 (PRMT1, the major arginine asymmetric dimethylation enzyme in mammals, is emerging as a potential drug target for cancer and cardiovascular disease. Understanding the catalytic mechanism of PRMT1 will facilitate inhibitor design. However, detailed mechanisms of the methyl transfer process and substrate deprotonation of PRMT1 remain unclear. In this study, we present a theoretical study on PRMT1 catalyzed arginine dimethylation by employing molecular dynamics (MD simulation and quantum mechanics/molecular mechanics (QM/MM calculation. Ternary complex models, composed of PRMT1, peptide substrate, and S-adenosyl-methionine (AdoMet as cofactor, were constructed and verified by 30-ns MD simulation. The snapshots selected from the MD trajectory were applied for the QM/MM calculation. The typical SN2-favored transition states of the first and second methyl transfers were identified from the potential energy profile. Deprotonation of substrate arginine occurs immediately after methyl transfer, and the carboxylate group of E144 acts as proton acceptor. Furthermore, natural bond orbital analysis and electrostatic potential calculation showed that E144 facilitates the charge redistribution during the reaction and reduces the energy barrier. In this study, we propose the detailed mechanism of PRMT1-catalyzed asymmetric dimethylation, which increases insight on the small-molecule effectors design, and enables further investigations into the physiological function of this family.

  10. High expression of DNA methyltransferases in primary human medulloblastoma.

    Science.gov (United States)

    Pócza, T; Krenács, T; Turányi, E; Csáthy, J; Jakab, Z; Hauser, P

    2016-01-01

    Epigenetic alterations have been implicated in cancer development. DNA methylation modulates gene expression, which is catalyzed by DNA methyltransferases (DNMTs). The objective of our study was to evaluate expression of DNMTs in medulloblastoma and analyze its correlation with clinical features. Nuclear expression of DNMT1, DNMT3A and DNMT3B was analyzed in human primary medulloblastoma of 44 patients using immunohistochemistry. Correlation of expression of DNMT levels with classical histological subtypes, novel molecular subgroups and survival of patients was analyzed. Elevated expression of DNMT1, DNMT3A and DNMT3B was observed in 63.64%, 68.18% and 72.73% of all cases, respectively. None of them showed a correlation with classical histology or survival. Concerning molecular subtypes, significantly higher expression of DNMT1 was observed in the SHH group compared to non-SHH samples (p = 0.02), but without significant difference in DNMT3A or DNMT3B levels between any subtypes. In conclusion, DNMT1, DNMT3A and DNMT3B are highly expressed in human medulloblastoma samples, suggesting that promoter hypermethylation may play a role in medulloblastoma development. Demethylation of tumor suppressor gene promoters may be considered as a possible future target in therapy of medulloblastoma.

  11. Germinal center dysregulation by histone methyltransferase EZH2 promotes lymphomagenesis

    Science.gov (United States)

    Caganova, Marieta; Carrisi, Chiara; Varano, Gabriele; Mainoldi, Federica; Zanardi, Federica; Germain, Pierre-Luc; George, Laura; Alberghini, Federica; Ferrarini, Luca; Talukder, Asoke K.; Ponzoni, Maurilio; Testa, Giuseppe; Nojima, Takuya; Doglioni, Claudio; Kitamura, Daisuke; Toellner, Kai-M.; Su, I-hsin; Casola, Stefano

    2013-01-01

    Protection against deadly pathogens requires the production of high-affinity antibodies by B cells, which are generated in germinal centers (GCs). Alteration of the GC developmental program is common in many B cell malignancies. Identification of regulators of the GC response is crucial to develop targeted therapies for GC B cell dysfunctions, including lymphomas. The histone H3 lysine 27 methyltransferase enhancer of zeste homolog 2 (EZH2) is highly expressed in GC B cells and is often constitutively activated in GC-derived non-Hodgkin lymphomas (NHLs). The function of EZH2 in GC B cells remains largely unknown. Herein, we show that Ezh2 inactivation in mouse GC B cells caused profound impairment of GC responses, memory B cell formation, and humoral immunity. EZH2 protected GC B cells against activation-induced cytidine deaminase (AID) mutagenesis, facilitated cell cycle progression, and silenced plasma cell determinant and tumor suppressor B-lymphocyte–induced maturation protein 1 (BLIMP1). EZH2 inhibition in NHL cells induced BLIMP1, which impaired tumor growth. In conclusion, EZH2 sustains AID function and prevents terminal differentiation of GC B cells, which allows antibody diversification and affinity maturation. Dysregulation of the GC reaction by constitutively active EZH2 facilitates lymphomagenesis and identifies EZH2 as a possible therapeutic target in NHL and other GC-derived B cell diseases. PMID:24200695

  12. The Role of Protein Arginine Methyltransferases in Inflammatory Responses

    Directory of Open Access Journals (Sweden)

    Ji Hye Kim

    2016-01-01

    Full Text Available Protein arginine methyltransferases (PRMTs mediate the methylation of a number of protein substrates of arginine residues and serve critical functions in many cellular responses, including cancer development, progression, and aggressiveness, T-lymphocyte activation, and hepatic gluconeogenesis. There are nine members of the PRMT family, which are divided into 4 types (types I–IV. Although most PRMTs do not require posttranslational modification (PTM to be activated, fine-tuning modifications, such as interactions between cofactor proteins, subcellular compartmentalization, and regulation of RNA, via micro-RNAs, seem to be required. Inflammation is an essential defense reaction of the body to eliminate harmful stimuli, including damaged cells, irritants, or pathogens. However, chronic inflammation can eventually cause several types of diseases, including some cancers, atherosclerosis, rheumatoid arthritis, and periodontitis. Therefore, inflammation responses should be well modulated. In this review, we briefly discuss the role of PRMTs in the control of inflammation. More specifically, we review the roles of four PRMTs (CARM1, PRMT1, PRMT5, and PRMT6 in modulating inflammation responses, particularly in terms of modulating the transcriptional factors or cofactors related to inflammation. Based on the regulatory roles known so far, we propose that PRMTs should be considered one of the target molecule groups that modulate inflammatory responses.

  13. Mapping the conformational space accessible to catechol-O-methyltransferase.

    Science.gov (United States)

    Ehler, Andreas; Benz, Jörg; Schlatter, Daniel; Rudolph, Markus G

    2014-08-01

    Methylation catalysed by catechol-O-methyltransferase (COMT) is the main pathway of catechol neurotransmitter deactivation in the prefrontal cortex. Low levels of this class of neurotransmitters are held to be causative of diseases such as schizophrenia, depression and Parkinson's disease. Inhibition of COMT may increase neurotransmitter levels, thus offering a route for treatment. Structure-based drug design hitherto seems to be based on the closed enzyme conformation. Here, a set of apo, semi-holo, holo and Michaelis form crystal structures are described that define the conformational space available to COMT and that include likely intermediates along the catalytic pathway. Domain swaps and sizeable loop movements around the active site testify to the flexibility of this enzyme, rendering COMT a difficult drug target. The low affinity of the co-substrate S-adenosylmethionine and the large conformational changes involved during catalysis highlight significant energetic investment to achieve the closed conformation. Since each conformation of COMT is a bona fide target for inhibitors, other states than the closed conformation may be promising to address. Crystallographic data for an alternative avenue of COMT inhibition, i.e. locking of the apo state by an inhibitor, are presented. The set of COMT structures may prove to be useful for the development of novel classes of inhibitors.

  14. Catechol-O-methyltransferase inhibitors in Parkinson's disease.

    Science.gov (United States)

    Müller, Thomas

    2015-02-01

    Inhibitors of catechol-O-methyltransferase (COMT) are commonly used as an adjunct to levodopa in patients with Parkinson's disease (PD) for the amelioration of wearing-off symptoms. This narrative review aims to discuss the role of COMT inhibitors on peripheral levodopa metabolism and continuous brain delivery of levodopa, and to describe their metabolic properties. Oral application of levodopa formulations with a dopa decarboxylase inhibitor (DDI) results in fluctuating levodopa plasma concentrations, predominantly due to the short half-life of levodopa and its slowing of gastric emptying. Following transport across the blood-brain barrier and its metabolic conversion to dopamine, these peripheral 'ups and downs' of levodopa are reflected in fluctuating dopamine levels in the synaptic cleft between presynaptic and postsynaptic dopaminergic neurons of the nigrostriatal system. As a result, pulsatile postsynaptic dopaminergic stimulation takes place and results in the occurrence of motor complications, such as wearing-off and dyskinesia. More continuous plasma behaviour was observed after the combination of levodopa/DDI formulations with COMT inhibitors. These compounds also weaken a levodopa/DDI-related homocysteine increase, as biomarker for an impaired methylation capacity, which is involved in an elevated oxidative stress exposure. These findings favour the concept of chronic levodopa/DDI application with concomitant inhibition of COMT and monoamine oxidase, since deamination of dopamine via this enzyme also generates free radicals. This triple combination is suggested as standard levodopa application in patients with PD who need levodopa, if they will tolerate it.

  15. Targeting histone methyltransferase EZH2 as cancer treatment.

    Science.gov (United States)

    Kondo, Yutaka

    2014-11-01

    It is widely accepted that epigenetic alterations are associated with different stages of tumour formation and progression in many cancers. Therefore, epigenetic abnormalities in cancers are emerging as important biomarkers and may have therapeutic potential. The polycomb repressive complex 2 (PRC2) is a key epigenetic regulator that catalyses trimethylation of lysine 27 on histone H3 (H3K27me3) via the histone methyltransferase, EZH2, which confers stemness and regulates differentiation during embryonic development. Given these roles of EZH2 and H3K27me3, plastic and dynamic features of cancer cells, especially cancer stem cells (CSCs), may be closely associated with this epigenetic mechanism. In addition, recent sequencing technology revealed that there are many recurrent mutations in polycomb-related genes, including EZH2, in different types of cancers. Therefore, researchers focused on targeting EZH2 as a novel cancer treatment and identified small compounds that inhibit EZH2 activity. Some of them are now under clinical trial in B-cell lymphoma. However, the underlying mechanisms by which PRC2 precisely regulate epigenetic alterations at certain genomic loci under different cellular conditions remain unclear. In this review, I focus on the recent advancements in EZH2 research, especially its dynamic regulation of epigenetic alterations in tumour cells, including the CSC population, and discuss perspectives and challenges for cancer treatment in the near future. © The Authors 2014. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

  16. Characterization of DNA methyltransferase and demethylase genes in Fragaria vesca.

    Science.gov (United States)

    Gu, Tingting; Ren, Shuai; Wang, Yuanhua; Han, Yuhui; Li, Yi

    2016-06-01

    DNA methylation is an epigenetic modification essential for gene regulations in plants, but understanding on how it is involved in fruit development, especially in non-climacteric fleshy fruit, is limited. The diploid woodland strawberry (Fragaria vesca) is an important model for non-climacteric fruit crops. In this study, we identified DNA methyltransferase genes and demethylase genes in Fragaria vesca and other angiosperm species. In accordance with previous studies, our phylogenetic analyses of those DNA methylation modifiers support the clustering of those genes into several classes. Our data indicate that whole-genome duplications and tandem duplications contributed to the expansion of those DNA methylation modifiers in angiosperms. We have further demonstrated that some DNA methylase and demethylase genes reach their highest expression levels in strawberry fleshy fruits when turning from white to red, suggesting that DNA methylation might undergo a dramatic change at the onset of fleshy fruit-ripening process. In addition, we have observed that expression of some DNA demethylase genes increases in response to various abiotic stresses including heat, cold, drought and salinity. Collectively, our study indicates a regulatory role of DNA methylation in the turning stage of non-climacteric fleshy fruit and responses to environment stimuli, and would facilitate functional studies of DNA methylation in the growth and development of non-climacteric fruits.

  17. Catechol-O-methyltransferase gene polymorphism and vulvar pain in women with vulvodynia.

    Science.gov (United States)

    Patanwala, Insiyyah Y; Lamvu, Georgine; Ledger, William J; Witzeman, Kathryn; Marvel, Richard; Rapkin, Andrea; Bongiovanni, Ann Marie; Feranec, Jessica; Witkin, Steven S

    2017-04-01

    The underlying causes of vulvar pain in women with vulvodynia remain poorly understood. Catechol-O-methyltransferase, an enzyme that metabolizes catecholamines, is a neuromodulator that is involved with perception and sensitivity to pain. The catechol-O-methyltransferase gene is polymorphic, and a single nucleotide polymorphism is associated with low activity and heightened pain sensitivity. The variant allele that encodes this polymorphism commonly is called the "L allele" because of its low enzyme activity as opposed to the normal H (high activity) allele. The methionine-containing catechol-O-methyltransferase protein coded by the L allele results in elevated catecholamine levels, reduced inactivation of the dopaminergic and adrenergic systems, and increased sensitivity to pain. This polymorphism not only may decrease the pain threshold in response to acute pain but also may facilitate the development of chronic pain. Therefore, the objective of our study was to assess whether a variation in the catechol-O-methyltransferase genotype is involved in increased pain sensitivity in women with vulvodynia. We conducted a prospective cohort study. Buccal swabs were collected from 167 white women with vulvodynia and 107 control subjects; the DNA was tested for a single nucleotide polymorphism at position 158 (rs4680) in the catechol-O-methyltransferase gene. Women with vulvodynia had a marginally increased, yet not significant, prevalence of the catechol-O-methyltransferase genotype that is associated with high activity of the coded protein: 32.9% in the women with vulvodynia, as opposed to 21.5% in the control subjects (odds ratio, 1.80; 95% confidence interval, 1.02-3.15). Subgrouping the cases based on pain frequency revealed that the elevated occurrence of this catechol-O-methyltransferase genotype was present in 40.6% of the subset of women who experienced pain only with sexual intercourse vs only 21.5% of control subjects (odds ratio, 2.50; 95% confidence interval

  18. Biotechnological Production of Dimethoxyflavonoids Using a Fusion Flavonoid O-Methyltransferase Possessing Both 3'- and 7-O-Methyltransferase Activities.

    Science.gov (United States)

    Lee, Danbi; Park, Hye Lin; Lee, Sang-Won; Bhoo, Seong Hee; Cho, Man-Ho

    2017-05-26

    Although they are less abundant in nature, methoxyflavonoids have distinct physicochemical and pharmacological properties compared to common nonmethylated flavonoids. Thus, enzymatic conversion and biotransformation using genetically engineered microorganisms of flavonoids have been attempted for the efficient production of methoxyflavonoids. Because of their regiospecificity, more than two flavonoid O-methyltransferases (FOMTs) and enzyme reactions are required to biosynthesize di(or poly)-methoxyflavonoids. For the one-step biotechnological production of bioactive di-O-methylflavonoids, we generated a multifunctional FOMT fusing a 3'-OMT (SlOMT3) and a 7-OMT (OsNOMT). The SlOMT3/OsNOMT fusion enzyme possessed both 3'- and 7-OMT activities to diverse flavonoid substrates, which were comparable to those of individual SlOMT3 and OsNOMT. The SlOMT3/OsNOMT enzyme also showed 3'- and 7-OMT activity for 7- or 3'-O-methylflavonoids, respectively, suggesting that the fusion enzyme can sequentially methylate flavonoids into di-O-methylflavonoids. The biotransformation of the flavonoids quercetin, luteolin, eriodictyol, and taxifolin using SlOMT3/OsNOMT-transformed Escherichia coli generated corresponding di-O-methylflavonoids, rhamnazin, velutin, 3',7-di-O-methyleriodictyol, and 3',7-di-O-methyltaxifolin, respectively. These results indicate that dimethoxyflavonoids may be efficiently produced from nonmethylated flavonoid precursors through a one-step biotransformation using the engineered E. coli harboring the SlOMT3/OsNOMT fusion gene.

  19. Case Study for the Evaluation of Current Treatment Recommendations of Guanidinoacetate Methyltransferase Deficiency: Ineffectiveness of Sodium Benzoate

    NARCIS (Netherlands)

    Mercimek-Mahmutoglu, S.; Salomons, G.S.; Chan, A.

    2014-01-01

    BACKGROUND: Guanidinoacetate methyltransferase deficiency is an autosomal recessively inherited disorder of creatine biosynthesis. We report a new patient with guanidinoacetate methyltransferase deficiency and her >3-year treatment outcome. Patient This is a 6-year-old girl who was diagnosed with

  20. Catechol-O-methyltransferase association with hemoglobin A1c.

    Science.gov (United States)

    Hall, Kathryn T; Jablonski, Kathleen A; Chen, Ling; Harden, Maegan; Tolkin, Benjamin R; Kaptchuk, Ted J; Bray, George A; Ridker, Paul M; Florez, Jose C; Mukamal, Kenneth J; Chasman, Daniel I

    2016-07-01

    Catecholamines have metabolic effects on blood pressure, insulin sensitivity and blood glucose. Genetic variation in catechol-O-methyltransferase (COMT), an enzyme that degrades catecholamines, is associated with cardiometabolic risk factors and incident cardiovascular disease (CVD). Here we examined COMT effects on glycemic function and type 2 diabetes. We tested whether COMT polymorphisms were associated with baseline HbA1c in the Women's Genome Health Study (WGHS), and Meta-Analyses of Glucose and Insulin-related traits Consortium (MAGIC), and with susceptibility to type 2 diabetes in WGHS, DIAbetes Genetics Replication And Meta-analysis consortium (DIAGRAM), and the Diabetes Prevention Program (DPP). Given evidence that COMT modifies some drug responses, we examined association with type 2 diabetes and randomized metformin and aspirin treatment. COMT rs4680 high-activity G-allele was associated with lower HbA1c in WGHS (β=-0.032% [0.012], p=0.008) and borderline significant in MAGIC (β=-0.006% [0.003], p=0.07). Combined COMT per val allele effects on type 2 diabetes were significant (OR=0.98 [0.96-0.998], p=0.03) in fixed-effects analyses across WGHS, DIAGRAM, and DPP. Similar results were obtained for 2 other COMT SNPs rs4818 and rs4633. In the DPP, the rs4680 val allele was borderline associated with lower diabetes incidence among participants randomized to metformin (HR=0.81 [0.65-1.00], p=0.05). COMT rs4680 high-activity G-allele was associated with lower HbA1c and modest protection from type 2 diabetes. The directionality of COMT associations was concordant with those previously observed for cardiometabolic risk factors and CVD. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. Catechol-O-methyltransferase, dopamine, and sleep-wake regulation.

    Science.gov (United States)

    Dauvilliers, Yves; Tafti, Mehdi; Landolt, Hans Peter

    2015-08-01

    Sleep and sleep disorders are complex and highly variable phenotypes regulated by many genes and environment. The catechol-O-methyltransferase (COMT) gene is an interesting candidate, being one of the major mammalian enzymes involved in the catabolism of catecholamines. The activity of COMT enzyme is genetically polymorphic due to a guanine-to-adenine transition at codon 158, resulting in a valine (Val) to methionine (Met) substitution. Individuals homozygous for the Val allele show higher COMT activity, and lower dopaminergic signaling in prefrontal cortex (PFC) than subjects homozygous for the Met allele. Since COMT has a crucial role in metabolising dopamine, it was suggested that the common functional polymorphism in the COMT gene impacts on cognitive function related to PFC, sleep-wake regulation, and potentially on sleep pathologies. The COMT Val158Met polymorphism may predict inter-individual differences in brain electroencephalography (EEG) alpha oscillations and recovery processes resulting from partial sleep loss in healthy individuals. The Val158Met polymorphism also exerts a sexual dimorphism and has a strong effect on objective daytime sleepiness in patients with narcolepsy-cataplexy. Since the COMT enzyme inactivates catecholamines, it was hypothesized that the response to stimulant drugs differs between COMT genotypes. Modafinil maintained executive functioning performance and vigilant attention throughout sleep deprivation in subjects with Val/Val genotype, but less in those with Met/Met genotype. Also, homozygous Met/Met patients with narcolepsy responded to lower doses of modafinil compared to Val/Val carriers. We review here the critical role of the common functional COMT gene polymorphism, COMT enzyme activity, and the prefrontal dopamine levels in the regulation of sleep and wakefulness in normal subjects, in narcolepsy and other sleep-related disorders, and its impact on the response to psychostimulants. Copyright © 2014 Elsevier Ltd. All

  2. Inhibitors of catechol-O-methyltransferase sensitize mice to pain

    Science.gov (United States)

    Kambur, O; Talka, R; Ansah, OB; Kontinen, VK; Pertovaara, A; Kalso, E; Männistö, PT

    2010-01-01

    BACKGROUND AND PURPOSE Catechol-O-methyltransferase (COMT) inhibitors are used in Parkinson's disease in which pain is an important symptom. COMT polymorphisms modulate pain and opioid analgesia in humans. In rats, COMT inhibitors have been shown to be pro-nociceptive in acute pain models, but also to attenuate allodynia and hyperalgesia in a model of diabetic neuropathy. Here, we have assessed the effects of acute and repeated administrations of COMT inhibitors on mechanical, thermal and carrageenan-induced nociception in male mice. EXPERIMENTAL APPROACH We used single and repeated administration of a peripherally restricted, short-acting (nitecapone) and also a centrally acting (3,5-dinitrocatechol, OR-486) COMT inhibitor. We also tested CGP 28014, an indirect inhibitor of COMT enzyme. Effects of OR-486 on thermal nociception were also studied in COMT deficient mice. Effects on spinal pathways were assessed in rats given intrathecal nitecapone. KEY RESULTS After single administration, both nitecapone and OR-486 reduced mechanical nociceptive thresholds and thermal nociceptive latencies (hot plate test) at 2 and 3 h, regardless of their brain penetration. These effects were still present after chronic treatment with COMT inhibitors for 5 days. Intraplantar injection of carrageenan reduced nociceptive latencies and both COMT inhibitors potentiated this reduction without modifying inflammation. CGP 28014 shortened paw flick latencies. OR-486 did not modify hot plate times in Comt gene deficient mice. Intrathecal nitecapone modified neither thermal nor mechanical nociception. CONCLUSIONS AND IMPLICATIONS Pro-nociceptive effects of COMT inhibitors were confirmed. The pro-nociceptive effects were primarily mediated via mechanisms acting outside the brain and spinal cord. COMT protein was required for these actions. PMID:20726980

  3. Catechol-o-methyltransferase and 3,4-({+/-})-methylenedioxymethamphetamine toxicity.

    Science.gov (United States)

    Herndon, Joseph M; Cholanians, Aram B; Lizarraga, Lucina E; Lau, Serrine S; Monks, Terrence J

    2014-05-01

    Metabolism of 3,4-(±)-methylenedioxymethamphetamine (MDMA) is necessary to elicit its neurotoxic effects. Perturbations in phase I and phase II hepatic enzymes can alter the neurotoxic profile of systemically administered MDMA. In particular, catechol-O-methyltransferase (COMT) plays a critical role in determining the fraction of MDMA that is converted to potentially neurotoxic metabolites. Thus, cytochrome P450 mediated demethylenation of MDMA, or its N-demethylated metabolite, 3,4-(±)-methylenedioxyamphetamine, give rise to the catechols, N-methyl-α-methyldopamine and α-methyldopamine, respectively. Methylation of these catechols by COMT limits their oxidation and conjugation to glutathione, a process that ultimately gives rise to neurotoxic metabolites. We therefore determined the effects of modulating COMT, a critical enzyme involved in determining the fraction of MDMA that is converted to potentially neurotoxic metabolites, on MDMA-induced toxicity. Pharmacological inhibition of COMT in the rat potentiated MDMA-induced serotonin deficits and exacerbated the acute MDMA-induced hyperthermic response. Using a genetic mouse model of COMT deficiency, in which mice lack a functional COMT gene, such mice displayed greater reductions in dopamine concentrations relative to their wild-type (WT) counterparts. Neither WT nor COMT deficient mice were susceptible to MDMA-induced decreases in serotonin concentrations. Interestingly, mice devoid of COMT were far more susceptible to the acute hyperthermic effects of MDMA, exhibiting greater increases in body temperature that ultimately resulted in death. Our findings support the view that COMT plays a pivotal role in determining the toxic response to MDMA.

  4. Structural Basis of Substrate Recognition in Thiopurine S-Methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Yi; Feng, Qiping; Wilk, Dennis; Adjei, Araba A.; Salavaggione, Oreste E.; Weinshilboum, Richard M.; Yee, Vivien C. (Case Western); (MCCM)

    2008-09-23

    Thiopurine S-methyltransferase (TPMT) modulates the cytotoxic effects of thiopurine prodrugs such as 6-mercaptopurine by methylating them in a reaction using S-adenosyl-l-methionine as the donor. Patients with TPMT variant allozymes exhibit diminished levels of protein and/or enzyme activity and are at risk for thiopurine drug-induced toxicity. We have determined two crystal structures of murine TPMT, as a binary complex with the product S-adenosyl-l-homocysteine and as a ternary complex with S-adenosyl-l-homocysteine and the substrate 6-mercaptopurine, to 1.8 and 2.0 {angstrom} resolution, respectively. Comparison of the structures reveals that an active site loop becomes ordered upon 6-mercaptopurine binding. The positions of the two ligands are consistent with the expected S{sub N}2 reaction mechanism. Arg147 and Arg221, the only polar amino acids near 6-mercaptopurine, are highlighted as possible participants in substrate deprotonation. To probe whether these residues are important for catalysis, point mutants were prepared in the human enzyme. Substitution of Arg152 (Arg147 in murine TPMT) with glutamic acid decreases V{sub max} and increases K{sub m} for 6-mercaptopurine but not K{sub m} for S-adenosyl-l-methionine. Substitution at this position with alanine or histidine and similar substitutions of Arg226 (Arg221 in murine TPMT) result in no effect on enzyme activity. The double mutant Arg152Ala/Arg226Ala exhibits a decreased V{sub max} and increased K{sub m} for 6-mercaptopurine. These observations suggest that either Arg152 or Arg226 may participate in some fashion in the TPMT reaction, with one residue compensating when the other is altered, and that Arg152 may interact with substrate more directly than Arg226, consistent with observations in the murine TPMT crystal structure.

  5. Histone methyltransferase Setdb1 is indispensable for Meckel's cartilage development.

    Science.gov (United States)

    Yahiro, Kohei; Higashihori, Norihisa; Moriyama, Keiji

    2017-01-22

    The histone methyltransferase Setdb1 represses gene expression by catalyzing lysine 9 of histone H3 trimethylation. Given that the conventional knockout of Setdb1 is embryo-lethal at the implantation stage, its role in craniofacial development is poorly understood. Here, we investigated the role of Setdb1, using conditional knockout mice-in which Setdb1 was deleted in the Meckel's cartilage (Setdb1 CKO)-and the mouse chondrogenic cell line ATDC5-in which Setdb1 was inhibited by siRNA. Deletion of Setdb1 in Meckel's cartilage, the supportive tissue in the embryonic mandible, led to its enlargement, instead of the degeneration that normally occurs. Chondrocytes from the Meckel's cartilage of Setdb1 CKO mice showed increased size. Furthermore, at embryonic days 16.5 and 18.5, part of the perichondrium was disrupted and mineralization was observed in the Meckel's cartilage. Proliferation analysis showed that inhibition of Setdb1 caused increased proliferation in chondrocytes in the Meckel's cartilage as well as in ATDC5 cells. Quantitative RT-PCR showed decreased expression of chondrogenic genes, such as Sox9, Mmp13, Collagen II, and Aggrecan, as a result of Setdb1 inhibition in ATDC5 cells. Along with these phenomenons, SMAD-dependent BMP signaling was significantly increased by the loss of Setdb1 in both the Meckel's cartilage of Setdb1 CKO mice and ATDC5 cells. Therefore, the abnormal development of Meckel's cartilage in Setdb1 CKO mice is partly due to the enhanced SMAD-dependent BMP signaling. Overall, to our knowledge, the present study is the first to show that epigenetic regulation by Setdb1 is indispensable for the embryonic development of Meckel's cartilage. Copyright © 2016. Published by Elsevier Inc.

  6. Effects of nickel on DNA methyltransferase activity and genomic DNA methylation levels.

    Science.gov (United States)

    Lee, Y W; Broday, L; Costa, M

    1998-07-31

    Methylation of DNA plays an important role in organizing the genome into transcriptionally active and inactive zones. Nickel compounds cause chromatin condensation and DNA methylation in the transgenic gpt+ Chinese hamster cell line (G12). Here we show that nickel is an inhibitor of cytosine 5-methyltransferase activity in vivo and in vitro. In living cells, this inhibition is transient and following a recovery period after nickel treatment, Mtase activity slightly rebounds. Genomic DNA methylation levels are also somewhat decreased following nickel treatment, but with time, there is an elevation of total DNA methylation above basal levels and before any rebound of methyltransferase activity. These results suggest that nickel exposure can elevate total genomic DNA methylation levels even when DNA methyltransferase activity is depressed. These findings may explain the hypermethylation of senescence and tumor suppressor genes found during nickel carcinogenesis and support the model of a direct effect of Ni2+ on chromatin leading to de novo DNA methylation.

  7. Purification and characterization of recombinant baculovirus-expressed mouse DNA methyltransferase.

    Science.gov (United States)

    Glickman, J F; Flynn, J; Reich, N O

    1997-01-13

    DNA methylation is essential for normal embryonic development in mice. An understanding of how DNA methylation is controlled is largely dependent upon the isolation and characterization of the cellular components of the DNA methylation system. The enzyme which methylates DNA in eukaryotic cells is a C-5 cytosine DNA methyltransferase. Historically, the characterization of this enzyme has been limited by its availability and purity. Here, we present a single-step purification of 4 mg of baculovirus-expressed mouse DNA methyltransferase containing a nickel-affinity leader peptide. The recombinant DNA methyltransferase copurified with inhibitory RNA which was removed by treatment with ribonuclease A. Like its non-recombinant counterpart, the recombinant enzyme is activated by hemi-methylation. A direct steady-state kinetic comparison between the recombinant baculovirus-expressed enzyme with its MEL cell-derived counterpart is presented.

  8. Rapid and sensitive single-step radiochemical assay for catechol-O-methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Zuercher, G.; Da Prada, M. (Hoffmann-La Roche (F.) and Co., Basel (Switzerland))

    1982-01-01

    A simple, rapid and reliable radiometric assay for the determination of catechol-O-methyltransferase activity is described. The method is based on the conversion of catechol to (/sup 3/H)guaiacol by catechol-O-methyltransferase in the presence of Mg/sup 2 +/, adenosine deaminase and S-adenosyl L-(methyl-/sup 3/H)methionine. Incubation and direct extraction of (/sup 3/H)guaiacol into organic scintillation fluid, as well as counting, are performed in the same standard scintillation vial. The assay is easy to perform and more sensitive than previous analogous procedures. The method has been applied to the assay of catechol-O-methyltransferase activity in discrete brain areas and also peripheral organs of rat and in human erythrocytes.

  9. Loving Sam.

    Science.gov (United States)

    Lasky, Kathryn

    1998-01-01

    Describes how the author came to love Mark Twain's writings, taught "Huckleberry Finn" and other of Twain's writings to a ninth-grade class in a Hasidic Jewish Academy, and learned to really love Samuel Clemens. Describes how this love inspired the author to write two books celebrating what she has valued most in 40 years of reading Mark…

  10. SAM] FORUM

    African Journals Online (AJOL)

    Iararnillo-Ayerbe F, Vallejo—Contreras ]. Frequency and clinical and dermatoscopic features of volar and ungual pigmented melanocytic lesions: a study in school children of Manizales,. Colombia. Peaiatr Dernzatol 2004; 21: 218-222. 18. Ragnarsson-Olding BK. Primary malignant melanoma of the vulva — an aggressive ...

  11. SAMS--a systems architecture for developing intelligent health information systems.

    Science.gov (United States)

    Yılmaz, Özgün; Erdur, Rıza Cenk; Türksever, Mustafa

    2013-12-01

    In this paper, SAMS, a novel health information system architecture for developing intelligent health information systems is proposed and also some strategies for developing such systems are discussed. The systems fulfilling this architecture will be able to store electronic health records of the patients using OWL ontologies, share patient records among different hospitals and provide physicians expertise to assist them in making decisions. The system is intelligent because it is rule-based, makes use of rule-based reasoning and has the ability to learn and evolve itself. The learning capability is provided by extracting rules from previously given decisions by the physicians and then adding the extracted rules to the system. The proposed system is novel and original in all of these aspects. As a case study, a system is implemented conforming to SAMS architecture for use by dentists in the dental domain. The use of the developed system is described with a scenario. For evaluation, the developed dental information system will be used and tried by a group of dentists. The development of this system proves the applicability of SAMS architecture. By getting decision support from a system derived from this architecture, the cognitive gap between experienced and inexperienced physicians can be compensated. Thus, patient satisfaction can be achieved, inexperienced physicians are supported in decision making and the personnel can improve their knowledge. A physician can diagnose a case, which he/she has never diagnosed before, using this system. With the help of this system, it will be possible to store general domain knowledge in this system and the personnel's need to medical guideline documents will be reduced.

  12. Genome mining for radical SAM protein determinants reveals multiple sactibiotic-like gene clusters.

    Directory of Open Access Journals (Sweden)

    Kiera Murphy

    Full Text Available Thuricin CD is a two-component bacteriocin produced by Bacillus thuringiensis that kills a wide range of clinically significant Clostridium difficile. This bacteriocin has recently been characterized and consists of two distinct peptides, Trnβ and Trnα, which both possess 3 intrapeptide sulphur to α-carbon bridges and act synergistically. Indeed, thuricin CD and subtilosin A are the only antimicrobials known to possess these unusual structures and are known as the sactibiotics (sulplur to alpha carbon-containing antibiotics. Analysis of the thuricin CD-associated gene cluster revealed the presence of genes encoding two highly unusual SAM proteins (TrnC and TrnD which are proposed to be responsible for these unusual post-translational modifications. On the basis of the frequently high conservation among enzymes responsible for the post-translational modification of specific antimicrobials, we performed an in silico screen for novel thuricin CD-like gene clusters using the TrnC and TrnD radical SAM proteins as driver sequences to perform an initial homology search against the complete non-redundant database. Fifteen novel thuricin CD-like gene clusters were identified, based on the presence of TrnC and TrnD homologues in the context of neighbouring genes encoding potential bacteriocin structural peptides. Moreover, metagenomic analysis revealed that TrnC or TrnD homologs are present in a variety of metagenomic environments, suggesting a widespread distribution of thuricin-like operons in a variety of environments. In-silico analysis of radical SAM proteins is sufficient to identify novel putative sactibiotic clusters.

  13. Postmodernism and Language in Sam Shepard's True West and Tooth of Crime

    Directory of Open Access Journals (Sweden)

    Bahar Mehrabi

    2012-09-01

    Full Text Available Sam Shepard has gained a reputation as one of America's foremost living playwrights. In over forty plays, Shepard has broken down traditional notions of dramaturgy in combining both modernist notions of the absurd and familiar icons from the American cultural landscape with an energy tinged by anarchy and violence. Moreover, Shepard has been considered by many critics as a postmodern dramatist. Hassan (1987 piles up a lengthy list of artists from various disciplines whose names epitomize postmodernism for him. The playwrights are: Samuel Beckett, Eugene Ionesco, Harold Pinter and only Sam Shepard and August Wilson from America. One characteristic of postmodern literature in general, is a focus on the instability of meaning and the inadequacy of language to completely and accurately represent truth, along with an irony and playfulness in the treatment of linguistic constructs. In other words, there is a questioning of language as a medium of perception and communication. Language can name the pain but it can’t be the pain; language cannot reach the actual individual feeling. Language is not strong enough to convey the intense emotion.  In postmodern poetics, there is a paradigmatic shift from the idea that language is transparent to the disclosure of its physicality, its intimacy, its obdurate persistence, and its paradoxical fragility. Thus, language is an insufficient means for transforming the ideas that exist in one's mind. The aim of the present article would be to present the ways in which the dramatic language of Sam Shepard, as a postmodern drama, demonstrates inadequacy of language in communication. In order to reach this goal, however, a number of his early dramas will be brought under scrutiny with regard to their language and style from the postmodern point of view.

  14. PhnJ – A novel radical SAM enzyme from the C–P lyase complex

    Directory of Open Access Journals (Sweden)

    Siddhesh S. Kamat

    2015-03-01

    Full Text Available PhnJ from the C–P lyase complex catalyzes the cleavage of the carbon–phosphorus bond in ribose-1-phosphonate-5-phosphate (PRPn to produce methane and ribose-1,2-cyclic-phosphate-5-phosphate (PRcP. This protein is a novel radical SAM enzyme that uses glycyl and thiyl radicals as reactive intermediates in the proposed reaction mechanism. The overall reaction is initiated with the reductive cleavage of S-adenosylmethionine (SAM by a reduced [4Fe–4S]1+-cluster to form an Ado-CH2∙ radical intermediate. This intermediate abstracts the proR hydrogen from Gly-32 of PhnJ to form Ado-CH3 and a glycyl radical. In the next step, there is hydrogen atom transfer from Cys-272 to the Gly-32 radical to generate a thiyl radical. The thiyl radical attacks the phosphorus center of the substrate, PRPn, to form a transient thiophosphonate radical intermediate. This intermediate collapses via homolytic C–P bond cleavage and hydrogen atom transfer from the proS hydrogen of Gly-32 to produce a thiophosphate intermediate, methane, and a radical intermediate at Gly-32. The final product, PRcP, is formed by nucleophilic attack of the C2-hydroxyl on the transient thiophosphate intermediate. This reaction regenerates the free thiol group of Cys-272. After hydrogen atom transfer from Cys-272 to the Gly-32 radical, the entire process is repeated with another substrate molecule without the use of another molecule of SAM or involvement from the [4Fe–4S]-cluster again.

  15. Related bifunctional restriction endonuclease-methyltransferase triplets: TspDTI, Tth111II/TthHB27I and TsoI with distinct specificities.

    Science.gov (United States)

    Zylicz-Stachula, Agnieszka; Zolnierkiewicz, Olga; Lubys, Arvydas; Ramanauskaite, Danute; Mitkaite, Goda; Bujnicki, Janusz M; Skowron, Piotr M

    2012-04-10

    We previously defined a family of restriction endonucleases (REases) from Thermus sp., which share common biochemical and biophysical features, such as the fusion of both the nuclease and methyltransferase (MTase) activities in a single polypeptide, cleavage at a distance from the recognition site, large molecular size, modulation of activity by S-adenosylmethionine (SAM), and incomplete cleavage of the substrate DNA. Members include related thermophilic REases with five distinct specificities: TspGWI, TaqII, Tth111II/TthHB27I, TspDTI and TsoI. TspDTI, TsoI and isoschizomers Tth111II/TthHB27I recognize different, but related sequences: 5'-ATGAA-3', 5'-TARCCA-3' and 5'-CAARCA-3' respectively. Their amino acid sequences are similar, which is unusual among REases of different specificity. To gain insight into this group of REases, TspDTI, the prototype member of the Thermus sp. enzyme family, was cloned and characterized using a recently developed method for partially cleaving REases. TspDTI, TsoI and isoschizomers Tth111II/TthHB27I are closely related bifunctional enzymes. They comprise a tandem arrangement of Type I-like domains, like other Type IIC enzymes (those with a fusion of a REase and MTase domains), e.g. TspGWI, TaqII and MmeI, but their sequences are only remotely similar to these previously characterized enzymes. The characterization of TspDTI, a prototype member of this group, extends our understanding of sequence-function relationships among multifunctional restriction-modification enzymes.

  16. A new prototype IIS/IIC/IIG endonuclease-methyltransferase TsoI from the thermophile Thermus scotoductus, recognising 5'-TARCCA(N11/9)-3' sequences.

    Science.gov (United States)

    Jezewska-Frackowiak, Joanna; Lubys, Arvydas; Vitkute, Jolanta; Zakareviciene, Laimute; Zebrowska, Joanna; Krefft, Daria; Skowron, Marta A; Zylicz-Stachula, Agnieszka; Skowron, Piotr M

    2015-01-20

    The Thermus sp. family of IIS/IIG/IIC enzymes includes the thermostable, bifunctional, fused restriction endonuclease (REase)-methyltransferases (MTase): TaqII, Tth111II/TthHB27I, TspGWI, TspDTI and TsoI. The enzymes are large proteins (approximately 120kDa), their enzymatic activities are affected by S-adenosylmethionine (SAM), they recognise similar asymmetric cognate sites and cleave at a distance of 11/9 nucleotides (nt). The enzymes exhibit similarities of their amino acid (aa) sequences and DNA catalytic motifs. Thermus sp. enzymes are an example of functional aa sequence homologies among REases recognising different, yet related DNA sequences. The family consists of TspGWI- and TspDTI-subfamilies. TsoI appears to be a non-identical 'triplet', related to TspDTI and Tth111II/TthHB27I. The discovery of TsoI, purified from Thermus scotoductus, is described. This prototype, displaying a novel specificity, which was determined by: (i) cleavage of a reference plasmid and bacteriophage DNA, (ii) cleavage of custom PCR DNA substrates, (iii) run-off sequencing of cleavage products and (iv) shotgun cloning and sequencing of bacteriophage lambda (λ) DNA digested with TsoI. The enzyme recognises a degenerated 5'-TARCCA-3' sequence, whereas DNA strands are cut 11/9 nt downstream. The discovery of the TsoI prototype is of practical importance in biotechnology, as it extends the palette of cleavage specificities for gene cloning. Copyright © 2014 Elsevier B.V. All rights reserved.

  17. Steric Clash in the SET Domain of Histone Methyltransferase NSD1 as a Cause of Sotos Syndrome and Its Genetic Heterogeneity in a Brazilian Cohort

    Directory of Open Access Journals (Sweden)

    Kyungsoo Ha

    2016-11-01

    Full Text Available Most histone methyltransferases (HMTase harbor a predicted Su(var3–9, Enhancer-of-zeste, Trithorax (SET domain, which transfers a methyl group to a lysine residue in their substrates. Mutations of the SET domains were reported to cause intellectual disability syndromes such as Sotos, Weaver, or Kabuki syndromes. Sotos syndrome is an overgrowth syndrome with intellectual disability caused by haploinsufficiency of the nuclear receptor binding SET domain protein 1 (NSD1 gene, an HMTase at 5q35.2–35.3. Here, we analyzed NSD1 in 34 Brazilian Sotos patients and identified three novel and eight known mutations. Using protein modeling and bioinformatic approaches, we evaluated the effects of one novel (I2007F and 21 previously reported missense mutations in the SET domain. For the I2007F mutation, we observed conformational change and loss of structural stability in Molecular Dynamics (MD simulations which may lead to loss-of-function of the SET domain. For six mutations near the ligand-binding site we observed in simulations steric clashes with neighboring side chains near the substrate S-Adenosyl methionine (SAM binding site, which may disrupt the enzymatic activity of NSD1. These results point to a structural mechanism underlying the pathology of the NSD1 missense mutations in the SET domain in Sotos syndrome. NSD1 mutations were identified in only 32% of the Brazilian Sotos patients in our study cohort suggesting other genes (including unknown disease genes underlie the molecular etiology for the majority of these patients. Our studies also found NSD1 expression to be profound in human fetal brain and cerebellum, accounting for prenatal onset and hypoplasia of cerebellar vermis seen in Sotos syndrome.

  18. A 7-Deazaadenosylaziridine Cofactor for Sequence-Specific Labeling of DNA by the DNA Cytosine-C5 Methyltransferase M.HhaI

    Directory of Open Access Journals (Sweden)

    Falk Kunkel

    2015-11-01

    Full Text Available DNA methyltransferases (MTases catalyze the transfer of the activated methyl group of the cofactor S-adenosyl-l-methionine (AdoMet or SAM to the exocyclic amino groups of adenine or cytosine or the C5 ring atom of cytosine within specific DNA sequences. The DNA adenine-N6 MTase from Thermus aquaticus (M.TaqI is also capable of coupling synthetic N-adenosylaziridine cofactor analogues to its target adenine within the double-stranded 5′-TCGA-3′ sequence. This M.TaqI-mediated coupling reaction was exploited to sequence-specifically deliver fluorophores and biotin to DNA using N-adenosylaziridine derivatives carrying reporter groups at the 8-position of the adenine ring. However, these 8-modified aziridine cofactors were poor substrates for the DNA cytosine-C5 MTase from Haemophilus haemolyticus (M.HhaI. Based on the crystal structure of M.HhaI in complex with a duplex oligodeoxynucleotide and the cofactor product, we synthesized a stable 7-deazaadenosylaziridine derivative with a biotin group attached to the 7-position via a flexible linker. This 7-modified aziridine cofactor can be efficiently used by M.HhaI for the direct, quantitative and sequence-specific delivery of biotin to the second cytosine within 5′-GCGC-3′ sequences in short duplex oligodeoxynucleotides and plasmid DNA. In addition, we demonstrate that biotinylation by M.HhaI depends on the methylation status of the target cytosine and, thus, could provide a method for cytosine-C5 DNA methylation detection in mammalian DNA.

  19. Systematic substrate adoption methodology (SAM) for future flexible, generic pharmaceutical production processes

    DEFF Research Database (Denmark)

    Singh, Ravendra; Godfrey, Andy; Gregertsen, Björn

    2013-01-01

    The discovery of an effective and safe pharmaceutical product is based on success in clinical trials. Often, several candidate compounds targeting the same disease area are tested in order to identify the most efficacious products. This involves the manufacture of small quantities of compounds...... methodology (SAM) for a series of substrates with similar molecular functionality has been developed. The objective is to achieve “flexible, fast and future” pharmaceutical production processes by adapting a generic modular process template. Application of the methodology is illustrated through a case study...... from the pharmaceutical industry. Use of computer-aided methods and tools as part of the methodology is also highlighted....

  20. A uniquely Canadian military moment: Sam Hughes and the No. 7 General Hospital, 1915-1916.

    Science.gov (United States)

    Coombs, Howard G

    2017-08-01

    Universities across Canada actively supported the call to arms in 1914, and Queen's University in Kingston, Ontario, was no different. Though a myriad of units composed of Queen's faculty and students were created, the university perceived the military hospital raised by the school's medical faculty to be among its most vital contributions to the First World War. This commentary describes the engagement of the No. 7 General Hospital with the Minister of Militia, Sam Hughes, which has become an almost unknown footnote to its illustrious story. This commentary has an Appendix, available at canjsurg.ca.

  1. Anodic Behavior of SAM2X5 Material Applied as Amorphous Coatings

    Energy Technology Data Exchange (ETDEWEB)

    Hailey, P D; Farmer, J C; Day, S D; Rebak, R B

    2007-08-10

    Iron-based amorphous alloys are desirable industrial materials since they are highly resistant to corrosion and possess enhanced hardness for wear resistance. The amorphous materials can be produced from the melt as powder and later spray deposited as coatings on large engineering structures. As a laboratory experiment, SAM2X5 powder was coated on electrochemical specimens of 304SS for testing. Results show that the coated specimens did not perform satisfactorily during the laboratory testing. This is because of partial devitrification during the deposition of the powder on the small specimen substrates.

  2. A novel Mg(2+)-dependent O-methyltransferase in the phenylpropanoid metabolism of Mesembryanthemum crystallinum.

    Science.gov (United States)

    Ibdah, Mwafaq; Zhang, Xing-Hai; Schmidt, Jürgen; Vogt, Thomas

    2003-11-07

    Upon irradiation with elevated light intensities, the ice plant (Mesembryanthemum crystallinum) accumulates a complex pattern of methylated and glycosylated flavonol conjugates in the upper epidermal layer. Identification of a flavonol methylating activity, partial purification of the enzyme, and sequencing of the corresponding peptide fragments revealed a novel S-adenosyl-l-methionine-dependent O-methyltransferase that was specific for flavonoids and caffeoyl-CoA. Cloning and functional expression of the corresponding cDNA verified that the new methyltransferase is a multifunctional 26.6-kDa Mg(2+)-dependent enzyme, which shows a significant sequence similarity to the cluster of caffeoyl coenzyme A-methylating enzymes. Functional analysis of highly homologous members from chickweed (Stellaria longipes), Arabidopsis thaliana, and tobacco (Nicotiana tabacum) demonstrated that the enzymes from the ice plant, chickweed, and A. thaliana possess a broader substrate specificity toward o-hydroquinone-like structures than previously anticipated for Mg(2+)-dependent O-methyltransferases, and are distinctly different from the tobacco enzyme. Besides caffeoyl-CoA and flavonols, a high specificity was also observed for caffeoylglucose, a compound never before reported to be methylated by any plant O-methyltransferase. Based on phylogenetic analysis of the amino acid sequence and differences in acceptor specificities among both animal and plant O-methyltransferases, we propose that the enzymes from the Centrospermae, along with the predicted gene product from A. thaliana, form a novel subclass within the caffeoyl coenzyme A-dependent O-methyltransferases, with potential divergent functions not restricted to lignin monomer biosynthesis.

  3. Evolved Gas Analyses of the Murray Formation in Gale Crater, Mars: Results of the Curiosity Rover's Sample Analysis at Mars (SAM) Instrument

    Science.gov (United States)

    Sutter, B.; McAdam, A. C.; Rampe, E. B.; Thompson, L. M.; Ming, D. W.; Mahaffy, P. R.; Navarro-Gonzalez, R.; Stern, J. C.; Eigenbrode, J. L.; Archer, P. D.

    2017-01-01

    The Sample Analysis at Mars (SAM) instrument aboard the Mars Science Laboratory rover has analyzed 13 samples from Gale Crater. All SAM-evolved gas analyses have yielded a multitude of volatiles (e.g., H2O, SO2, H2S, CO2, CO, NO, O2, HCl) [1- 6]. The objectives of this work are to 1) Characterize recent evolved SO2, CO2, O2, and NO gas traces of the Murray formation mudstone, 2) Constrain sediment mineralogy/composition based on SAM evolved gas analysis (SAM-EGA), and 3) Discuss the implications of these results relative to understanding the geological history of Gale Crater.

  4. A pilot study investigating the feasibility of symptom assessment manager (SAM), a Web-based real-time tool for monitoring challenging behaviors.

    Science.gov (United States)

    Loi, Samantha M; Wanasinghage, Sangeeth; Goh, Anita; Lautenschlager, Nicola T; Darby, David G; Velakoulis, Dennis

    2017-11-03

    Improving and minimizing challenging behaviors seen in psychiatric conditions, including behavioral and psychological symptoms of dementia are important in the care of people with these conditions. Yet there is a lack of systematic evaluation of these as a part of routine clinical care. The Neuropsychiatric Inventory is a validated and reliable tool for rating the severity and disruptiveness of challenging behaviors. We report on the evaluation of a Web-based symptom assessment manager (SAM), designed to address the limitation of previous tools using some of the Neuropsychiatric Inventory functions, to monitor behaviors by staff caring for people with dementia and other psychiatric conditions in inpatient and residential care settings. The SAM was piloted in an 8-bed inpatient neuropsychiatry unit over 5 months. Eleven nurses and 4 clinicians were trained in usage of SAM. Primary outcomes were usage of SAM and perceived usability, utility, and acceptance of SAM. Secondary outcomes were the frequencies of documented behavior. Usage data were analyzed using chi-square and logistic regression analyses. The SAM was used for all admitted patients regardless of diagnosis, with a usage rate of 64% for nurses regularly employed in the unit. Staff provided positive feedback regarding the utility of SAM. The SAM appeared to offer individualized behavior assessment by providing a quick, structured, and standardized platform for assessing behavior in a real-world setting. Further research would involve trialing SAM with more staff in alternative settings such as in home or residential care settings. Copyright © 2017 John Wiley & Sons, Ltd.

  5. Evidence for S-adenosyl-L-methionine (SAM-e) for the treatment of major depressive disorder.

    Science.gov (United States)

    Papakostas, George I

    2009-01-01

    Despite the increasingly large array of antidepressants available to treat major depressive disorder, patients continue to experience relatively modest response and remission rates. In addition, patients may experience adverse side effects from pharmacotherapy that not only hinder treatment compliance and adherence but, in some cases, may also contribute to increased disability, patient suffering, morbidity, and mortality. In order to enhance treatment efficacy and tolerability, patients and clinicians have become increasingly interested in nonpharmaceutical supplements for treating depression. One of the best-studied of these supplements is S-adenosyl-L-methionine (SAM-e), a naturally occurring molecule present in all living cells and a major methyl group donor in the human body. Controlled trials have found SAM-e to be more efficacious than placebo and equal in efficacy to the tricyclic antidepressants for treating major depressive disorder (MDD) when administered parenterally (either intravenously or intramuscularly). Less evidence supports the use of oral SAM-e, although some trials have demonstrated its efficacy as well. In addition, there is a paucity of evidence examining whether oral forms of SAM-e can be safe, well tolerated, and efficacious when used as adjunctive treatment for antidepressant nonresponders with MDD. Although preliminary data suggest SAM-e may be useful as an adjunctive therapy to antidepressants, controlled studies are needed to confirm or refute these preliminary findings. (c) Copyright 2009 Physicians Postgraduate Press, Inc.

  6. Potensi Tinggalan Arkeologis di Kawasan Bandar Udara Sam Ratulangi Manado: Upaya Pelestarian, Pemanfaatan, dan Pengembangan bagi Masyarakat

    Directory of Open Access Journals (Sweden)

    Irfanuddin W. Marzuki

    2016-08-01

    Full Text Available The area of Sam Ratulangi airport’s Manado has archaeological heritage which has been know as it is closed for public. This research used descriptive method, using inductive reasoning. Meanwhile, the analysis method used morphologyl, technology, and contextual analysis. This research aimed to find out the potential of archaeological heritage in Sam Ratulangi airport area of Manado. In addition to its strategy of preservation the haritage included veilbox, bungker, and waruga. The preservation can be conducted by doing protection, development, and utilization. The preservation both physical and non physical protection. The effort for its development and utilization was conducted for the purpose of science, education, culture, and tourism. Kawasan Bandar Udara Sam Ratulangi Manado mempunyai potensi tinggalan arkeologis yang selama ini tidak diketahui masyarakat luas, dikarenakan letak tinggalan yang berada dalam kawasan tertutup untuk umum. Penelitian menggunakan metode deskriptif dengan penalaran induktif. Metode analisis menggunakan analisis morfologi, teknologi dan kontekstual. Tujuan penelitian untuk mengetahui potensi tinggalan arkeologis yang terdapat di kawasan Bandar Udara Sam Ratulangi dan strategi pelestariannya. Tinggalan arkeologis yang terdapat di kawasan Bandar Udara Sam Ratulangi meliputi veilbox, bungker, dan waruga. Upaya pelestarian dapat dilakukan dengan cara perlindungan, pengembangan dan pemanfaatan. Upaya perlindungan meliputi perlindungan secara fisik dan non fisik. Upaya pengembangan dan pemanfaatan dilakukan untuk kepentingan ilmu pengetahuan, pendidikan, kebudayaan dan pariwisata.

  7. The RNA-methyltransferase Misu (NSun2 poises epidermal stem cells to differentiate.

    Directory of Open Access Journals (Sweden)

    Sandra Blanco

    2011-12-01

    Full Text Available Homeostasis of most adult tissues is maintained by balancing stem cell self-renewal and differentiation, but whether post-transcriptional mechanisms can regulate this process is unknown. Here, we identify that an RNA methyltransferase (Misu/Nsun2 is required to balance stem cell self-renewal and differentiation in skin. In the epidermis, this methyltransferase is found in a defined sub-population of hair follicle stem cells poised to undergo lineage commitment, and its depletion results in enhanced quiescence and aberrant stem cell differentiation. Our results reveal that post-transcriptional RNA methylation can play a previously unappreciated role in controlling stem cell fate.

  8. Cloning, expression, purification and crystallization of Schizosaccharomyces pombe Set7, a putative histone methyltransferase.

    Science.gov (United States)

    Mevius, Damiaan E H F; Shen, Yunpeng; Morishita, Masayo; di Luccio, Eric

    2016-04-01

    Dysfunction of histone-modifying enzymes affects chromatin regulation and is involved in carcinogenesis, tumour progression and other diseases. Histone methyltransferases are a family of key histone-modifying enzymes, but their structures, functions and mechanisms are incompletely understood, thus constraining drug-design efforts. Here, preliminary steps towards structure-function studies of Schizosaccharomyces pombe Set7, a putative histone methyltransferase and the first yeast full-length SET-domain-containing protein to be studied using X-ray crystallography, are reported. The methods from cloning to X-ray diffraction and phasing are discussed and the results will aid in prospective studies of histone-modifying enzymes.

  9. Structure and mechanism of an antibiotics-synthesizing 3-hydroxykynurenine C-methyltransferase.

    Science.gov (United States)

    Chen, Sheng-Chia; Huang, Chi-Hung; Lai, Shu-Jung; Liu, Jai-Shin; Fu, Pin-Kuei; Tseng, Shih-Ting; Yang, Chia Shin; Lai, Mei-Chin; Ko, Tzu-Ping; Chen, Yeh

    2015-05-11

    Streptosporangium sibiricum SibL catalyzes the methyl transfer from S-adenosylmethionine (SAM) to 3-hydroxykynurenine (3-HK) to produce S-adenosylhomocysteine (SAH) and 3-hydroxy-4-methyl-kynurenine for sibiromycin biosynthesis. Here, we present the crystal structures of apo-form Ss-SibL, Ss-SibL/SAH binary complex and Ss-SibL/SAH/3-HK ternary complex. Ss-SibL is a homodimer. Each subunit comprises a helical N-terminal domain and a Rossmann-fold C-terminal domain. SAM (or SAH) binding alone results in domain movements, suggesting a two-step catalytic cycle. Analyses of the enzyme-ligand interactions and further mutant studies support a mechanism in which Tyr134 serves as the principal base in the transferase reaction of methyl group from SAM to 3-HK.

  10. Radiometric assay for phenylethanolamine N-methyltransferase and catechol O-methyltransferase in a single tissue sample: application to rat hypothalamic nuclei, pineal gland, and heart

    Energy Technology Data Exchange (ETDEWEB)

    Culman, J.; Torda, T.; Weise, V.K.

    1987-08-01

    A simple and highly sensitive method for simultaneous assay of phenylethanolamine N-methyltransferase (PNMT) and catechol O-methyltransferase (COMT) is described. These enzymes are determined in a single tissue homogenate using S-(methyl-/sup 3/H) adenosyl-L-methionine as methyl donor and sequentially incubating with the substrates phenylethanolamine and epinephrine. The radioactive products of the enzymatic reactions, N-methylphenylethanolamine and metanephrine, are extracted and then separated by thin-layer chromatography. The identity of the reaction products has been established chromatographically and the conditions for both enzymatic reactions in the assay procedure have been defined. Measurement of PNMT activity in the rat pineal gland or in minute fragments of other tissues (e.g., brain nuclei) has not been possible using previously described methods. Activities of PNMT and COMT in the rat pineal gland, various hypothalamic nuclei, and the auricular and ventricular myocardia are herein reported.

  11. Expression of DNA Methyltransferases in Breast Cancer Patients and to Analyze the Effect of Natural Compounds on DNA Methyltransferases and Associated Proteins

    OpenAIRE

    Mirza, Sameer; Sharma, Gayatri; Parshad, Rajinder; Gupta, Sidhartha Datta; Pandya, Pranav; Ralhan, Ranju

    2013-01-01

    Purpose The DNA methylation mediated by specific DNA methyltransferases (DNMTs), results in the epigenetic silencing of multiple genes which are implicated in human breast cancer. We hypothesized that the natural compounds modulate the expression of DNMTs and their associated proteins in the breast cancer cell lines and affect the methylation mediated gene silencing. Methods The DNMTs transcript expression was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) in the tumors ...

  12. Resilience as positive coping appraisals: Testing the schematic appraisals model of suicide (SAMS).

    Science.gov (United States)

    Johnson, J; Gooding, P A; Wood, A M; Tarrier, N

    2010-03-01

    The Schematic Appraisals Model of Suicide (SAMS) suggests that positive self-appraisals may be important for buffering suicidal thoughts and behaviours, potentially providing a key source of resilience. The current study aimed to explore whether positive self-appraisals buffered individuals from suicidality in the face of stressful life events. 78 participants who reported experiencing some degree of suicidality were recruited from a student population. They completed a battery of questionnaires including measures of suicidality, stressful life events and positive self-appraisals. Positive self-appraisals moderated the association between stressful life events and suicidality. For those reporting moderate or high levels of positive self-appraisals, raised incidence of stressful life events did not lead to increases in suicidality. These results support the SAMS framework, and suggest that positive self-appraisals may confer resilience to suicide. Positive self-appraisals may be a promising avenue for further resilience research, and an important area to target for suicide interventions. 2009 Elsevier Ltd. All rights reserved.

  13. SAM - Sistema Automatizado del Método MECAP para Especificar Casos de Prueba

    Directory of Open Access Journals (Sweden)

    Kenyer Domínguez

    2010-12-01

    Full Text Available Existen cuatro elementos que son relevantes al momento de definir las pruebas: Confiabilidad, Costo, Tiempo y Calidad. El tiempo de desarrollo y el costo del producto se incrementan cuando se desean pruebas confiables y un software de calidad. Pero ¿qué se puede hacer para que los involucrados comprendan que las pruebas deben ser vistas como una red de seguridad? Si la calidad no se contempla antes de comenzar las pruebas, entonces ella no estará cuando se éstas terminen. El objetivo de este artículo es presentar la herramienta, SAM – Sistema Automatizado del Método MECAP que permite especificar Casos de Prueba a partir de Casos de Uso incorporando elementos que promueven la verificación y validación de la trazabilidad entre la Gestión de Requerimientos, el Análisis y Diseño y las Pruebas. SAM soporta el proceso de pruebas de forma automatizada, mejorando la confiabilidad de las mismas

  14. Absence of molecular slip on ultraclean and SAM-coated surfaces

    Science.gov (United States)

    Pye, Justin; Wood, Clay; Burton, Justin

    2016-11-01

    The liquid/solid boundary condition is a complex problem that is becoming increasingly important for the development of nanoscale fluidic devices. Many groups have now measured slip near an interface at nanoscale dimensions using a variety of experimental techniques. In simple systems, large slip lengths are generally measured for non-wetting liquid/solid combinations, but many conflicting measurements and interpretations remain. We have developed a novel pseudo-differential technique using a quartz crystal microbalance (QCM) to measure slip lengths on various surfaces. A drop of one liquid is grown on the QCM in the presence of a second, ambient liquid. We have isolated any anomalous boundary effects such as interfacial slip by choosing two liquids which have identical bulk effects on the QCM frequency and dissipation in the presence of no-slip. Slip lengths are -less than 2 nm- for water (relative to undecane) on all surfaces measured, including plasma cleaned gold, SiO2, and two different self assembled monolayers (SAMs), regardless of contact angle. We also find that surface cleanliness is crucial to accurately measure slip lengths. Additionally, clean glass substrates appear to have a significant adsorbed water layer and SAM surfaces show excess dissipation, possibly associated with contact line motion. In addition to investigating other liquid pairs, future work will include extending this technique to surfaces with independently controllable chemistry and roughness, both of which are known to strongly affect interfacial hydrodynamics.

  15. Constraining the MSL-SAM methane detection source location Through Mars Regional Atmospheric Modeling System (MRAMS)

    Science.gov (United States)

    Pla Garcia, J.; Rafkin, S. C. R.

    2017-09-01

    The detection of methane by SAM instrument has garnered significant attention. There are many major unresolved questions regarding this detection: 1) Where is the release location? 2) How spatially extensive is the release? 3) For how long is CH4 released? In an effort to better address the potential mixing and remaining questions, atmospheric circulation studies of Gale Crater were performed with MRAMS mesoscale model, ideally suited for this investigation. The model was focused on rover locations using nested grids with a spacing of 330 meters on the innermost grid that is centered over the detection site. In order to characterize seasonal mixing changes throughout the Martian year, simulations were conducted at Ls0, 90, 180 and 270. The rise in CH4 concentration was reported to start around Ls336, peaked shortly after Ls82, and then dropped to background prior to Ls103. The aim of this work is to establish the amount of mixing during all seasons and to test whether CH4 releases inside or outside of Gale crater are consistent with SAM observations.

  16. Human and management factors in probabilistic risk analysis: the SAM approach and observations from recent applications

    Energy Technology Data Exchange (ETDEWEB)

    Elisabeth Pate-Cornell, M.; Murphy, Dean M

    1996-08-01

    Most severe industrial accidents have been shown to involve one or more human errors and these are generally rooted in management problems. The objective of this paper is to draw some conclusions from the experience that we have acquired from three different studies of this phenomenon: (1) the Piper Alpha accident including problems of operations management and fire risks on-board offshore platforms, (2) the management of the heat shield of the NASA space shuttle orbiter, and (3) the roots of patient risks in anaesthesia. This paper describes and illustrates the SAM approach (System-Action-Management) that was developed and used in these studies to link the probabilities of system failures to human and management factors. This SAM model includes: first, a probabilistic risk analysis of the physical system, second, an analysis of the decisions and actions that affect the probabilities of its basic events, and third, a study of the management factors that influence those decisions and actions. In the three initial studies, the analytical links (conditional probabilities) among these three submodels were coarsely quantified based on statistical data whenever available, or most often, on expert opinions. This paper describes some observations that were made across these three studies, for example, the importance of the informal reward system, the difficulties in the communication of uncertainties, the problems of managing resource constraints, and the safety implications of the short cuts that they often imply.

  17. Post-translational modification of ribosomally synthesized peptides by a radical SAM epimerase in Bacillus subtilis

    Science.gov (United States)

    Benjdia, Alhosna; Guillot, Alain; Ruffié, Pauline; Leprince, Jérôme; Berteau, Olivier

    2017-07-01

    Ribosomally synthesized peptides are built out of L-amino acids, whereas D-amino acids are generally the hallmark of non-ribosomal synthetic processes. Here we show that the model bacterium Bacillus subtilis is able to produce a novel type of ribosomally synthesized and post-translationally modified peptide that contains D-amino acids, and which we propose to call epipeptides. We demonstrate that a two [4Fe-4S]-cluster radical S-adenosyl-L-methionine (SAM) enzyme converts L-amino acids into their D-counterparts by catalysing Cα-hydrogen-atom abstraction and using a critical cysteine residue as the hydrogen-atom donor. Unexpectedly, these D-amino acid residues proved to be essential for the activity of a peptide that induces the expression of LiaRS, a major component of the bacterial cell envelope stress-response system. Present in B. subtilis and in several members of the human microbiome, these epipeptides and radical SAM epimerases broaden the landscape of peptidyl structures accessible to living organisms.

  18. A Globally Distributed Grid Monitoring System to Facilitate High-Performance Computing at D0/SAM-Grid

    Energy Technology Data Exchange (ETDEWEB)

    Rana, Abhishek S. [Texas U., Arlington

    2002-01-01

    A grid environment involves large scale sharing of resources that are distributed from a geographical or an administrative perspective. There is a need for systems that enable continuous discovery and monitoring of the components of a grid. In this work, we discuss the development and deployment of a monitoring system that has been designed as a prototype for the D0/SAM-Grid. We have developed a system that uses a layered architecture for information generation and processing, utilizes the various grid middleware tools, and implements Integration and Enquiry Protocols using existing Discovery Protocols to provide a user with a coherent view of all current activity in this grid - in the form of a web portal interface. The prototype system has been deployed for monitoring of 11 sites geographically distributed in 5 countries across 3 continents. This work focuses on the D0/SAM-Grid, and is based on the SAM system developed at Fermilab.

  19. Structure and mechanism of an antibiotics-synthesizing 3-hydroxykynurenine C-methyltransferase

    OpenAIRE

    Chen, Sheng-Chia; Huang, Chi-Hung; Lai, Shu-Jung; Liu, Jai-Shin; Fu, Pin-Kuei; Tseng, Shih-Ting; Yang, Chia Shin; Lai, Mei-Chin; Ko, Tzu-Ping; Chen, Yeh

    2015-01-01

    Streptosporangium sibiricum SibL catalyzes the methyl transfer from S-adenosylmethionine (SAM) to 3-hydroxykynurenine (3-HK) to produce S-adenosylhomocysteine (SAH) and 3-hydroxy-4-methyl-kynurenine for sibiromycin biosynthesis. Here, we present the crystal structures of apo-form Ss-SibL, Ss-SibL/SAH binary complex and Ss-SibL/SAH/3-HK ternary complex. Ss-SibL is a homodimer. Each subunit comprises a helical N-terminal domain and a Rossmann-fold C-terminal domain. SAM (or SAH) binding alone r...

  20. Structural and evolutionary bioinformatics of the SPOUT superfamily of methyltransferases

    Directory of Open Access Journals (Sweden)

    Purta Elzbieta

    2007-03-01

    Full Text Available Abstract Background SPOUT methyltransferases (MTases are a large class of S-adenosyl-L-methionine-dependent enzymes that exhibit an unusual alpha/beta fold with a very deep topological knot. In 2001, when no crystal structures were available for any of these proteins, Anantharaman, Koonin, and Aravind identified homology between SpoU and TrmD MTases and defined the SPOUT superfamily. Since then, multiple crystal structures of knotted MTases have been solved and numerous new homologous sequences appeared in the databases. However, no comprehensive comparative analysis of these proteins has been carried out to classify them based on structural and evolutionary criteria and to guide functional predictions. Results We carried out extensive searches of databases of protein structures and sequences to collect all members of previously identified SPOUT MTases, and to identify previously unknown homologs. Based on sequence clustering, characterization of domain architecture, structure predictions and sequence/structure comparisons, we re-defined families within the SPOUT superfamily and predicted putative active sites and biochemical functions for the so far uncharacterized members. We have also delineated the common core of SPOUT MTases and inferred a multiple sequence alignment for the conserved knot region, from which we calculated the phylogenetic tree of the superfamily. We have also studied phylogenetic distribution of different families, and used this information to infer the evolutionary history of the SPOUT superfamily. Conclusion We present the first phylogenetic tree of the SPOUT superfamily since it was defined, together with a new scheme for its classification, and discussion about conservation of sequence and structure in different families, and their functional implications. We identified four protein families as new members of the SPOUT superfamily. Three of these families are functionally uncharacterized (COG1772, COG1901, and COG4080

  1. Methyltransferase G9A Regulates Osteogenesis via Twist Gene Repression.

    Science.gov (United States)

    Higashihori, N; Lehnertz, B; Sampaio, A; Underhill, T M; Rossi, F; Richman, J M

    2017-09-01

    Here we investigate the role of epigenetic factors in controlling the timing of cranial neural crest cell differentiation. The gene coding for histone H3 lysine 9 methyltransferase G9A was conditionally deleted in neural crest cells with Wnt1-Cre. The majority of homozygous-null animals survived to birth but thereafter failed to thrive. Phenotypic analysis of postnatal animals revealed that the mutants displayed incomplete ossification and 20% shorter jaws as compared to their wild-type littermates. At E13.5, patterns of expression of the osteogenic transcription factor RUNX2 and the mesenchymal transcription factor TWIST are similar in controls and mutants; both overlap in areas of future intramembranous bone formation. At E14.5, the nonosteogenic mesenchyme expressed TWIST, whereas the ossification center had strong RUNX2 and osteopontin expression. In the mutants, TWIST protein was present in the osteogenic mesenchyme, while osteopontin was not expressed until E15.5. In addition, in mutants, small regions of TWIST-positive osteogenic mesenchyme were visible until E15.5. The delay in ossification and reduction in size of the ossification centers were correlated with an earlier decrease in proliferation. We used micromass cultures of the face to investigate the direct effects of G9A inhibition on skeletal differentiation. Addition of a small molecule inhibitor for G9A, BIX-01294, to wild-type cells upregulated Twist genes similar to what was observed in vivo. The inhibitor also caused decreases in several osteogenic markers. Chromatin immunoprecipitation analysis of primary osteogenic mesenchyme from calvaria revealed that Twist1 and Twist2 regulatory regions contain the repressive H3K9me2 marks catalyzed by G9A, which are removed when BIX-01294 is added. Our results establish a role for G9A and H3K9me2 in the regulation of Twist genes and provide novel insights into the significance of epigenetic mechanisms in controlling temporal and tissue-specific gene

  2. Fabrication and tribological properties of self-assembled monolayer of n-alkyltrimethoxysilane on silicon: Effect of SAM alkyl chain length

    Energy Technology Data Exchange (ETDEWEB)

    Huo, Lixia [National Key Laboratory of Science and Technology on Vacuum Technology and Physics, Lanzhou Institute of Physics, Lanzhou, Gansu 730010 (China); Institute of Polymer Science and Engineering, College of Chemistry and Chemical Engineering, Lanzhou University, Gansu 730000 (China); Du, Pengcheng [Institute of Polymer Science and Engineering, College of Chemistry and Chemical Engineering, Lanzhou University, Gansu 730000 (China); Zhou, Hui; Zhang, Kaifeng [National Key Laboratory of Science and Technology on Vacuum Technology and Physics, Lanzhou Institute of Physics, Lanzhou, Gansu 730010 (China); Liu, Peng, E-mail: pliu@lzu.edu.cn [Institute of Polymer Science and Engineering, College of Chemistry and Chemical Engineering, Lanzhou University, Gansu 730000 (China)

    2017-02-28

    Highlights: • n-Alkyltrimethoxysilanes with various chain lengths were self-assembled on silicon. • Effect of alkyl chain lengths (C6, C12, or C18) on the SAMs was investigated. • Surface roughness of the SAMs decreased with increasing the alkyl chain lengths. • The C{sub 12}-SAM possessed superior friction reduction and wear resistance. - Abstract: It is well known that the self-assembled organic molecules on a solid surface exhibit the friction-reducing performance. However, the effect of the molecular size of the self-assembled organic molecules has not been established. In the present work, self-assembled monolayers (SAMs) of n-alkyltrimethoxysilanes with different alkyl chain lengths (C{sub 6}, C{sub 12}, or C{sub 18}) were fabricated on silicon substrate. The water contact angles of the SAMs increased from 26.8° of the hydroxylated silicon substrate to near 60° after self-assembly. The atomic force microscopy (AFM) analysis results showed that the mean roughness (R{sub a}) of the SAMs decreased with increasing the alkyl chain length. The tribological properties of the SAMs sliding against Al{sub 2}O{sub 3} ball were evaluated on an UMT-2 tribometer, and the worn surfaces of the samples were analyzed by means of Nano Scratch Tester and surface profilometry. It was found that lowest friction coefficient and smallest width of wear were achieved with the SAMs of C{sub 12} alkyl chain (C{sub 12}-SAM). The superior friction reduction and wear resistance of the SAMs in comparison with the bare silicon substrate are attributed to good adhesion of the self-assembled films to the substrate, especially the C{sub 12}-SAM with desirable alkyl chain length.

  3. Franchising as a Potential Growth Strategy for a Small Business : A Case of Sam-Chi Fast Food Restaurant

    OpenAIRE

    Odunsi, Sadiq

    2015-01-01

    The purpose of this study was to find out whether Sam-Chi fast food restaurant can grow through franchising as well as to give the owners recommendations on how to effectively adopt the franchising business model as a means to grow their business. Sam-Chi restaurant is situated in Lagos, Nigeria and the restaurant is owned and operated by Samuel Okore and his wife Chichi Okore. The theoretical framework of this research is separated into two sections. The first section covers the growth of a ...

  4. Hierarchical Storage Management at the NASA Center for Computational Sciences: From UniTree to SAM-QFS

    Science.gov (United States)

    Salmon, Ellen; Tarshish, Adina; Palm, Nancy; Patel, Sanjay; Saletta, Marty; Vanderlan, Ed; Rouch, Mike; Burns, Lisa; Duffy, Daniel; Caine, Robert

    2004-01-01

    This paper presents the data management issues associated with a large center like the NCCS and how these issues are addressed. More specifically, the focus of this paper is on the recent transition from a legacy UniTree (Legato) system to a SAM-QFS (Sun) system. Therefore, this paper will describe the motivations, from both a hardware and software perspective, for migrating from one system to another. Coupled with the migration from UniTree into SAM-QFS, the complete mass storage environment was upgraded to provide high availability, redundancy, and enhanced performance. This paper will describe the resulting solution and lessons learned throughout the migration process.

  5. CLONING, EXPRESSION, AND CHARACTERIZATION OF RAT S-ADENOSYL-L-METHIONINE: ARSENIC (III) METHYLTRANSFERASE (CYT19)

    Science.gov (United States)

    CLONING, EXPRESSION, AND CHARACTERIZATION OF RAT S-ADENOSYL-L-METHIONINE: ARSENIC(III) METHYLTRANSFERASE (cyt19)Stephen B. Waters1 , Felicia Walton1 , Miroslav Styblo1 , Karen Herbin-Davis2, and David J. Thomas2 1 School of Medicine, University of North Carolina at Chape...

  6. The genome-wide identification and transcriptional levels of DNA methyltransferases and demethylases in globe artichoke.

    Directory of Open Access Journals (Sweden)

    Silvia Gianoglio

    Full Text Available Changes to the cytosine methylation status of DNA, driven by the activity of C5 methyltransferases (C5-MTases and demethylases, exert an important influence over development, transposon movement, gene expression and imprinting. Three groups of C5-MTase enzymes have been identified in plants, namely MET (methyltransferase 1, CMT (chromomethyltransferases and DRM (domains rearranged methyltransferases. Here the repertoire of genes encoding C5-MTase and demethylase by the globe artichoke (Cynara cardunculus var. scolymus is described, based on sequence homology, a phylogenetic analysis and a characterization of their functional domains. A total of ten genes encoding C5-MTase (one MET, five CMTs and four DRMs and five demethylases was identified. An analysis of their predicted product's protein structure suggested an extensive level of conservation has been retained by the C5-MTases. Transcriptional profiling based on quantitative real time PCR revealed a number of differences between the genes encoding maintenance and de novo methyltransferases, sometimes in a tissue- or development-dependent manner, which implied a degree of functional specialization.

  7. The genome-wide identification and transcriptional levels of DNA methyltransferases and demethylases in globe artichoke.

    Science.gov (United States)

    Gianoglio, Silvia; Moglia, Andrea; Acquadro, Alberto; Comino, Cinzia; Portis, Ezio

    2017-01-01

    Changes to the cytosine methylation status of DNA, driven by the activity of C5 methyltransferases (C5-MTases) and demethylases, exert an important influence over development, transposon movement, gene expression and imprinting. Three groups of C5-MTase enzymes have been identified in plants, namely MET (methyltransferase 1), CMT (chromomethyltransferases) and DRM (domains rearranged methyltransferases). Here the repertoire of genes encoding C5-MTase and demethylase by the globe artichoke (Cynara cardunculus var. scolymus) is described, based on sequence homology, a phylogenetic analysis and a characterization of their functional domains. A total of ten genes encoding C5-MTase (one MET, five CMTs and four DRMs) and five demethylases was identified. An analysis of their predicted product's protein structure suggested an extensive level of conservation has been retained by the C5-MTases. Transcriptional profiling based on quantitative real time PCR revealed a number of differences between the genes encoding maintenance and de novo methyltransferases, sometimes in a tissue- or development-dependent manner, which implied a degree of functional specialization.

  8. Guanidinoacetate Methyltransferase (GAMT) Deficiency: Late Onset of Movement Disorder and Preserved Expressive Language

    Science.gov (United States)

    O'Rourke, Declan J.; Ryan, Stephanie; Salomons, Gajja; Jakobs, Cornelis; Monavari, Ahmad; King, Mary D.

    2009-01-01

    Guanidinoacetate methyltransferase (GAMT) deficiency is a disorder of creatine biosynthesis, characterized by early-onset learning disability and epilepsy in most affected children. Severe expressive language delay is a constant feature even in the mildest clinical phenotypes. We report the clinical, biochemical, imaging, and treatment data of two…

  9. The histone methyltransferase and putative oncoprotein MMSET is overexpressed in a large variety of human tumors

    DEFF Research Database (Denmark)

    Hudlebusch, Heidi Rye; Santoni-Rugiu, Eric; Simon, Ronald

    2011-01-01

    Multiple myeloma SET (Suppressor of variegation, Enhancer of zeste, and Trithorax) domain (MMSET) is a histone lysine methyltransferase deregulated in a subgroup of multiple myelomas with the t(4;14)(p16;q32) translocation and poor prognosis. With the aim of understanding, if MMSET can be involved...

  10. Guanidinoacetate methyltransferase (GAMT) deficiency : Outcomes in 48 individuals and recommendations for diagnosis, treatment and monitoring

    NARCIS (Netherlands)

    Stockler-Ipsiroglu, Sylvia; van Karnebeek, Clara; Longo, Nicola; Korenke, G. Christoph; Mercimek-Mahmutoglu, Saadet; Marquart, Iris; Barshop, Bruce; Grolik, Christiane; Schlune, Andrea; Angle, Brad; Araujo, Helena Caldeira; Coskun, Turgay; Diogo, Luisa; Geraghty, Michael; Haliloglu, Goknur; Konstantopoulou, Vassiliki; Leuzzi, Vincenzo; Levtova, Alina; MacKenzie, Jennifer; Maranda, Bruno; Mhanni, Aizeddin A.; Mitchell, Grant; Morris, Andrew; Newlove, Theresa; Renaud, Deborah; Scaglia, Fernando; Valayannopoulos, Vassili; van Spronsen, Francjan J.; Verbruggen, Krijn T.; Yuskiv, Nataliya; Nyhan, William; Schulze, Andreas

    We collected data on 48 patients from 38 families with guanidinoacetate methyltransferase (GAMT) deficiency. Global developmental delay/intellectual disability (DD/ID) with speech/language delay and behavioral problems as the most affected domains was present in 44 participants, with additional

  11. Control of substrate specificity by a single active site residue of the KsgA methyltransferase.

    Science.gov (United States)

    O'Farrell, Heather C; Musayev, Faik N; Scarsdale, J Neel; Rife, Jason P

    2012-01-10

    The KsgA methyltransferase is universally conserved and plays a key role in regulating ribosome biogenesis. KsgA has a complex reaction mechanism, transferring a total of four methyl groups onto two separate adenosine residues, A1518 and A1519, in the small subunit rRNA. This means that the active site pocket must accept both adenosine and N(6)-methyladenosine as substrates to catalyze formation of the final product N(6),N(6)-dimethyladenosine. KsgA is related to DNA adenosine methyltransferases, which transfer only a single methyl group to their target adenosine residue. We demonstrate that part of the discrimination between mono- and dimethyltransferase activity lies in a single residue in the active site, L114; this residue is part of a conserved motif, known as motif IV, which is common to a large group of S-adenosyl-L-methionine-dependent methyltransferases. Mutation of the leucine to a proline mimics the sequence found in DNA methyltransferases. The L114P mutant of KsgA shows diminished overall activity, and its ability to methylate the N(6)-methyladenosine intermediate to produce N(6),N(6)-dimethyladenosine is impaired; this is in contrast to a second active site mutation, N113A, which diminishes activity to a level comparable to L114P without affecting the methylation of N(6)-methyladenosine. We discuss the implications of this work for understanding the mechanism of KsgA's multiple catalytic steps.

  12. Catechol-O-methyltransferase gene methylation and substance use in adolescents: The TRAILS study

    NARCIS (Netherlands)

    L.J. van der Knaap (Lisette); J.M. Schäfer (Johanna); I.H.A. Franken (Ingmar); F.C. Verhulst (Frank); F.V.A. van Oort (Floor); H. Riese (Harriëtte)

    2014-01-01

    textabstractSubstance use often starts in adolescence and poses a major problem for society and individual health. The dopamine system plays a role in substance use, and catechol-O-methyltransferase (COMT) is an important enzyme that degrades dopamine. The Val108/158Met polymorphism

  13. Catechol-O-methyltransferase gene methylation and substance use in adolescents : the TRAILS study

    NARCIS (Netherlands)

    van der Knaap, L. J.; Schaefer, J. M.; Franken, I. H. A.; Verhulst, F. C.; van Oort, F. V. A.; Riese, H.

    Substance use often starts in adolescence and poses a major problem for society and individual health. The dopamine system plays a role in substance use, and catechol-O-methyltransferase (COMT) is an important enzyme that degrades dopamine. The Val(108/158)Met polymorphism modulates COMT activity

  14. Loss of the histone methyltransferase EZH2 induces resistance to multiple drugs in acute myeloid leukemia

    DEFF Research Database (Denmark)

    Göllner, Stefanie; Oellerich, Thomas; Agrawal-Singh, Shuchi

    2017-01-01

    In acute myeloid leukemia (AML), therapy resistance frequently occurs, leading to high mortality among patients. However, the mechanisms that render leukemic cells drug resistant remain largely undefined. Here, we identified loss of the histone methyltransferase EZH2 and subsequent reduction of h...

  15. Functional characterization of cinnamyl alcohol dehydrogenase and caffeic acid O-methyltransferase in Brachypodium distachyon.

    Science.gov (United States)

    Lignin is a significant recalcitrant in the conversion of plant biomass to bioethanol. Cinnamyl alcohol dehydrogenase (CAD) and caffeic acid O-methyltransferase (COMT) catalyze key steps in the pathway of lignin monomer biosynthesis. Brown midrib mutants in Zea mays and Sorghum bicolor with impaired...

  16. Polymorphisms in the catechol-o-methyltransferase gene and delirium in the elderly

    NARCIS (Netherlands)

    van Munster, Barbara C.; Baas, Frank; Tanck, Michael W.; de Rooij, Sophia E. J. A.

    2011-01-01

    Catechol-O-methyltransferase, encoded by the COMT gene, is one of the enzymes that degrade dopamine. The aim of this study was to investigate whether polymorphisms in the COMT gene were associated with delirium. Patients aged 65 years and older, acutely admitted to the medical department or to the

  17. Catechol-O-methyltransferase Val158Met and Cognitive Function in Parkinson's Disease

    NARCIS (Netherlands)

    Hoogland, Jeroen; de Bie, Rob M. A.; Williams-Gray, Caroline H.; Muslimović, Dino; Schmand, Ben; Post, Bart

    2010-01-01

    Cognitive dysfunction is one of the most incapacitating non-motor symptoms of Parkinson's disease (PD). Some cognitive deficits are thought to be related to abnormal dopamine homeostasis. The latter is influenced by catechol-O-methyltransferase (COMT), an enzyme that degrades dopamine. Previous

  18. Polymorphisms in catechol-O-methyltransferase and methylenetetrahydrofolate reductase in relation to the risk of schizophrenia.

    NARCIS (Netherlands)

    Muntjewerff, J.W.; Gellekink, H.; Heijer, M. den; Hoogendoorn, M.L.; Kahn, R.S.; Sinke, R.J.; Blom, H.J.

    2008-01-01

    BACKGROUND: Evidence is emerging for the association of aberrant homocysteine-methylation cycle and increased risk of schizophrenia. METHODS: We examined the prevalence of the catechol-O-methyltransferase (COMT) 324G>A (Val108/158Met) and methylenetetrahydrofolate reductase (MTHFR) 677C>T

  19. Association of Catechol-O-Methyltransferase (COMT) Polymorphism and Academic Achievement in a Chinese Cohort

    Science.gov (United States)

    Yeh, Ting-Kuang; Chang, Chun-Yen; Hu, Chung-Yi; Yeh, Ting-Chi; Lin, Ming-Yeh

    2009-01-01

    Catechol-O-methyltransferase (COMT) is a methylation enzyme that catalyzes the degradation pathway and inactivation of dopamine. It is accepted widely as being involved in the modulation of dopaminergic physiology and prefrontal cortex (PFC) function. The COMT Val158Met polymorphism is associated with variation in COMT activity. COMT 158Met allele…

  20. Catechol-O-methyltransferase val158met and cognitive function in Parkinson's disease

    NARCIS (Netherlands)

    Hoogland, J.; de Bie, R.M.A.; Williams-Gray, C.H.; Muslimovic, D.; Schmand, B.; Post, B.

    2010-01-01

    Cognitive dysfunction is one of the most incapacitating non-motor symptoms of Parkinson's disease (PD). Some cognitive deficits are thought to be related to abnormal dopamine homeostasis. The latter is influenced by catechol-O-methyltransferase (COMT), an enzyme that degrades dopamine. Previous

  1. Catechol-O-methyltransferase: a method for autoradiographic visualization of isozymes in cellogel

    Energy Technology Data Exchange (ETDEWEB)

    Brahe, C.; Crosti, N.; Meera Khan, P.; Serra, A.

    1984-02-01

    An electrophoretic procedure for separating the molecular forms of catechol-O-methyltransferase in cellulose acetate gel is described; the zones of enzyme activity were revealed by autoradiography. The electrophoretic patterns of the enzyme in several tissues and cell lines derived from four different species are presented.

  2. Catechol-O-methyltransferase Val158Met and the Risk of Dyskinesias in Parkinson's Disease

    NARCIS (Netherlands)

    De Lau, L.M.L.; Verbaan, D.; Marinus, J.; Heutink, P.; van Hilten, J.J.

    2012-01-01

    Background:: The A-allele of the catechol-O-methyltransferase (COMT) Val158Met polymorphism is associated with decreased enzymatic activity and higher dopamine availability. Methods:: We studied 219 patients with PD who were free of dyskinesias at baseline and underwent thorough annual examinations.

  3. Catechol-O-methyltransferase val158met and cognitive function in Parkinson's disease.

    NARCIS (Netherlands)

    Hoogland, J.; Bie, R.M. de; Williams-Gray, C.H.; Muslimovic, D.; Schmand, B.A.; Post, B.

    2010-01-01

    Cognitive dysfunction is one of the most incapacitating non-motor symptoms of Parkinson's disease (PD). Some cognitive deficits are thought to be related to abnormal dopamine homeostasis. The latter is influenced by catechol-O-methyltransferase (COMT), an enzyme that degrades dopamine. Previous

  4. Catechol-O-methyltransferase (COMT) gene variants and pain in chronic pancreatitis

    NARCIS (Netherlands)

    Esch, A.A.J.; Vries, E. De; Morsche, R.H.M. te; Oijen, M.G.H. van; Jansen, J.B.M.J.; Drenth, J.P.H.

    2011-01-01

    BACKGROUND: Pain is the major symptom of chronic pancreatitis. The role of genetics in pancreatic pain is unclear. Catechol-O-methyltransferase (COMT) regulates enkephalin levels and influences pain perception. The COMT gene contains functional polymorphisms that have been found to influence human

  5. Local chromatin microenvironment determines DNMT activity : from DNA methyltransferase to DNA demethylase or DNA dehydroxymethylase

    NARCIS (Netherlands)

    van der Wijst, Monique G. P.; Venkiteswaran, Muralidhar; Chen, Hui; Xu, Guo-Liang; Plosch, Torsten; Rots, Marianne G.

    2015-01-01

    Insights on active DNA demethylation disproved the original assumption that DNA methylation is a stable epigenetic modification. Interestingly, mammalian DNA methyltransferases 3A and 3B (DNMT-3A and -3B) have also been reported to induce active DNA demethylation, in addition to their well-known

  6. DNA methyltransferase and alcohol dehydrogenase: gene-nutrient interactions in relation to risk of colorectal polyps.

    NARCIS (Netherlands)

    Jung, A.Y.; Poole, E.M.; Bigler, J.; Whitton, J.; Potter, J.D.; Ulrich, C.M.

    2008-01-01

    Disturbances in DNA methylation are a characteristic of colorectal carcinogenesis. Folate-mediated one-carbon metabolism is essential for providing one-carbon groups for DNA methylation via DNA methyltransferases (DNMTs). Alcohol, a folate antagonist, could adversely affect one-carbon metabolism. In

  7. Recognition elements in rRNA for the tylosin resistance methyltransferase RlmA(II)

    DEFF Research Database (Denmark)

    Lebars, Isabelle; Husson, Clotilde; Yoshizawa, Satoko

    2007-01-01

    antibiotics. We have previously solved the solution structure of hairpin 35 in the conformation that is recognized by the RlmA(II) methyltransferase from Streptococcus pneumoniae. It was shown that while essential recognition elements are located in hairpin 35, the interactions between RlmA(II) and hairpin 35...

  8. Recognition of RNA cap in the Wesselsbron virus NS5 methyltransferase domain: implications for RNA-capping mechanisms in Flavivirus.

    Science.gov (United States)

    Bollati, Michela; Milani, Mario; Mastrangelo, Eloise; Ricagno, Stefano; Tedeschi, Gabriella; Nonnis, Simona; Decroly, Etienne; Selisko, Barbara; de Lamballerie, Xavier; Coutard, Bruno; Canard, Bruno; Bolognesi, Martino

    2009-01-09

    The mRNA-capping process starts with the conversion of a 5'-triphosphate end into a 5'-diphosphate by an RNA triphosphatase, followed by the addition of a guanosine monophosphate unit in a 5'-5' phosphodiester bond by a guanylyltransferase. Methyltransferases are involved in the third step of the process, transferring a methyl group from S-adenosyl-l-methionine to N7-guanine (cap 0) and to the ribose 2'OH group (cap 1) of the first RNA nucleotide; capping is essential for mRNA stability and proper replication. In the genus Flavivirus, N7-methyltransferase and 2'O-methyltransferase activities have been recently associated with the N-terminal domain of the viral NS5 protein. In order to further characterize the series of enzymatic reactions that support capping, we analyzed the crystal structures of Wesselsbron virus methyltransferase in complex with the S-adenosyl-l-methionine cofactor, S-adenosyl-l-homocysteine (the product of the methylation reaction), Sinefungin (a molecular analogue of the enzyme cofactor), and three different cap analogues (GpppG, (N7Me)GpppG, and (N7Me)GpppA). The structural results, together with those on other flaviviral methyltransferases, show that the capped RNA analogues all bind to an RNA high-affinity binding site. However, lack of specific interactions between the enzyme and the first nucleotide of the RNA chain suggests the requirement of a minimal number of nucleotides following the cap to strengthen protein/RNA interaction. Our data also show that, following incubation with guanosine triphosphate, Wesselsbron virus methyltransferase displays a guanosine monophosphate molecule covalently bound to residue Lys28, hinting at possible implications for the transfer of a guanine group to ppRNA. The structures of the Wesselsbron virus methyltransferase complexes obtained are discussed in the context of a model for N7-methyltransferase and 2'O-methyltransferase activities.

  9. Evolution of the Phosphatidylcholine Biosynthesis Pathways in Green Algae: Combinatorial Diversity of Methyltransferases.

    Science.gov (United States)

    Hirashima, Takashi; Toyoshima, Masakazu; Moriyama, Takashi; Sato, Naoki

    2018-01-12

    Phosphatidylcholine (PC) is one of the most common phospholipids in eukaryotes, although some green algae such as Chlamydomonas reinhardtii are known to lack PC. Recently, we detected PC in four species in the genus Chlamydomonas: C. applanata NIES-2202, C. asymmetrica NIES-2207, C. debaryana NIES-2212, and C. sphaeroides NIES-2242. To reveal the PC biosynthesis pathways in green algae and the evolutionary scenario involved in their diversity, we analyzed the PC biosynthesis genes in these four algae using draft genome sequences. Homology searches suggested that PC in these species is synthesized by phosphoethanolamine-N-methyltransferase (PEAMT) and/or phosphatidylethanolamine-N-methyltransferase (PEMT), both of which are absent in C. reinhardtii. Recombinant PEAMTs from these algae showed methyltransferase activity for phosphoethanolamine but not for monomethyl phosphoethanolamine in vitro, in contrast to land plant PEAMT, which catalyzes the three methylations from phosphoethanolamine to phosphocholine. This suggested an involvement of other methyltransferases in PC biosynthesis. Here, we characterized the putative phospholipid-N-methyltransferase (PLMT) genes of these species by genetic and phylogenetic analysis. Complementation assays using a PC biosynthesis-deficient yeast suggested that the PLMTs of these algae can synthesize PC from phosphatidylethanolamine. These results indicated that the PC biosynthesis pathways in green algae differ from those of land plants, although the enzymes involved are homologous. Phylogenetic analysis suggested that the PEAMTs and PLMTs in these algae were inherited from the common ancestor of green algae. The absence of PC biosynthesis in many Chlamydomonas species is likely a result of parallel losses of PEAMT and PLMT in this genus.

  10. NaCl stress induces CsSAMs gene expression in Cucumis sativus by mediating the binding of CsGT-3b to the GT-1 element within the CsSAMs promoter.

    Science.gov (United States)

    Wang, Li-Wei; He, Mei-Wen; Guo, Shi-Rong; Zhong, Min; Shu, Sheng; Sun, Jin

    2017-05-01

    The CsSAMs promoter is a salt-stress-inducible promoter containing three GT-1 elements that are sufficient for the salt-stress response. The transcription factor CsGT-3b was found to bind to the GT-1 element. The S-adenosyl-L-methionine synthase (SAMs) gene is among the functional genes induced during environmental stress. However, little is known about the regulatory mechanism and upstream regulators of this salt-inducible gene in cucumber plants. Thus, it is necessary to understand the characteristics of the SAMs gene by analyzing its promoter and transcription factors. In this study, we isolated and functionally analyzed a 1743-bp flanking fragment of the CsSAMs gene from Cucumis sativus. To examine promoter activity, the full-length promoter, as well as different promoter fragments, were fused to the β-glucuronidase (GUS) reporter gene and introduced into the tobacco genome. The full-length promoter displayed maximal promoter activity, whereas the P4 promoter, containing 321 bp of upstream sequence, showed no basal promoter activity. In addition, the CsSAMs promoter exhibited stress-inducible regulation rather than tissue-specific activity in transgenic tobacco. Histochemical analysis revealed strong GUS staining in leaves, stems, and roots, especially in the veins of leaves, the vascular bundle of stems, and root tip zones following NaCl stress. A transient expression assay confirmed that the 242-bp region (-1743 to -1500) was sufficient for the NaCl-stress response. Yeast one-hybrid assays further revealed interaction between the NaCl-response protein CsGT-3b and the GT-1 (GAAAAA) element within the 242-bp region. Taken together, we revealed the presence of four salt-stress-responsive elements (GT-1 cis-elements) in the CsSAMs promoter and identified a transcription factor, CsGT-3b, that specifically binds to this sequence. These results might help us better understand the intricate regulatory network of the cucumber SAMs gene.

  11. Protein Arginine Methyltransferase 5 as a Driver of Lymphomagenesis

    Science.gov (United States)

    Smith, Porsha Latrice

    Over the past decade, it has become clear that oncogenesis is a process driven by a wide variety of triggers including gene mutations, gene amplifications, inflammation, and immune deficiency. The growing pool of data collected from whole genome and epigenome studies of both solid and blood cancers has pointed toward dysregulation of chromatin remodelers as a unique class of cancer drivers. Next generation sequencing studies of lymphomas have identified a wide array of somatic mutations affecting enzymes that regulate epigenetic control of gene expression. Lymphoma is a type of cancer that originates in secondary lymphoid organs and manifests as an outgrowth of transformed lymphocytes, or white blood cells (WBCs) in the blood. The majority of lymphoma cases can be grouped into the Non-Hodgkins lymphoma (NHL) subset and mainly occurs in B-cells. B-cell NHL is a heterogeneous set of cancers that would benefit from new therapies to improve patient progression-free survival. Cancers such as NHL typically present with a combination of genetic and epigenetic aberrations that contribute to the malignancy program. The epigenetic modifier protein arginine methyltransferase 5 (PRMT5) is required for B-cell transformation following Epstein-Barr virus (EBV) infection, and is overexpressed in various subsets of B-cell NHL. Based on these data we hypothesized that PRMT5 is a major driver of B-cell lymphomagenesis. To explore the role of PRMT5 in the development and progression of B-cell NHL we created a small molecule inhibitors targeted to PRMT5. Using the NHL subset mantle cell lymphoma (MCL) as a model we tested the efficacy of the drug. We discovered that PRMT5 was overexpressed in MCL primary samples and cell lines as compared to normal resting B cells. Furthermore, use of the small molecule inhibitor decreased the proliferation and viability in these cells without affecting the normal B-cells. Additionally, use of inhibitors caused G2/M cell cycle and decreased the

  12. The Investigation of Identity Construction: A Foucauldian Reading of Sam Shepard's Buried Child

    Directory of Open Access Journals (Sweden)

    Saeidabadi S.

    2016-01-01

    Full Text Available Shepard is peculiarly powerful in his symbolic family problem plays: True West, Buried Child and Curse of the Starving Class. He allegorizes the American experience and undermines the myth of America as the New Eden. The present study seeks to critically explore Sam Shepard's Buried Child in terms of Foucauldian conception of identity construction. Shepard is depicting a dystopian world with its bewildered characters; however he has still got a romantic view of individuals trying to grapple with the society in order to get unity and order. This Shephardian attitude towards human beings is seemingly a free agent that overlaps the Foucauldian view which establishes a philosophy focusing on the relationship between the self and the society. The present essay attempts to demonstrate the complicated relationship between the self and the opposing forces.

  13. SAGE I and SAM II measurements of 1 micron aerosol extinction in the free troposphere

    Science.gov (United States)

    Kent, G. S.; Farrukh, U. O.; Wang, P. H.; Deepak, A.

    1988-01-01

    The SAGE-I and SAM-II satellite sensors were designed to measure, with global coverage, the 1 micron extinction produced by the stratospheric aerosol. In the absence of high altitude clouds, similar measurements may be made for the free tropospheric aerosol. Median extinction values at middle and high latitudes in the Northern Hemisphere, for altitudes between 5 and 10 km, are found to be one-half to one order of magnitude greater than values at corresponding latitudes in the Southern Hemisphere. In addition, a seasonal increase by a factor of 1.5-2 was observed in both hemispheres, in 1979-80, in local spring and summer. Following major volcanic eruptions, a long-lived enhancement of the aerosol extinction is observed for altitudes above 5 km.

  14. SAGE 1 and SAM 2 measurements of 1 micron aerosol extinction in the free troposphere

    Science.gov (United States)

    Kent, G. S.; Farrukh, U. O.; Wang, P. H.; Deepak, A.

    1988-01-01

    The SAGE 1 and SAM 2 satellite sensors were designed to measure, with global coverage, the 1 micron extinction produced by the stratospheric aerosol. In the absence of high altitude cloud, similar measurements may be made for the free tropospheric aerosol. Median extinction values in the Northern Hemisphere, for altitudes between 5 and 10 km, are found to be one-half to one order of magnitude greater than values at corresponding latitudes in the Southern Hemisphere. In addition, a seasonal increase by a factor of 1.5 yields 2 is observed in both hemispheres in local spring and summer. Following major volcanic eruptions, a long-lived enhancement of the aerosol extinction is observed for altitudes above 5 km.

  15. Mechanistic Investigations of Lysine-Tryptophan Crosslink Formation Catalyzed by Streptococcal Radical SAM Enzymes.

    Science.gov (United States)

    Schramma, Kelsey R; Forneris, Clarissa C; Caruso, Alessio; Seyedsayamdost, Mohammad R

    2018-01-10

    Streptide is a ribosomally synthesized and post-translationally modified peptide with a unique cyclization motif consisting of an intramolecular lysine-tryptophan crosslink. Three radical SAM enzymes, StrB, AgaB, and SuiB from different Streptococci, have been shown to install this modification onto their respective precursor peptides in a leader-dependent fashion. Herein we conduct detailed investigations to differentiate between several plausible mechanistic proposals, specifically addressing radical versus electrophilic addition to the indole during crosslink formation, the role of substrate side-chains in binding in the enzyme active site, and the identity of the catalytic base in the reaction cycle. Our results are consistent with a radical electrophilic aromatic substitution mechanism for the key carbon-carbon bond-forming step. They also elaborate on other mechanistic features that underpin this unique and synthetically challenging post-translational modification.

  16. Sam Thompson, Stewart Parker, and the lineage of northern Irish drama

    Directory of Open Access Journals (Sweden)

    Marilynn Richtarik

    2010-11-01

    Full Text Available Northern Irish dramatist Stewart Parker (1941-1988 wrote plays typically valued for their wit, intellectual content, and formal experimentation. Nonetheless, he was profoundly influenced as a young man by a very different sort of playwright. Sam Thompson (1916-1965, who began his working life in the Belfast shipyards, squarely confronted Northern Irish sectarianism in his plays. His sense of the political potential of drama left an enduring mark on Parker, who organized and edited Thompson's manuscripts several years after his untimely death. Although their dramatic writings bear little resemblance to each other, the two writers should be regarded as united in a common Northern Irish dramatic tradition by virtue of their shared socialist outlook, belief in the importance of individual stands against conformity, and sense of theatre's social mission.

  17. Charge dependent asphaltene adsorption onto metal substrate : electrochemistry and AFM, STM, SAM, SEM analysis

    Energy Technology Data Exchange (ETDEWEB)

    Batina, N.; Morales-Martinez, J. [Univ. Autonoma Metropolitana-Iztapalapa (Mexico). Lab. de Nanotecnologia e Ingenieria Molecular; Ivar-Andersen, S. [Technical Univ. of Denmark (Denmark). Dept. Hem. Eng; Lira-Galeana, C. [Inst. Mexicano del Petroleo, Lazaro (Mexico). Molecular Simulation Research Program; De la Cruz-Hernandez, W.; Cota-Araiza, L.; Avalos-Borja, M. [Univ. Nacional Autonoma de Mexico (Mexico)

    2008-07-01

    Asphaltenes have been identified as the main component of pipeline molecular deposits that cause plugging of oil wells. In this study, Atomic Force Microscopy (AFM), Scanning Tunneling Microscopy (STM), Scanning Auger Microprobe Spectroscopy (SAM) and Scanning Electron Microscopy (SEM) were used to characterized molecular deposits of Mexican crude oil and asphaltenes formed at a charged metal surface. The qualitative and quantitative characterization involved determining the size and shape of adsorbed molecules and aggregates, and the elemental analysis of all components in molecular films. Samples were prepared by electrolytic deposition under galvanostatic or potentiostatic conditions directly from the crude oil or asphaltene in toluene solutions. The study showed that the formation of asphaltene deposit depends on the metal substrate charge. Asphaltenes as well as crude oil readily adsorbed at the negatively charged metal surface. Two elements were present, notably carbon and sulfur. Their content ratio varied depending on the metal substrate charge.

  18. Structural investigation of a C-terminal EphA2 receptor mutant: Does mutation affect the structure and interaction properties of the Sam domain?

    Science.gov (United States)

    Mercurio, Flavia A; Costantini, Susan; Di Natale, Concetta; Pirone, Luciano; Guariniello, Stefano; Scognamiglio, Pasqualina L; Marasco, Daniela; Pedone, Emilia M; Leone, Marilisa

    2017-09-01

    Ephrin A2 receptor (EphA2) plays a key role in cancer, it is up-regulated in several types of tumors and the process of ligand-induced receptor endocytosis, followed by degradation, is considered as a potential path to diminish tumor malignancy. Protein modulators of this mechanism are recruited at the cytosolic Sterile alpha motif (Sam) domain of EphA2 (EphA2-Sam) through heterotypic Sam-Sam associations. These interactions engage the C-terminal helix of EphA2 and close loop regions (the so called End Helix side). In addition, several studies report on destabilizing mutations in EphA2 related to cataract formation and located in/or close to the Sam domain. Herein, we analyzed from a structural point of view, one of these mutants characterized by the insertion of a novel 39 residue long polypeptide at the C-terminus of EphA2-Sam. A 3D structural model was built by computational methods and revealed partial disorder in the acquired C-terminal tail and a few residues participating in an α-helix and two short β-strands. We investigated by CD and NMR studies the conformational properties in solution of two peptides encompassing the whole C-terminal tail and its predicted helical region, respectively. NMR binding experiments demonstrated that these peptides do not interact relevantly with either EphA2-Sam or its interactor Ship2-Sam. Molecular dynamics (MD) simulations further indicated that the EphA2 mutant could be represented only through a conformational ensemble and that the C-terminal tail should not largely wrap the EphA2-Sam End-Helix interface and affect binding to other Sam domains. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Boron (B) deprivation increases plasma homocysteine and decreases liver S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH) in rats

    Science.gov (United States)

    The diverse effects of B deprivation suggest that B affects a biomolecule involved in a variety of biochemical reactions. An experiment was conducted to determine whether dietary B affects the liver concentration of SAM, a frequently used enzyme substrate, especially for methylation reactions that y...

  20. Reactions Involving Calcium and Magnesium Sulfates as Potential Sources of Sulfur Dioxide During MSL SAM Evolved Gas Analyses

    Science.gov (United States)

    McAdam, A. C.; Knudson, C. A.; Sutter, B.; Franz, H. B.; Archer, P. D., Jr.; Eigenbrode, J. L.; Ming, D. W.; Morris, R. V.; Hurowitz, J. A.; Mahaffy, P. R.; hide

    2016-01-01

    The Sample Analysis at Mars (SAM) and Chemistry and Mineralogy (CheMin) instruments on the Mars Science Laboratory (MSL) have analyzed several subsamples of 860 C). Sulfides or Fe sulfates were detected by CheMin (e.g., CB, MJ, BK) and could contribute to the high temperature SO2 evolution, but in most cases they are not present in enough abundance to account for all of the SO2. This additional SO2 could be largely associated with x-ray amorphous material, which comprises a significant portion of all samples. It can also be attributed to trace S phases present below the CheMin detection limit, or to reactions which lower the temperatures of SO2 evolution from sulfates that are typically expected to thermally decompose at temperatures outside the SAM temperature range (e.g., Ca and Mg sulfates). Here we discuss the results of SAM-like laboratory analyses targeted at understanding this last possibility, focused on understanding if reactions of HCl or an HCl evolving phase (oxychlorine phases, chlorides, etc.) and Ca and Mg sulfates can result in SO2 evolution in the SAM temperature range.

  1. An Evaluation and Development Environment for an ARM7-Based Autopilot Using the Atmel SAM7S256 Microcontroller

    Science.gov (United States)

    2009-08-01

    size. These microcontrollers provide more computational ability than smaller Microchip peripheral interface controllers or AVR microcontrollers , while...An Evaluation and Development Environment for an ARM7-Based Autopilot Using the Atmel SAM7S256 Microcontroller by Justin L. Shumaker and... Microcontroller Justin L. Shumaker Vehicle Technology Directorate, ARL Ainsmar X. Brown National Institute of Aerospace

  2. Vejsalg, gårdbutikker og fritidsfiskere - den informelle økonomi på Samsø

    DEFF Research Database (Denmark)

    Rasmussen, Rasmus Ole

    2007-01-01

    Bidraget behandler den uformelle økonomi på Samsø, med udgangspunkt i den første kortlægning nogensinde af den del af den uformelle økonomi som er forbundet med salg af produkter som er relaterede til naturgrundlaget. De mange vejboder er et iøjnefaldende element i landskabet, og nogle af dem har...

  3. Response of Antarctic ice shelf melt to SAM trend and possible feedbacks with the ice-dynamics

    Science.gov (United States)

    Donat-Magnin, Marion; Jourdain, Nicolas C.; Gallée, Hubert; Spence, Paul; Cornford, Stephen L.; Le Sommer, Julien; Durand, Gaël

    2017-04-01

    The observed positive trend in the Southern Annular Mode (SAM) may warm the Southern Ocean sub-surface through decreased Ekman downward pumping. Subsequent change in ice-shelves melt has been suggested to trigger glacier acceleration in West Antarctica. Here we use a regional ocean model configuration of the Amundsen Sea that includes interactive ice-shelf cavities. Our results show that the inclusion of ice-shelves changes the ocean response to the projected SAM trend, i.e. it typically inhibits a part of the SAM-induced subsurface warming. Heat budget analysis has been used to propose responsible mechanisms. Regarding Thwaites and Pine Island, sub ice-shelf melt increases above 400m by approximately 40% for Thwaites and 10% for Pine Island and decreases by up to 10% below in response to ocean temperature changes driven by the projected SAM trend. The melt sensitivity to poleward shifting winds is nonetheless small compared to the sensitivity to an ice-sheet instability, i.e. to a projected change in the shape of ice-shelf cavities. For instance, the sub ice-shelf melt are doubled near the grounding line of some glaciers in response to the largest grounding line retreat projected for 2100. Large increase in basal melt close to the grounding line could largely impact instability and glacier acceleration. Our work suggests the need for including ice shelves into ocean models, and to couple ocean models to ice-sheet models in climate projections.

  4. Carbon and Sulfur Isotopic Composition of Yellowknife Bay Sediments: Measurements by the Sample Analysis at Mars (SAM) Quadrupole Mass Spectrometer

    Science.gov (United States)

    Franz, H. B.; Mahaffy, P. R.; Stern, J. C.; Eigenbrode, J. L.; Steele, A.; Ming, D. W.; McAdam, A. C.; Freissinet, C.; Glavin, D. P.; Archer, P. D.; hide

    2014-01-01

    Since landing at Gale Crater in Au-gust 2012, the Sample Analysis at Mars (SAM) instru-ment suite on the Mars Science Laboratory (MSL) “Curiosity” rover has analyzed solid samples from the martian regolith in three locations, beginning with a scoop of aeolian deposits from the Rocknest (RN) sand shadow. Curiosity subsequently traveled to Yellowknife Bay, where SAM analyzed samples from two separate holes drilled into the Sheepbed Mudstone, designated John Klein (JK) and Cumberland (CB). Evolved gas analysis (EGA) of all samples revealed the presence of H2O as well as O-, C- and S-bearing phas-es, in most cases at abundances below the detection limit of the CheMin instrument. In the absence of definitive mineralogical identification by CheMin, SAM EGA data can help provide clues to the mineralogy of volatile-bearing phases through examination of tem-peratures at which gases are evolved from solid sam-ples. In addition, the isotopic composition of these gas-es may be used to identify possible formation scenarios and relationships between phases. Here we report C and S isotope ratios for CO2 and SO2 evolved from the JK and CB mudstone samples as measured with SAM’s quadrupole mass spectrometer (QMS) and draw com-parisons to RN.

  5. Optimisation and Characterisation of Anti-Fouling Ternary SAM Layers for Impedance-Based Aptasensors

    Directory of Open Access Journals (Sweden)

    Anna Miodek

    2015-09-01

    Full Text Available An aptasensor with enhanced anti-fouling properties has been developed. As a case study, the aptasensor was designed with specificity for human thrombin. The sensing platform was developed on screen printed electrodes and is composed of a self-assembled monolayer made from a ternary mixture of 15-base thiolated DNA aptamers specific for human thrombin co-immobilised with 1,6-hexanedithiol (HDT and further passivated with 1-mercapto-6-hexanol (MCH. HDT binds to the surface by two of its thiol groups forming alkyl chain bridges and this architecture protects from non-specific attachment of molecules to the electrode surface. Using Electrochemical Impedance Spectroscopy (EIS, the aptasensor is able to detect human thrombin as variations in charge transfer resistance (Rct upon protein binding. After exposure to a high concentration of non-specific Bovine Serum Albumin (BSA solution, no changes in the Rct value were observed, highlighting the bio-fouling resistance of the surface generated. In this paper, we present the optimisation and characterisation of the aptasensor based on the ternary self-assembled monolayer (SAM layer. We show that anti-fouling properties depend on the type of gold surface used for biosensor construction, which was also confirmed by contact angle measurements. We further studied the ratio between aptamers and HDT, which can determine the specificity and selectivity of the sensing layer. We also report the influence of buffer pH and temperature used for incubation of electrodes with proteins on detection and anti-fouling properties. Finally, the stability of the aptasensor was studied by storage of modified electrodes for up to 28 days in different buffers and atmospheric conditions. Aptasensors based on ternary SAM layers are highly promising for clinical applications for detection of a range of proteins in real biological samples.

  6. Mouse arsenic (+3 oxidation state) methyltransferase genotype affects metabolism and tissue dosimetry of arsenicals after arsenite administration in drinking water

    Science.gov (United States)

    Arsenic (+3 oxidation state) methyltransferase (As3mt) catalyzes methylation of inorganic arsenic producing a number of methylated arsenic metabolites. Although methylation has been commonly considered a pathway for detoxification of arsenic, some highly reactive methylated ars...

  7. A conformational switch in the active site of BT_2972, a methyltransferase from an antibiotic resistant pathogen B. thetaiotaomicron.

    Directory of Open Access Journals (Sweden)

    Veerendra Kumar

    Full Text Available Methylation is one of the most common biochemical reactions involved in cellular and metabolic functions and is catalysed by the action of methyltransferases. Bacteroides thetaiotaomicron is an antibiotic-resistant bacterium that confers resistance through methylation, and as yet, there is no report on the structure of methyltransferases from this bacterium. Here, we report the crystal structure of an AdoMet-dependent methyltransferase, BT_2972 and its complex with AdoMet and AdoHcy for B. thetaiotaomicron VPI-5482 strain along with isothermal titration calorimetric assessment of the binding affinities. Comparison of the apo and complexed BT_2972 structures reveals a significant conformational change between open and closed forms of the active site that presumably regulates the association with cofactors and may aid interaction with substrate. Together, our analysis suggests that BT_2972 is a small molecule methyltransferase and might catalyze two O-methylation reaction steps involved in the ubiquinone biosynthesis pathway.

  8. The expression platform and the aptamer: cooperativity between Mg2+ and ligand in the SAM-I riboswitch

    Science.gov (United States)

    Hennelly, Scott P.; Novikova, Irina V.; Sanbonmatsu, Karissa Y.

    2013-01-01

    Riboswitch operation involves the complex interplay between the aptamer domain and the expression platform. During transcription, these two domains compete against each other for shared sequence. In this study, we explore the cooperative effects of ligand binding and Magnesium interactions in the SAM-I riboswitch in the context of aptamer collapse and anti-terminator formation. Overall, our studies show the apo-aptamer acts as (i) a pre-organized aptamer competent to bind ligand and undergo structural collapse and (ii) a conformation that is more accessible to anti-terminator formation. We show that both Mg2+ ions and SAM are required for a collapse transition to occur. We then use competition between the aptamer and expression platform for shared sequence to characterize the stability of the collapsed aptamer. We find that SAM and Mg2+ interactions in the aptamer are highly cooperative in maintaining switch polarity (i.e. aptamer ‘off-state’ versus anti-terminator ‘on-state’). We further show that the aptamer off-state is preferentially stabilized by Mg2+ and similar divalent ions. Furthermore, the functional switching assay was used to select for phosphorothioate interference, and identifies potential magnesium chelation sites while characterizing their coordinated role with SAM in aptamer stabilization. In addition, we find that Mg2+ interactions with the apo-aptamer are required for the full formation of the anti-terminator structure, and that higher concentrations of Mg2+ (>4 mM) shift the equilibrium toward the anti-terminator on-state even in the presence of SAM. PMID:23258703

  9. Cloning, sequencing, and expression of the uroporphyrinogen III methyltransferase cobA gene of Propionibacterium freudenreichii (shermanii).

    OpenAIRE

    Sattler, I; Roessner, C A; Stolowich, N J; Hardin, S H; Harris-Haller, L W; Yokubaitis, N T; Murooka, Y; Hashimoto, Y; Scott, A I

    1995-01-01

    We cloned, sequenced, and overexpressed cobA, the gene encoding uroporphyrinogen III methyltransferase in Propionibacterium freudenreichii, and examined the catalytic properties of the enzyme. The methyltransferase is similar in mass (27 kDa) and homologous to the one isolated from Pseudomonas denitrificans. In contrast to the much larger isoenzyme encoded by the cysG gene of Escherichia coli (52 kDa), the P. freudenreichii enzyme does not contain the additional 22-kDa peptide moiety at its N...

  10. Vibrational Stark Effect of the Electric-Field Reporter 4-Mercaptobenzonitrile as a Tool for Investigating Electrostatics at Electrode/SAM/Solution Interfaces

    Directory of Open Access Journals (Sweden)

    Peter Hildebrandt

    2012-06-01

    Full Text Available 4-mercaptobenzonitrile (MBN in self-assembled monolayers (SAMs on Au and Ag electrodes was studied by surface enhanced infrared absorption and Raman spectroscopy, to correlate the nitrile stretching frequency with the local electric field exploiting the vibrational Stark effect (VSE. Using MBN SAMs in different metal/SAM interfaces, we sorted out the main factors controlling the nitrile stretching frequency, which comprise, in addition to external electric fields, the metal-MBN bond, the surface potential, and hydrogen bond interactions. On the basis of the linear relationships between the nitrile stretching and the electrode potential, an electrostatic description of the interfacial potential distribution is presented that allows for determining the electric field strengths on the SAM surface, as well as the effective potential of zero-charge of the SAM-coated metal. Comparing this latter quantity with calculated values derived from literature data, we note a very good agreement for Au/MBN but distinct deviations for Ag/MBN which may reflect either the approximations and simplifications of the model or the uncertainty in reported structural parameters for Ag/MBN. The present electrostatic model consistently explains the electric field strengths for MBN SAMs on Ag and Au as well as for thiophenol and mercaptohexanoic acid SAMs with MBN incorporated as a VSE reporter.

  11. Crystal Structure of Bicc1 SAM Polymer and Mapping of Interactions between the Ciliopathy-Associated Proteins Bicc1, ANKS3, and ANKS6.

    Science.gov (United States)

    Rothé, Benjamin; Leettola, Catherine N; Leal-Esteban, Lucia; Cascio, Duilio; Fortier, Simon; Isenschmid, Manuela; Bowie, James U; Constam, Daniel B

    2017-12-26

    Head-to-tail polymers of sterile alpha motifs (SAM) can scaffold large macromolecular complexes. Several SAM-domain proteins that bind each other are mutated in patients with cystic kidneys or laterality defects, including the Ankyrin (ANK) and SAM domain-containing proteins ANKS6 and ANKS3, and the RNA-binding protein Bicc1. To address how their interactions are regulated, we first determined a high-resolution crystal structure of a Bicc1-SAM polymer, revealing a canonical SAM polymer with a high degree of flexibility in the subunit interface orientations. We further mapped interactions between full-length and distinct domains of Bicc1, ANKS3, and ANKS6. Neither ANKS3 nor ANKS6 alone formed macroscopic homopolymers in vivo. However, ANKS3 recruited ANKS6 to Bicc1, and the three proteins together cooperatively generated giant macromolecular complexes. Thus, the giant assemblies are shaped by SAM domains, their flanking sequences, and SAM-independent protein-protein and protein-mRNA interactions. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Biochemical Studies of Mycobacterial Fatty Acid Methyltransferase: A Catalyst for the Enzymatic Production of Biodiesel.

    Science.gov (United States)

    Petronikolou, Nektaria; Nair, Satish K

    2015-11-19

    Transesterification of fatty acids yields the essential component of biodiesel, but current processes are cost-prohibitive and generate waste. Recent efforts make use of biocatalysts that are effective in diverting products from primary metabolism to yield fatty acid methyl esters in bacteria. These biotransformations require the fatty acid O-methyltransferase (FAMT) from Mycobacterium marinum (MmFAMT). Although this activity was first reported in the literature in 1970, the FAMTs have yet to be biochemically characterized. Here, we describe several crystal structures of MmFAMT, which highlight an unexpected structural conservation with methyltransferases that are involved in plant natural product metabolism. The determinants for ligand recognition are analyzed by kinetic analysis of structure-based active-site variants. These studies reveal how an architectural fold employed in plant natural product biosynthesis is used in bacterial fatty acid O-methylation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  13. EmtA, a rRNA methyltransferase conferring high-level evernimicin resistance

    DEFF Research Database (Denmark)

    Mann, P. A.; Xiong, L.; Mankin, A. S.

    2001-01-01

    Enterococcus faecium strain 9631355 was isolated from animal sources on the basis of its resistance to the growth promotant avilamycin. The strain also exhibited high-level resistance to evernimicin, a drug undergoing evaluation as a therapeutic agent in humans. Ribosomes from strain 9631355...... exhibited a dramatic reduction in evernimicin binding, shown by both cell-free translation assays and direct-binding assays. The resistance determinant was cloned from strain 9631355; sequence alignments suggested it was a methyltransferase and therefore it was designated emtA for evernimicin...... methyltransferase. Evernimicin resistance was transmissible and emtA was localized to a plasmid-borne insertion element. Purified EmtA methylated 50S subunits from an evernimicin-sensitive strain 30-fold more efficiently than those from a resistant strain. Reverse transcription identified a pause site...

  14. Super-resolution optical DNA Mapping via DNA methyltransferase-directed click chemistry

    DEFF Research Database (Denmark)

    Vranken, Charlotte; Deen, Jochem; Dirix, Lieve

    2014-01-01

    We demonstrate an approach to optical DNA mapping, which enables near single-molecule characterization of whole bacteriophage genomes. Our approach uses a DNA methyltransferase enzyme to target labelling to specific sites and copper-catalysed azide-alkyne cycloaddition to couple a fluorophore...... to the DNA. We achieve a labelling efficiency of ∼70% with an average labelling density approaching one site every 500 bp. Such labelling density bridges the gap between the output of a typical DNA sequencing experiment and the long-range information derived from traditional optical DNA mapping. We lay...... the foundations for a wider-scale adoption of DNA mapping by screening 11 methyltransferases for their ability to direct sequence-specific DNA transalkylation; the first step of the DNA labelling process and by optimizing reaction conditions for fluorophore coupling via a click reaction. Three of 11 enzymes...

  15. Crystallization and preliminary X-ray diffraction studies of a catechol-O-methyltransferase/inhibitor complex

    Energy Technology Data Exchange (ETDEWEB)

    Rodrigues, M. L. [Instituto de Tecnologia Química e Biológica (ITQB), Universidade Nova de Lisboa, Av. República, Apt. 127, 2781-901 Oeiras (Portugal); Bonifácio, M. J.; Soares-da-Silva, P. [Department of Research and Development, BIAL, 4785 S. Mamede do Coronado (Portugal); Carrondo, M. A.; Archer, M., E-mail: archer@itqb.unl.pt [Instituto de Tecnologia Química e Biológica (ITQB), Universidade Nova de Lisboa, Av. República, Apt. 127, 2781-901 Oeiras (Portugal)

    2005-01-01

    Catechol-O-methyltransferase has been co-crystallized with a novel inhibitor, which has potential therapeutic application in the Parkinson’s disease therapy. Inhibitors of the enzyme catechol-O-methyltransferase (COMT) are used as co-adjuvants in the therapy of Parkinson’s disease. A recombinant form of the soluble cytosolic COMT from rat has been co-crystallized with a new potent inhibitor, BIA 8-176 [(3,4-dihydroxy-2-nitrophenyl)phenylmethanone], by the vapour-diffusion method using PEG 6K as precipitant. Crystals diffract to 1.6 Å resolution on a synchrotron-radiation source and belong to the monoclinic space group P2{sub 1}, with unit-cell parameters a = 52.77, b = 79.63, c = 61.54 Å, β = 91.14°.

  16. Analysis of DNA methyltransferase 3A gene mutations in patients with Philadelphia-negative myeloproliferative neoplasms

    OpenAIRE

    Neda Ketabchi; Mostafa Paridar; Javad Mohammadi-Asl; Alireza Abooali; Maria Kavianpour; Najmaldin Saki

    2017-01-01

    Context: Philadelphia (Ph)-negative myeloproliferative neoplasms (MPNs), including essential thrombocythemia (ET), polycythemia vera (PV), and primary myelofibrosis (PMF) from a group of disorders characterized by dysregulated JAK-STAT functionality, abnormal hematopoiesis, as well as increased production of proliferative cytokines. In addition to JAK2V617F mutation, additional gene alterations that are involved in epigenetic mechanisms, particularly de novo DNA methyltransferase 3A (DNMT3A),...

  17. Identification and Biochemical Characterization of Four Wood-Associated Glucuronoxylan Methyltransferases in Populus

    Science.gov (United States)

    Yuan, Youxi; Teng, Quincy; Zhong, Ruiqin; Ye, Zheng-Hua

    2014-01-01

    Wood is one of the promising bioenergy feedstocks for lignocellulosic biofuel production. Understanding how wood components are synthesized will help us design strategies for better utilization of wood for biofuel production. One of the major wood components is xylan, in which about 10% of xylosyl residues are substituted with glucuronic acid (GlcA) side chains. All the GlcA side chains of xylan in wood of Populus trichocarpa are methylated, which is different from Arabidopsis xylan in which about 60% of GlcA side chains are methylated. Genes responsible for methylation of GlcA side chains in Populus xylan have not been identified. Here, we report genetic and biochemical analyses of four DUF579 domain-containing proteins, PtrGXM1, PtrGXM2, PtrGXM3 and PtrGXM4, from Populus trichocarpa and their roles in GlcA methylation in xylan. The PtrGXM genes were found to be highly expressed in wood-forming cells and their encoded proteins were shown to be localized in the Golgi. When overexpressed in the Arabidopsis gxm1/2/3 triple mutant, PtrGXMs were able to partially complement the mutant phenotypes including defects in glucuronoxylan methyltransferase activity and GlcA methylation in xylan, indicating that PtrGXMs most likely function as glucuronoxylan methyltransferases. Direct evidence was provided by enzymatic analysis of recombinant PtrGXM proteins showing that they possessed a methyltransferase activity capable of transferring the methyl group onto GlcA-substituted xylooligomers. Kinetic analysis showed that PtrGXMs exhibited differential affinities toward the GlcA-substituted xylooligomer acceptor with PtrGXM3 and PtrGXM4 having 10 times higher Km values than PtrGXM1 and PtrGXM2. Together, these findings indicate that PtrGXMs are methyltransferases mediating GlcA methylation in Populus xylan during wood formation. PMID:24523868

  18. Functional Oligomerization of the Saccharomyces cerevisiae Isoprenylcysteine Carboxyl Methyltransferase, Ste14p

    OpenAIRE

    Griggs, Amy M.; Hahne, Kalub; Hrycyna, Christine A.

    2010-01-01

    The isoprenylcysteine carboxyl methyltransferase (Icmt) from Saccharomyces cerevisiae, also designated Ste14p, is a 26-kDa integral membrane protein that contains six transmembrane spanning segments. This protein is localized to the endoplasmic reticulum membrane where it performs the methylation step of the CAAX post-translational processing pathway. Sequence analysis reveals a putative GXXXG dimerization motif located in transmembrane 1 of Ste14p, but it is not known whether Ste14p forms or...

  19. Sequence and structural evolution of the KsgA/Dim1 methyltransferase family

    Directory of Open Access Journals (Sweden)

    Rife Jason P

    2008-10-01

    Full Text Available Abstract Background One of the 60 or so genes conserved in all domains of life is the ksgA/dim1 orthologous group. Enzymes from this family perform the same post-transcriptional nucleotide modification in ribosome biogenesis, irrespective of organism. Despite this common function, divergence has enabled some family members to adopt new and sometimes radically different functions. For example, in S. cerevisiae Dim1 performs two distinct functions in ribosome biogenesis, while human mtTFB is not only an rRNA methyltransferase in the mitochondria but also a mitochondrial transcription factor. Thus, these proteins offer an unprecedented opportunity to study evolutionary aspects of structure/function relationships, especially with respect to our recently published work on the binding mode of a KsgA family member to its 30S subunit substrate. Here we compare and contrast KsgA orthologs from bacteria, eukaryotes, and mitochondria as well as the paralogous ErmC enzyme. Results By using structure and sequence comparisons in concert with a unified ribosome binding model, we have identified regions of the orthologs that are likely related to gains of function beyond the common methyltransferase function. There are core regions common to the entire enzyme class that are associated with ribosome binding, an event required in rRNA methylation activity, and regions that are conserved in subgroups that are presumably related to non-methyltransferase functions. Conclusion The ancient protein KsgA/Dim1 has adapted to cellular roles beyond that of merely an rRNA methyltransferase. These results provide a structural foundation for analysis of multiple aspects of ribosome biogenesis and mitochondrial transcription.

  20. Catecholamine-o-methyltransferase polymorphisms are associated with postoperative pain intensity.

    LENUS (Irish Health Repository)

    Lee, Peter J

    2011-02-01

    single nucleotide polymorphisms (SNPs) in the genes for catecholamine-O-methyltransferase (COMT), μ-opioid receptor and GTP cyclohydrolase (GCH1) have been linked to acute and chronic pain states. COMT polymorphisms are associated with experimental pain sensitivity and a chronic pain state. No such association has been identified perioperatively. We carried out a prospective observational clinical trial to examine associations between these parameters and the development of postoperative pain in patients undergoing third molar (M3) extraction.

  1. A glutamate/aspartate switch controls product specificity in a protein arginine methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Debler, Erik W.; Jain, Kanishk; Warmack, Rebeccah A.; Feng, You; Clarke, Steven G.; Blobel, Günter; Stavropoulos, Pete

    2016-02-08

    Trypanosoma brucei PRMT7 (TbPRMT7) is a protein arginine methyltransferase (PRMT) that strictly monomethylates various substrates, thus classifying it as a type III PRMT. However, the molecular basis of its unique product specificity has remained elusive. Here, we present the structure of TbPRMT7 in complex with its cofactor product S-adenosyl-L-homocysteine (AdoHcy) at 2.8 Å resolution and identify a glutamate residue critical for its monomethylation behavior. TbPRMT7 comprises the conserved methyltransferase and β-barrel domains, an N-terminal extension, and a dimerization arm. The active site at the interface of the N-terminal extension, methyltransferase, and β-barrel domains is stabilized by the dimerization arm of the neighboring protomer, providing a structural basis for dimerization as a prerequisite for catalytic activity. Mutagenesis of active-site residues highlights the importance of Glu181, the second of the two invariant glutamate residues of the double E loop that coordinate the target arginine in substrate peptides/proteins and that increase its nucleophilicity. Strikingly, mutation of Glu181 to aspartate converts TbPRMT7 into a type I PRMT, producing asymmetric dimethylarginine (ADMA). Isothermal titration calorimetry (ITC) using a histone H4 peptide showed that the Glu181Asp mutant has markedly increased affinity for monomethylated peptide with respect to the WT, suggesting that the enlarged active site can favorably accommodate monomethylated peptide and provide sufficient space for ADMA formation. In conclusion, these findings yield valuable insights into the product specificity and the catalytic mechanism of protein arginine methyltransferases and have important implications for the rational (re)design of PRMTs.

  2. Regulation of protein stability of DNA methyltransferase 1 by post-translational modifications

    OpenAIRE

    Scott, Anthony; Song, Jing; Ewing, Rob; Wang, Zhenghe

    2014-01-01

    DNA methylation is an important epigenetic mechanism that ensures correct gene expression and maintains genetic stability. DNA methyltransferase 1 (DNMT1) is the primary enzyme that maintains DNA methylation during replication. Dysregulation of DNMT1 is implicated in a variety of diseases. DNMT1 protein stability is regulated via various post-translational modifications, such as acetylation and ubiquitination, but also through protein–protein interactions. These mechanisms ensure DNMT1 is pro...

  3. Mutations in the DNA methyltransferase gene DNMT3A cause an overgrowth syndrome with intellectual disability

    DEFF Research Database (Denmark)

    Tatton-Brown, Katrina; Seal, Sheila; Ruark, Elise

    2014-01-01

    Overgrowth disorders are a heterogeneous group of conditions characterized by increased growth parameters and other variable clinical features such as intellectual disability and facial dysmorphism. To identify new causes of human overgrowth, we performed exome sequencing in ten proband...... and histone binding. Similar mutations were not present in 1,000 UK population controls (13/152 cases versus 0/1,000 controls; P intellectual disability and greater height. DNMT3A encodes a DNA methyltransferase essential for establishing...

  4. Lack of involvement of known DNA methyltransferases in familial hydatidiform mole implies the involvement of other factors in establishment of imprinting in the human female germline

    OpenAIRE

    Picton H M; Huntriss J; Hodge D; Judson H; de Vos M; Hayward B E; Sheridan E; Bonthron DT

    2003-01-01

    Abstract Background Differential methylation of the two alleles is a hallmark of imprinted genes. Correspondingly, loss of DNA methyltransferase function results in aberrant imprinting and abnormal post-fertilization development. In the mouse, mutations of the oocyte-specific isoform of the DNA methyltransferase Dnmt1 (Dnmt1o) and of the methyltransferase-like Dnmt3L gene result in specific failures of imprint establishment or maintenance, at multiple loci. We have previously shown in humans ...

  5. Cloning and characterization of mr-s, a novel SAM domain protein, predominantly expressed in retinal photoreceptor cells

    Directory of Open Access Journals (Sweden)

    Koike Chieko

    2006-03-01

    Full Text Available Abstract Background Sterile alpha motif (SAM domains are ~70 residues long and have been reported as common protein-protein interaction modules. This domain is found in a large number of proteins, including Polycomb group (PcG proteins and ETS family transcription factors. In this work, we report the cloning and functional characterization of a novel SAM domain-containing protein, which is predominantly expressed in retinal photoreceptors and the pineal gland and is designated mouse mr-s (major retinal SAM domain protein. Results mr-s is evolutionarily conserved from zebrafish through human, organisms through which the mechanism of photoreceptor development is also highly conserved. Phylogenetic analysis suggests that the SAM domain of mr-s is most closely related to a mouse polyhomeotic (ph ortholog, Mph1/Rae28, which is known as an epigenetic molecule involved in chromatin modifications. These findings provide the possibility that mr-s may play a critical role by regulating gene expression in photoreceptor development. mr-s is preferentially expressed in the photoreceptors at postnatal day 3–6 (P3-6, when photoreceptors undergo terminal differentiation, and in the adult pineal gland. Transcription of mr-s is directly regulated by the cone-rod homeodomain protein Crx. Immunoprecipitation assay showed that the mr-s protein self-associates mainly through the SAM domain-containing region as well as ph. The mr-s protein localizes mainly in the nucleus, when mr-s is overexpressed in HEK293T cells. Moreover, in the luciferase assays, we found that mr-s protein fused to GAL4 DNA-binding domain functions as a transcriptional repressor. We revealed that the repression activity of mr-s is not due to a homophilic interaction through its SAM domain but to the C-terminal region. Conclusion We identified a novel gene, mr-s, which is predominantly expressed in retinal photoreceptors and pineal gland. Based on its expression pattern and biochemical analysis

  6. Near-isogenic lines for measuring phenotypic effects of DIMBOA-Glc methyltransferase activity in maize.

    Science.gov (United States)

    Mijares, Valeria; Meihls, Lisa N; Jander, Georg; Tzin, Vered

    2013-10-01

    Three O-methyltransferases (BX10a, b, c) catalyze the conversion of 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one glucoside (DIM BOA-Glc) to 2-hydroxy-4,7-dimethoxy-1,4-benzoxazin-3-one glucoside (HDMBOA -Glc) in maize (Zea mays). Variation in benzoxazinoid accumulation and resistance to Rhopalosiphum maidis (corn leaf aphid) was attributed to a natural CACTA family transposon insertion that inactivates Bx10c. Whereas maize inbred line B73 has this transposon insertion, line CM L277 does not. To characterize the phenotypic effects of DIM BOA-Glc methyltransferase activity, we created near-isogenic lines derived from B73 and CM L277 that do or do not contain the transposon insertion. Bx10c inactivation causes high DIM BOA -Glc, low HDMBOA-Glc, and decreased aphid reproduction relative to near-isogenic lines that have a functional Bx10c gene. These results confirm the importance of this locus in maize aphid resistance. The availability of Bx10c near-isogenic lines will facilitate further research on the function of different benzoxazinoids and DIM BOA-Glc methyltransferase activity in maize defense against herbivores and pathogens.

  7. Independent Recruitment of an O-Methyltransferase for Syringyl Lignin Biosynthesis in Selaginella moellendorffii[W

    Science.gov (United States)

    Weng, Jing-Ke; Akiyama, Takuya; Ralph, John; Chapple, Clint

    2011-01-01

    Syringyl lignin, an important component of the secondary cell wall, has traditionally been considered to be a hallmark of angiosperms because ferns and gymnosperms in general lack lignin of this type. Interestingly, syringyl lignin was also detected in Selaginella, a genus that represents an extant lineage of the most basal of the vascular plants, the lycophytes. In angiosperms, syringyl lignin biosynthesis requires the activity of ferulate 5-hydroxylase (F5H), a cytochrome P450-dependent monooxygenase, and caffeic acid/5-hydroxyferulic acid O-methyltransferase (COMT). Together, these two enzymes divert metabolic flux from the biosynthesis of guaiacyl lignin, a lignin type common to all vascular plants, toward syringyl lignin. Selaginella has independently evolved an alternative lignin biosynthetic pathway in which syringyl subunits are directly derived from the precursors of p-hydroxyphenyl lignin, through the action of a dual specificity phenylpropanoid meta-hydroxylase, Sm F5H. Here, we report the characterization of an O-methyltransferase from Selaginella moellendorffii, COMT, the coding sequence of which is clustered together with F5H at the adjacent genomic locus. COMT is a bifunctional phenylpropanoid O-methyltransferase that can methylate phenylpropanoid meta-hydroxyls at both the 3- and 5-position and function in concert with F5H in syringyl lignin biosynthesis in S. moellendorffii. Phylogenetic analysis reveals that Sm COMT, like F5H, evolved independently from its angiosperm counterparts. PMID:21742988

  8. Identification of a Pseudomonas aeruginosa PAO1 DNA Methyltransferase, Its Targets, and Physiological Roles

    Directory of Open Access Journals (Sweden)

    Sebastian Doberenz

    2017-02-01

    Full Text Available DNA methylation is widespread among prokaryotes, and most DNA methylation reactions are catalyzed by adenine DNA methyltransferases, which are part of restriction-modification (R-M systems. R-M systems are known for their role in the defense against foreign DNA; however, DNA methyltransferases also play functional roles in gene regulation. In this study, we used single-molecule real-time (SMRT sequencing to uncover the genome-wide DNA methylation pattern in the opportunistic pathogen Pseudomonas aeruginosa PAO1. We identified a conserved sequence motif targeted by an adenine methyltransferase of a type I R-M system and quantified the presence of N6-methyladenine using liquid chromatography-tandem mass spectrometry (LC-MS/MS. Changes in the PAO1 methylation status were dependent on growth conditions and affected P. aeruginosa pathogenicity in a Galleria mellonella infection model. Furthermore, we found that methylated motifs in promoter regions led to shifts in sense and antisense gene expression, emphasizing the role of enzymatic DNA methylation as an epigenetic control of phenotypic traits in P. aeruginosa. Since the DNA methylation enzymes are not encoded in the core genome, our findings illustrate how the acquisition of accessory genes can shape the global P. aeruginosa transcriptome and thus may facilitate adaptation to new and challenging habitats.

  9. Transcriptome profiling of Set5 and Set1 methyltransferases: Tools for visualization of gene expression

    Directory of Open Access Journals (Sweden)

    Glòria Mas Martín

    2014-12-01

    Full Text Available Cells regulate transcription by coordinating the activities of multiple histone modifying complexes. We recently identified the yeast histone H4 methyltransferase Set5 and discovered functional overlap with the histone H3 methyltransferase Set1 in gene expression. Specifically, using next-generation RNA sequencing (RNA-Seq, we found that Set5 and Set1 function synergistically to regulate specific transcriptional programs at subtelomeres and transposable elements. Here we provide a comprehensive description of the methodology and analysis tools corresponding to the data deposited in NCBI's Gene Expression Omnibus (GEO under the accession number GSE52086. This data complements the experimental methods described in Mas Martín G et al. (2014 and provides the means to explore the cooperative functions of histone H3 and H4 methyltransferases in the regulation of transcription. Furthermore, a fully annotated R code is included to enable researchers to use the following computational tools: comparison of significant differential expression (SDE profiles; gene ontology enrichment of SDE; and enrichment of SDE relative to chromosomal features, such as centromeres, telomeres, and transposable elements. Overall, we present a bioinformatics platform that can be generally implemented for similar analyses with different datasets and in different organisms.

  10. Kinetic analysis of Yersinia pestis DNA adenine methyltransferase activity using a hemimethylated molecular break light oligonucleotide.

    Directory of Open Access Journals (Sweden)

    Robert J Wood

    Full Text Available BACKGROUND: DNA adenine methylation plays an important role in several critical bacterial processes including mismatch repair, the timing of DNA replication and the transcriptional control of gene expression. The dependence of bacterial virulence on DNA adenine methyltransferase (Dam has led to the proposal that selective Dam inhibitors might function as broad spectrum antibiotics. METHODOLOGY/PRINCIPAL FINDINGS: Herein we report the expression and purification of Yersinia pestis Dam and the development of a continuous fluorescence based assay for DNA adenine methyltransferase activity that is suitable for determining the kinetic parameters of the enzyme and for high throughput screening against potential Dam inhibitors. The assay utilised a hemimethylated break light oligonucleotide substrate containing a GATC methylation site. When this substrate was fully methylated by Dam, it became a substrate for the restriction enzyme DpnI, resulting in separation of fluorophore (fluorescein and quencher (dabcyl and therefore an increase in fluorescence. The assays were monitored in real time using a fluorescence microplate reader in 96 well format and were used for the kinetic characterisation of Yersinia pestis Dam, its substrates and the known Dam inhibitor, S-adenosylhomocysteine. The assay has been validated for high throughput screening, giving a Z-factor of 0.71+/-0.07 indicating that it is a sensitive assay for the identification of inhibitors. CONCLUSIONS/SIGNIFICANCE: The assay is therefore suitable for high throughput screening for inhibitors of DNA adenine methyltransferases and the kinetic characterisation of the inhibition.

  11. miR-29 Represses the Activities of DNA Methyltransferases and DNA Demethylases

    Directory of Open Access Journals (Sweden)

    Izuho Hatada

    2013-07-01

    Full Text Available Members of the microRNA-29 (miR-29 family directly target the DNA methyltransferases, DNMT3A and DNMT3B. Disturbances in the expression levels of miR-29 have been linked to tumorigenesis and tumor aggressiveness. Members of the miR-29 family are currently thought to repress DNA methylation and suppress tumorigenesis by protecting against de novo methylation. Here, we report that members of the miR-29 family repress the activities of DNA methyltransferases and DNA demethylases, which have opposing roles in control of DNA methylation status. Members of the miR-29 family directly inhibited DNA methyltransferases and two major factors involved in DNA demethylation, namely tet methylcytosine dioxygenase 1 (TET1 and thymine DNA glycosylase (TDG. Overexpression of miR-29 upregulated the global DNA methylation level in some cancer cells and downregulated DNA methylation in other cancer cells, suggesting that miR-29 suppresses tumorigenesis by protecting against changes in the existing DNA methylation status rather than by preventing de novo methylation of DNA.

  12. Structural insights into mechanisms of the small RNA methyltransferase HEN1

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Ying; Ji, Lijuan; Huang, Qichen; Vassylyev, Dmitry G.; Chen, Xuemei; Ma, Jin-Biao; (UAB); (UCR)

    2010-02-22

    RNA silencing is a conserved regulatory mechanism in fungi, plants and animals that regulates gene expression and defence against viruses and transgenes. Small silencing RNAs of {approx}20-30 nucleotides and their associated effector proteins, the Argonaute family proteins, are the central components in RNA silencing. A subset of small RNAs, such as microRNAs and small interfering RNAs (siRNAs) in plants, Piwi-interacting RNAs in animals and siRNAs in Drosophila, requires an additional crucial step for their maturation; that is, 2'-O-methylation on the 3' terminal nucleotide. A conserved S-adenosyl-L-methionine-dependent RNA methyltransferase, HUA ENHANCER 1 (HEN1), and its homologues are responsible for this specific modification. Here we report the 3.1 {angstrom} crystal structure of full-length HEN1 from Arabidopsis in complex with a 22-nucleotide small RNA duplex and cofactor product S-adenosyl-L-homocysteine. Highly cooperative recognition of the small RNA substrate by multiple RNA binding domains and the methyltransferase domain in HEN1 measures the length of the RNA duplex and determines the substrate specificity. Metal ion coordination by both 2' and 3' hydroxyls on the 3'-terminal nucleotide and four invariant residues in the active site of the methyltransferase domain suggests a novel Mg{sup 2+}-dependent 2'-O-methylation mechanism.

  13. MtrA of the sodium ion pumping methyltransferase binds cobalamin in a unique mode.

    Science.gov (United States)

    Wagner, Tristan; Ermler, Ulrich; Shima, Seigo

    2016-06-21

    In the three domains of life, vitamin B12 (cobalamin) is primarily used in methyltransferase and isomerase reactions. The methyltransferase complex MtrA-H of methanogenic archaea has a key function in energy conservation by catalysing the methyl transfer from methyl-tetrahydromethanopterin to coenzyme M and its coupling with sodium-ion translocation. The cobalamin-binding subunit MtrA is not homologous to any known B12-binding proteins and is proposed as the motor of the sodium-ion pump. Here, we present crystal structures of the soluble domain of the membrane-associated MtrA from Methanocaldococcus jannaschii and the cytoplasmic MtrA homologue/cobalamin complex from Methanothermus fervidus. The MtrA fold corresponds to the Rossmann-type α/β fold, which is also found in many cobalamin-containing proteins. Surprisingly, the cobalamin-binding site of MtrA differed greatly from all the other cobalamin-binding sites. Nevertheless, the hydrogen-bond linkage at the lower axial-ligand site of cobalt was equivalently constructed to that found in other methyltransferases and mutases. A distinct polypeptide segment fixed through the hydrogen-bond linkage in the relaxed Co(III) state might be involved in propagating the energy released upon corrinoid demethylation to the sodium-translocation site by a conformational change.

  14. Identification of des-methyl-DIF-1 methyltransferase in Dictyostelium purpureum.

    Science.gov (United States)

    Motohashi, Kazunori A; Morita, Naoki; Kato, Atsushi; Saito, Tamao

    2012-01-01

    The signalling molecule 1-(3,5-dichloro-2,6-dihydroxy-4-methoxyphenyl) hexan-1-one (DIF-1) is required for differentiation and pattern formation in Dictyostelium discoideum development. DIF-1 is synthesized by three enzymes, a hybrid polyketide synthase, a flavin-dependent halogenase, and a des-methyl-DIF-1 methyltransferase. The genome data on the related species D. purpureum are now public. Using this genome information, des-methyl-DIF-1 methyltransferase of D. purpureum was identified, and was named Dp dmtA. Overexpression of Dp dmtA complemented the defects in basal disc formation and lower cup formation in a dmtA knock-out mutant of D. discoideum. This indicates that Dp dmtA has the same function as D. discoideum dmtA and compensates for loss of the dmtA gene in the D. discoideum dmtA mutant. The materials released in the medium by D. purpureum contained stalk-inducing activity with the same retention time as that of DIF-1 in HPLC fractionation. This indicates that the stalk-inducing signal of DIF-1 and des-methyl-DIF-1 methyltransferase are conserved in D. purpureum.

  15. Mechanistic insights into small RNA recognition and modification by the HEN1 methyltransferase.

    Science.gov (United States)

    Plotnikova, Alexandra; Baranauskė, Simona; Osipenko, Aleksandr; Klimašauskas, Saulius; Vilkaitis, Giedrius

    2013-07-15

    The HEN1 methyltransferase from Arabidopsis thaliana modifies the 3'-terminal nucleotides of small regulatory RNAs. Although it is one of the best characterized members of the 2'-O-methyltransferase family, many aspects of its interactions with the cofactor and substrate RNA remained unresolved. To better understand the substrate interactions and contributions of individual steps during HEN1 catalysis, we studied the binding and methylation kinetics of the enzyme using a series of unmethylated, hemimethylated and doubly methylated miRNA and siRNA substrates. The present study shows that HEN1 specifically binds double-stranded unmethylated or hemimethylated miR173/miR173* substrates with a subnanomolar affinity in a cofactor-dependent manner. Kinetic studies under single turnover and pre-steady state conditions in combination with isotope partitioning analysis showed that the binary HEN1-miRNA/miRNA* complex is catalytically competent; however, successive methylation of the two strands in a RNA duplex occurs in a non-processive (distributive) manner. We also find that the observed moderate methylation strand preference is largely exerted at the RNA-binding step and is fairly independent of the nature of the 3'-terminal nucleobase, but shows some dependency on proximal nucleotide mispairs. The results of the present study thus provide novel insights into the mechanism of RNA recognition and modification by a representative small RNA 2'-O-methyltransferase.

  16. The activity of a yeast Family 16 methyltransferase, Efm2, is affected by a conserved tryptophan and its N-terminal region.

    Science.gov (United States)

    Hamey, Joshua J; Hart-Smith, Gene; Erce, Melissa A; Wilkins, Marc R

    2016-12-01

    The Family 16 methyltransferases are a group of eukaryotic nonhistone protein methyltransferases. Sixteen of these have recently been described in yeast and human, but little is known about their sequence and structural features. Here we investigate one of these methyltransferases, Saccharomyces cerevisiae elongation factor methyltransferase 2 (Efm2), by site-directed mutagenesis and truncation. We show that an active site-associated tryptophan, invariant in Family 16 methyltransferases and at position 222 in Efm2, is important for methyltransferase activity. A second highly conserved tryptophan, at position 318 in Efm2, is likely involved in S-adenosyl methionine binding but is of lesser consequence for catalysis. By truncation analysis, we show that the N-terminal 50-200 amino acids of Efm2 are critical for its methyltransferase activity. As N-terminal regions are variable among Family 16 methyltransferases, this suggests a possible role in determining substrate specificity. This is consistent with recently solved structures that show the core of Family 16 methyltransferases to be near-identical but the N termini to be structurally quite different. Finally, we show that Efm2 can exist as an oligomer but that its N terminus is not necessary for oligomerisation to occur.

  17. Rainfall declines over Queensland from 1951-2007 and links to the Subtropical Ridge and the SAM

    Energy Technology Data Exchange (ETDEWEB)

    Cottrill, D A [Bureau of Meteorology, 700 Collins St, Docklands, Melbourne, Victoria (Australia); Ribbe, J, E-mail: a.cottrill@bom.gov.a [University of Southern Queensland, Toowoomba, Queensland (Australia)

    2010-08-15

    Much of southern and eastern Australia including Queensland have experienced rainfall declines over recent decades affecting agricultural production and accelerating water infrastructure development. Rainfall declines from southern Australia have now been directly related to changes in the Southern Annular Mode (SAM) and the subtropical ridge. In southern and coastal Queensland, the rainfall declines have occurred mostly in the austral summer and autumn. Observations from this region reveal the rainfall decline is correlated to an increase in the mean sea level pressure (MSLP) at many stations. The largest increases in MSLP are over southeast Queensland and coastal regions, where some of the largest rainfall declines occur. This study indicates the subtropical ridge as one of the main factors in the rainfall decline over this region. SAM is also likely to be important, although its seasonal influence, apart from winter, is harder to determine.

  18. Benchmark Simulations of the Thermal-Hydraulic Responses during EBR-II Inherent Safety Tests using SAM

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Rui; Sumner, Tyler S.

    2016-04-17

    An advanced system analysis tool SAM is being developed for fast-running, improved-fidelity, and whole-plant transient analyses at Argonne National Laboratory under DOE-NE’s Nuclear Energy Advanced Modeling and Simulation (NEAMS) program. As an important part of code development, companion validation activities are being conducted to ensure the performance and validity of the SAM code. This paper presents the benchmark simulations of two EBR-II tests, SHRT-45R and BOP-302R, whose data are available through the support of DOE-NE’s Advanced Reactor Technology (ART) program. The code predictions of major primary coolant system parameter are compared with the test results. Additionally, the SAS4A/SASSYS-1 code simulation results are also included for a code-to-code comparison.

  19. Sulphur-bearing Compounds Detected by MSL SAM Evolved Gas Analysis of Materials from Yellowknife Bay, Gale Crater, Mars

    Science.gov (United States)

    McAdam, A. C.; Franz, H. B.; Archer, P. D. Jr.; Sutter, B.; Eigenbrode, J. L.; Freissinet, C.; Atreya, S. K.; Bish, D. L.; Blake, D. F.; Brunner, A.; hide

    2014-01-01

    The Sample Analysis at Mars (SAM) and Chemistry and Mineralogy (CheMin) instruments on the Mars Science Laboratory (MSL) analysed several subsamples of sample fines (<150 µm) from three sites in Yellowknife Bay, an aeolian bedform termed Rocknest (hereafter "RN") and two samples drilled from the Sheepbed mudstone at sites named John Klein ("JK") and Cumberland ("CB"). SAM's evolved gas analysis (EGA) mass spectrometry detected H2O, CO2, O2, H2, SO2, H2S, HCl, NO, OCS, CS2 and other trace gases. The identity of evolved gases and temperature (T) of evolution can support mineral detection by CheMin and place constraints on trace volatile-bearing phases present below the CheMin detection limit or difficult to characterize with XRD (e.g., X-ray amorphous phases). Here, we focus on potential constraints on phases that evolved SO2, H2S, OCS, and CS2 during thermal analysis.

  20. Mapping of Post-translational Modifications of Transition Proteins, TP1 and TP2, and Identification of Protein Arginine Methyltransferase 4 and Lysine Methyltransferase 7 as Methyltransferase for TP2*

    Science.gov (United States)

    Gupta, Nikhil; Madapura, M. Pradeepa; Bhat, U. Anayat; Rao, M. R. Satyanarayana

    2015-01-01

    In a unique global chromatin remodeling process during mammalian spermiogenesis, 90% of the nucleosomal histones are replaced by testis-specific transition proteins, TP1, TP2, and TP4. These proteins are further substituted by sperm-specific protamines, P1 and P2, to form a highly condensed sperm chromatin. In spermatozoa, a small proportion of chromatin, which ranges from 1 to 10% in mammals, retains the nucleosomal architecture and is implicated to play a role in transgenerational inheritance. However, there is still no mechanistic understanding of the interaction of chromatin machinery with histones and transition proteins, which facilitate this selective histone replacement from chromatin. Here, we report the identification of 16 and 19 novel post-translational modifications on rat endogenous transition proteins, TP1 and TP2, respectively, by mass spectrometry. By in vitro assays and mutational analysis, we demonstrate that protein arginine methyltransferase PRMT4 (CARM1) methylates TP2 at Arg71, Arg75, and Arg92 residues, and lysine methyltransferase KMT7 (Set9) methylates TP2 at Lys88 and Lys91 residues. Further studies with modification-specific antibodies that recognize TP2K88me1 and TP2R92me1 modifications showed that they appear in elongating to condensing spermatids and predominantly associated with the chromatin-bound TP2. This work establishes the repertoire of post-translational modifications that occur on TP1 and TP2, which may play a significant role in various chromatin-templated events during spermiogenesis and in the establishment of the sperm epigenome. PMID:25818198

  1. Mapping of Post-translational Modifications of Transition Proteins, TP1 and TP2, and Identification of Protein Arginine Methyltransferase 4 and Lysine Methyltransferase 7 as Methyltransferase for TP2.

    Science.gov (United States)

    Gupta, Nikhil; Madapura, M Pradeepa; Bhat, U Anayat; Rao, M R Satyanarayana

    2015-05-08

    In a unique global chromatin remodeling process during mammalian spermiogenesis, 90% of the nucleosomal histones are replaced by testis-specific transition proteins, TP1, TP2, and TP4. These proteins are further substituted by sperm-specific protamines, P1 and P2, to form a highly condensed sperm chromatin. In spermatozoa, a small proportion of chromatin, which ranges from 1 to 10% in mammals, retains the nucleosomal architecture and is implicated to play a role in transgenerational inheritance. However, there is still no mechanistic understanding of the interaction of chromatin machinery with histones and transition proteins, which facilitate this selective histone replacement from chromatin. Here, we report the identification of 16 and 19 novel post-translational modifications on rat endogenous transition proteins, TP1 and TP2, respectively, by mass spectrometry. By in vitro assays and mutational analysis, we demonstrate that protein arginine methyltransferase PRMT4 (CARM1) methylates TP2 at Arg(71), Arg(75), and Arg(92) residues, and lysine methyltransferase KMT7 (Set9) methylates TP2 at Lys(88) and Lys(91) residues. Further studies with modification-specific antibodies that recognize TP2K88me1 and TP2R92me1 modifications showed that they appear in elongating to condensing spermatids and predominantly associated with the chromatin-bound TP2. This work establishes the repertoire of post-translational modifications that occur on TP1 and TP2, which may play a significant role in various chromatin-templated events during spermiogenesis and in the establishment of the sperm epigenome. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. Possible Worlds in Sam Shepard’s Fool for Love and Its Cinematic Adaptation

    Directory of Open Access Journals (Sweden)

    Omid Amani

    2017-07-01

    Full Text Available This paper is an attempt to compare Sam Shepard’s Fool for Love, published in 1983, and its cinematic adaptation directed by Robert Altman, released in 1985, in terms of the notion of possible worlds (in the philosophically-oriented branch of cognitive poetics. In her discussion of possible worlds, Marie-Laure Ryan proposes a typology of accessibility relations (identity of properties, identity of inventory, compatibility of inventory, chronological, physical, taxonomic, logical, analytical, and linguistic compatibility in order to account for how the sense of the reality of our actual world reverberates in fictional worlds. Ryan also offers another typology, that is, the internal structure of the fictional world (knowledge worlds, intention worlds, wish worlds, obligation worlds, and fantasy worlds to show how the characters’ different conceptions of the world define and build up the narrative structure of fiction. It is argued that the change of medium – drama into film in this case – results in some changes in the possible worlds projected, since verbality and visuality give rise to some differences in terms of modes of perception. Also discussed is the significance in this regard of the spatio-temporal scope, which is confined in the case of drama, a genre often conceived to be performed, whereas in a movie the director is able to expand this scope as the occasion demands.

  3. Lysine-Tryptophan-Crosslinked Peptides Produced by Radical SAM Enzymes in Pathogenic Streptococci.

    Science.gov (United States)

    Schramma, Kelsey R; Seyedsayamdost, Mohammad R

    2017-04-21

    Macrocycles represent a common structural framework in many naturally occurring peptides. Several strategies exist for macrocyclization, and the enzymes that incorporate them are of great interest, as they enhance our repertoire for creating complex molecules. We recently discovered a new peptide cyclization reaction involving a crosslink between the side chains of lysine and tryptophan that is installed by a radical SAM enzyme. Herein, we characterize relatives of this metalloenzyme from the pathogens Streptococcus agalactiae and Streptococcus suis. Our results show that the corresponding enzymes, which we call AgaB and SuiB, contain multiple [4Fe-4S] clusters and catalyze Lys-Trp crosslink formation in their respective substrates. Subsequent high-resolution-MS and 2D-NMR analyses located the site of macrocyclization. Moreover, we report that AgaB can accept modified substrates containing natural or unnatural amino acids. Aside from providing insights into the mechanism of this unusual modification, the substrate promiscuity of AgaB may be exploited to create diverse macrocyclic peptides.

  4. Cloning and expressing a highly functional and substrate specific farnesoic acid o-methyltransferase from the Asian citrus psyllid (Diaphorina citri Kuwayama)

    Science.gov (United States)

    Van Ekert, Evelien; Shatters, Robert G.; Rougé, Pierre; Powell, Charles A.; Smagghe, Guy; Borovsky, Dov

    2015-01-01

    The Asian citrus psyllid, Diaphorina citri, transmits a phloem-limited bacterium, Candidatus ‘Liberibacter’ asiaticus that causes citrus greening disease. Because juvenile hormone (JH) plays an important role in adult and nymphal development, we studied the final steps in JH biosynthesis in D. citri. A putative JH acid methyltransferase ortholog gene (jmtD) and its cognate cDNA were identified by searching D. citri genome database. Expression analysis shows expression in all life stages. In adults, it is expressed in the head-thorax, (containing the corpora allata), and the abdomen (containing ovaries and male accessory glands). A 3D protein model identified the catalytic groove with catalytically active amino acids and the S-adenosyl methionine (SAM)-binding loop. The cDNA was expressed in Escherichia coli cells and the purified enzyme showed high preference for farnesoic acid (FA) and homoFA (kcat of 0.752 × 10−3 and 0.217 × 10−3 s−1, respectively) as compared to JH acid I (JHA I) (cis/trans/cis; 2Z, 6E, 10cis), JHA III (2E, 6E, 10cis), and JHA I (trans/cis/cis; 2E, 2Z, 10cis) (kcat of 0.081 × 10−3, 0.013 × 10−3, and 0.003 × 10−3 s−1, respectively). This suggests that this ortholog is a DcFA-o-methyl transferase gene (fmtD), not a jmtD, and that JH biosynthesis in D. citri proceeds from FA to JH III through methyl farnesoate (MF). DcFA-o-MT does not require Ca2+, Mg2+ or Zn2+, however, Zn2+ (1 mM) completely inhibits the enzyme probably by binding H115 at the active groove. This represents the first purified FA-o-MT from Hemiptera with preferred biological activity for FA and not JHA. PMID:25893162

  5. Kto samõi bogatõi v Estonii / Sten-Aleks Pihlak, Kärt Blumberg, Lemmi Kann

    Index Scriptorium Estoniae

    Pihlak, Sten-Aleks

    2008-01-01

    Eesti laevatranspordi kolmik - Ain Hanschmidt, Enn Pant ja Kalev Järvelill on Äripäeva rikaste edetabelis esimesed. Viimaste kuude suure aktsiahinnalanguse tõttu on paljud Äripäeva rikaste edetabelis olijad kaotanud igast kolmest kroonist kaks. Artiklis selgitatakse miks on Äripäeva Rikaste TOPis vähe naisi. Lisad: Iz TOP-500 samõhh bogatõhh ljudei v Estonii; Reitingi bogatshei za prezhnije godõ; Metodika

  6. Pengaruh Penggunaan Media Sosial Snapchat Terhadap Pengungkapan Diri Mahasiswa Jurusan Ilmu Komunikasi Fakultas Ilmu Sosial Dan Politik Universitas Sam Ratulangi

    OpenAIRE

    Mailoor, Adrian; Senduk, J. J; Londa, J. W

    2017-01-01

    Rumusan masalah dari penelitian ini adalah : Apakah Penggunaan Media Sosial Snapchat mempunyai pengaruh terhadap Pengungkapan Diri Mahasiswa Jurusan Ilmu Komunikasi Fakultas Ilmu Sosial dan Politik Universitas Sam Ratulangi. Menggunakan landasan Teori Self Disclosure. Joseph Luft dan Harry Ingham mengemukakan teori self disclosure yang didasarkan pada model interaksi manusia. Asumsi ini membawa Joseph Luft dan Harry Ingham menciptakan suatu teori atau model sebagai salah satu cara untuk melih...

  7. Is Samsāra actually the Same as Nirvāṇa ? A Critical Examination of ...

    African Journals Online (AJOL)

    A recent trend in interpreting Nāgārjuna's claim that samsāra is the same as nirvāṇa is to highlight the identity of dependent origination and emptiness in Nāgārjuna's writings. Although this trend is correct in foregrounding the identity claim in this central Mahāyāna thinker's philosophy, the complexity of the claim requires ...

  8. SAMS/Nimbus-7 Level 1 Radiance Data from CD-ROM V001 (SAMSN7L1RAD_CDROM) at GES DISC

    Data.gov (United States)

    National Aeronautics and Space Administration — SAMSN7L1RAD_CDROM is the gridded Nimbus-7 Stratospheric and Mesospheric Sounder (SAMS) Level 1 Radiance Data Product. The radiances were selected to derive gas...

  9. HPA and SAM axis responses as correlates of self- vs parental ratings of anxiety in boys with an Autistic Disorder.

    Science.gov (United States)

    Bitsika, Vicki; Sharpley, Christopher F; Sweeney, John A; McFarlane, James R

    2014-03-29

    Anxiety and Autistic Disorder (AD) are both neurological conditions and both disorders share some features that make it difficult to precisely allocate specific symptoms to each disorder. HPA and SAM axis activities have been conclusively associated with anxiety, and may provide a method of validating anxiety rating scale assessments given by parents and their children with AD about those children. Data from HPA axis (salivary cortisol) and SAM axis (salivary alpha amylase) responses were collected from a sample of 32 high-functioning boys (M age=11yr) with an Autistic Disorder (AD) and were compared with the boys' and their mothers' ratings of the boys' anxiety. There was a significant difference between the self-ratings given by the boys and ratings given about them by their mothers. Further, only the boys' self-ratings of their anxiety significantly predicted the HPA axis responses and neither were significantly related to SAM axis responses. Some boys showed cortisol responses which were similar to that previously reported in children who had suffered chronic and severe anxiety arising from stressful social interactions. As well as suggesting that some boys with an AD can provide valid self-assessments of their anxiety, these data also point to the presence of very high levels of chronic HPA-axis arousal and consequent chronic anxiety in these boys. Copyright © 2014 Elsevier Inc. All rights reserved.

  10. Against All Odds—“Sam Hall” and “The Man in Black”: From British Social Disparagement to American Defiant Individualism

    OpenAIRE

    Eva-Sabine Zehelein

    2007-01-01

    This article traces the development of the “Sam Hall” topos from its 18th or 17th century British origins to William Blake, to the Dubliners’ version of a Celtic traditional, to Hayes’ Harvard version, and finally to Johnny Cash. As an expression of socio-cultural criticism, the outlaw Sam Hall has been formed into a prototype of American defiant individualism “against all odds.”

  11. Against All Odds—“Sam Hall” and “The Man in Black”: From British Social Disparagement to American Defiant Individualism

    Directory of Open Access Journals (Sweden)

    Eva-Sabine Zehelein

    2007-12-01

    Full Text Available This article traces the development of the “Sam Hall” topos from its 18th or 17th century British origins to William Blake, to the Dubliners’ version of a Celtic traditional, to Hayes’ Harvard version, and finally to Johnny Cash. As an expression of socio-cultural criticism, the outlaw Sam Hall has been formed into a prototype of American defiant individualism “against all odds.”

  12. Selective translational repression of HIV-1 RNA by Sam68DeltaC occurs by altering PABP1 binding to unspliced viral RNA

    Directory of Open Access Journals (Sweden)

    Soros Vanessa

    2008-10-01

    Full Text Available Abstract HIV-1 structural proteins are translated from incompletely spliced 9 kb and 4 kb mRNAs, which are transported to the cytoplasm by Crm1. It has been assumed that once in the cytoplasm, translation of incompletely spliced HIV-1 mRNAs occurs in the same manner as host mRNAs. Previous analyses have demonstrated that Sam68 and a mutant thereof, Sam68ΔC, have dramatic effects on HIV gene expression, strongly enhancing and inhibiting viral structural protein synthesis, respectively. While investigating the inhibition of incompletely spliced HIV-1 mRNAs by Sam68ΔC, we determined that the effect was independent of the perinuclear bundling of the viral RNA. Inhibition was dependent upon the nuclear export pathway used, as translation of viral RNA exported via the Tap/CTE export pathway was not blocked by Sam68ΔC. We demonstrate that inhibition of HIV expression by Sam68ΔC is correlated with a loss of PABP1 binding with no attendant change in polyadenosine tail length of the affected RNAs. The capacity of Sam68ΔC to selectively inhibit translation of HIV-1 RNAs exported by Crm1 suggests that it is able to recognize unique characteristics of these viral RNPs, a property that could lead to new therapeutic approaches to controlling HIV-1 replication.

  13. The catechol-O-methyltransferase inhibitory potential of Z-vallesiachotamine by in silicoand in vitro approaches

    Directory of Open Access Journals (Sweden)

    Carolina dos Santos Passos

    Full Text Available AbstractZ-Vallesiachotamine is a monoterpene indole alkaloid that has a β-N-acrylate group in its structure. This class of compounds has already been described in different Psychotriaspecies. Our research group observed that E/Z-vallesiachotamine exhibits a multifunctional feature, being able to inhibit targets related to neurodegeneration, such as monoamine oxidase A, sirtuins 1 and 2, and butyrylcholinesterase enzymes. Aiming at better characterizing the multifunctional profile of this compound, its effect on cathecol-O-methyltransferase activity was investigated. The cathecol-O-methyltransferase activity was evaluated in vitro by a fluorescence-based method, using S-(5′-adenosyl-l-methionine as methyl donor and aesculetin as substrate. The assay optimization was performed varying the concentrations of methyl donor (S-(5′-adenosyl-l-methionine and enzyme. It was observed that the highest concentrations of both factors (2.25 U of the enzyme and 100 µM of S-(5′-adenosyl-l-methionine afforded the more reproducible results. The in vitro assay demonstrated that Z-vallesiachotamine was able to inhibit the cathecol-O-methyltransferase activity with an IC50 close to 200 µM. Molecular docking studies indicated that Z-vallesiachotamine can bind the catechol pocket of catechol-O-methyltransferase enzyme. The present work demonstrated for the first time the inhibitory properties of Z-vallesiachotamine on cathecol-O-methyltransferase enzyme, affording additional evidence regarding its multifunctional effects in targets related to neurodegenerative diseases.

  14. Bulk chemical analyses of petrographic thin sections of rocks of the Samli area, western Turkey

    Science.gov (United States)

    Lindsay, James R.; Leo, Gerhard W.

    1976-01-01

    Petrographic thin sections have been analyzed for their major-element composition using a fusion-dilution technique and measuring the intensity of X-rays by means of the electron microprobe. The balsam-mounted thin sections were removed from the glass slides by soaking them in methylene chloride. The freed sections were mixed with twice their weight of Li2B4O7, and fused at 1100° C. A fragment of the resulting glasslike bead was mounted for probe analysis. Both wavelength and energy-dispersive detector systems were used for quantitative determinations of elements sodium through iron. Because the samples and standards are diluted and fused, powdered rock standards may be used as reference materials. The calibration curves obtained by plotting X-ray intensity versus concentration of the analyte are linear over the concentration ranges used in this work and have an overall range of error of 2 to 8 percent. Although some of the analytical values show excessive scatter for petrographically similar rocks, in general the analyses are acceptable given the sample size and analytical uncertainties. Bulk chemical analyses were made of 18 thin sections, including altered diabase, amphibolite, and calc-silicate hornfels produced by progressive contact metamorphism and associated with metasomatic magnetite deposits in the Samli area, western Turkey. Normative plots indicate that (1) diabase and amphibolite are compositionally related and are similar to average compositions of basalts and orthoamphibolites, and (2) calc-silicate hornfels appears to have been derived in part from amphibolite and in part from crystalline limestone that underlies much of the region.

  15. Intrinsic resistance to aminoglycosides in Enterococcus faecium is conferred by the 16S rRNA m5C1404-specific methyltransferase EfmM

    DEFF Research Database (Denmark)

    Galimand, Marc; Schmitt, Emmanuelle; Panvert, Michel

    2011-01-01

    structure determination of EfmM at 2.28 Å resolution reveals an N-terminal domain connected to a central methyltransferase domain that is linked by a flexible lysine-rich region to two C-terminal subdomains. Mutagenesis of the methyltransferase domain established that two cysteines at specific tertiary...

  16. Crystal structure of the Escherichia coli 23S rRNA:m5C methyltransferase RlmI (YccW) reveals evolutionary links between RNA modification enzymes

    DEFF Research Database (Denmark)

    Sunita, S; Tkaczuk, Karolina L; Purta, Elzbieta

    2008-01-01

    Methylation is the most common RNA modification in the three domains of life. Transfer of the methyl group from S-adenosyl-l-methionine (AdoMet) to specific atoms of RNA nucleotides is catalyzed by methyltransferase (MTase) enzymes. The rRNA MTase RlmI (rRNA large subunit methyltransferase gene I...

  17. Arabidopsis DNA methyltransferase AtDNMT2 associates with histone deacetylase AtHD2s activity

    Energy Technology Data Exchange (ETDEWEB)

    Song, Yuan [Key Laboratory of Arid and Grassland Agroecology, Ministry of Education, School of Life Science, Lanzhou University, Lanzhou 730000 (China); Southern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, 1391 Sandford Street, London, ON, Canada N5V4T3 (Canada); Wu, Keqiang [Institute of Plant Biology, National Taiwan University, Taipei 106, Taiwan (China); Dhaubhadel, Sangeeta [Southern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, 1391 Sandford Street, London, ON, Canada N5V4T3 (Canada); An, Lizhe, E-mail: lizhean@lzu.edu.cn [Key Laboratory of Arid and Grassland Agroecology, Ministry of Education, School of Life Science, Lanzhou University, Lanzhou 730000 (China); Tian, Lining, E-mail: tianl@agr.gc.ca [Southern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, 1391 Sandford Street, London, ON, Canada N5V4T3 (Canada)

    2010-05-28

    DNA methyltransferase2 (DNMT2) is always deemed to be enigmatic, because it contains highly conserved DNA methyltransferase motifs but lacks the DNA methylation catalytic capability. Here we show that Arabidopsis DNA methyltransferase2 (AtDNMT2) is localized in nucleus and associates with histone deacetylation. Bimolecular fluorescence complementation and pull-down assays show AtDNMT2 interacts with type-2 histone deacetylases (AtHD2s), a unique type of histone deacetylase family in plants. Through analyzing the expression of AtDNMT2: ss-glucuronidase (GUS) fusion protein, we demonstrate that AtDNMT2 has the ability to repress gene expression at transcription level. Meanwhile, the expression of AtDNMT2 gene is altered in athd2c mutant plants. We propose that AtDNMT2 possibly involves in the activity of histone deacetylation and plant epigenetic regulatory network.

  18. Epigenetic changes of Arabidopsis genome associated with altered DNA methyltransferase and demethylase expressions after gamma irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Ji Eun; Cho, Eun Ju; Kim, Ji Hong; Chung, Byung Yeoup; Kim, Jin Hong [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

    2012-05-15

    DNA methylation at carbon 5 of cytosines is a hall mark of epigenetic inactivation and heterochromatin in both plants and mammals. In Arabidopsis, DNA methylation has two roles that protect the genome from selfish DNA elements and regulate gene expression. Plant genome has three types of DNA methyltransferase, METHYLTRANSFERASE 1 (MET1), DOMAINREARRANGED METHYLASE (DRM) and CHROMOMETHYLASE 3 (CMT3) that are capable of methylating CG, CHG (where H is A, T, or C) and CHH sites, respectively. MET1 is a maintenance DNA methyltransferase that controls CG methylation. Two members of the DRM family, DRM1 and DRM2, are responsible for de novo methylation of CG, CHG, and CHH sites but show a preference for CHH sites. Finally, CMT3 principally carries out CHG methylation and is involved in both de novo methylation and maintenance. Alternatively, active DNA demethylation may occur through the glycosylase activity by removing the methylcytosines from DNA. It may have essential roles in preventing transcriptional silencing of transgenes and endogenous genes and in activating the expression of imprinted genes. DNA demetylation in Arabidopsis is mediated by the DEMETER (DME) family of bifunctional DNA glycosylase. Three targets of DME are MEA (MEDEA), FWA (FLOWERING WAGENINGEN), and FIS2 (FERTILIZATION INDEPENDENT SEED 2). The DME family contains DEMETER-LIKE 2 (DML2), DML3, and REPRESSOR OF SILENING 1 (ROS1). DNA demetylation by ROS1, DML2, and DML3 protect the hypermethylation of specific genome loci. ROS1 is necessary to suppress the promoter methylation and the silencing of endogenous genes. In contrast, the function of DML2 and DML3 has not been reported. Several recent studies have suggested that epigenetic alterations such as change in DNA methylation and histone modification should be caused in plant genomes upon exposure to ionizing radiation. However, there is a lack of data exploring the underlying mechanisms. Therefore, the present study aims to characterize and

  19. Crystal structure of phosphoethanolamine methyltransferase from Plasmodium falciparum in complex with amodiaquine

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Soon Goo; Alpert, Tara D.; Jez, Joseph M. (WU)

    2012-07-17

    Phosphoethanolamine N-methyltransferase (PMT) is essential for phospholipid biogenesis in the malarial parasite Plasmodium falciparum. PfPMT catalyzes the triple methylation of phosphoethanolamine to produce phosphocholine, which is then used for phosphatidylcholine synthesis. Here we describe the 2.0 {angstrom} resolution X-ray crystal structure of PfPMT in complex with amodiaquine. To better characterize inhibition of PfPMT by amodiaquine, we determined the IC{sub 50} values of a series of aminoquinolines using a direct radiochemical assay. Both structural and functional analyses provide a possible approach for the development of new small molecule inhibitors of PfPMT.

  20. A fluorescence resonance energy transfer-based method for histone methyltransferases

    DEFF Research Database (Denmark)

    Devkota, Kanchan; Lohse, Brian; Nyby Jakobsen, Camilla

    2015-01-01

    A simple dye–quencher fluorescence resonance energy transfer (FRET)-based assay for methyltransferases was developed and used to determine kinetic parameters and inhibitory activity at EHMT1 and EHMT2. Peptides mimicking the truncated histone H3 tail were functionalized in each end with a dye...... and a quencher, respectively. When lysine-9 residues in the peptides were methylated, they were protected from cleavage by endoproteinase–EndoLysC, whereas unmethylated peptides were cleaved, resulting in an increase in fluorescent intensity....

  1. Turning a Substrate Peptide into a Potent Inhibitor for the Histone Methyltransferase SETD8

    Energy Technology Data Exchange (ETDEWEB)

    Judge, Russell A.; Zhu, Haizhong; Upadhyay, Anup K.; Bodelle, Pierre M.; Hutchins, Charles W.; Torrent, Maricel; Marin, Violeta L.; Yu, Wenyu; Vedadi, Masoud; Li, Fengling; Brown, Peter J.; Pappano, William N.; Sun, Chaohong; Petros, Andrew M.

    2016-12-08

    SETD8 is a histone H4–K20 methyltransferase that plays an essential role in the maintenance of genomic integrity during mitosis and in DNA damage repair, making it an intriguing target for cancer research. While some small molecule inhibitors for SETD8 have been reported, the structural binding modes for these inhibitors have not been revealed. Using the complex structure of the substrate peptide bound to SETD8 as a starting point, different natural and unnatural amino acid substitutions were tested, and a potent (Ki 50 nM, IC50 0.33 μM) and selective norleucine containing peptide inhibitor has been obtained.

  2. A rat brain cytosolic N-methyltransferase(s) activity converting phosphorylethanolamine into phosphorylcholine.

    Science.gov (United States)

    Andriamampandry, C; Massarelli, R; Freysz, L; Kanfer, J N

    1990-09-14

    It had been previously speculated upon but never proved that the methylation of phosphorylethanolamine could contribute to the production of choline containing compounds. However, experimental evidence obtained with neuronal cultures was interpreted as showing that the stepwise methylation of phosphobases may be an important route for this biosynthesis. We demonstrate that cytosolic fraction from rat brain possesses a N-methyltransferase activity capable of methylating phosphorylethanolamine and its mono- and dimethyl-derivatives into phosphorylcholine. The level of activity detectable in rat liver cytosol is only 18% of that found in the brain cytosol.

  3. Synthesis and optimization of N-heterocyclic pyridinones as catechol-O-methyltransferase (COMT) inhibitors.

    Science.gov (United States)

    Zhao, Zhijian; Harrison, Scott T; Schubert, Jeffrey W; Sanders, John M; Polsky-Fisher, Stacey; Zhang, Nanyan Rena; McLoughlin, Debra; Gibson, Christopher R; Robinson, Ronald G; Sachs, Nancy A; Kandebo, Monika; Yao, Lihang; Smith, Sean M; Hutson, Pete H; Wolkenberg, Scott E; Barrow, James C

    2016-06-15

    A series of N-heterocyclic pyridinone catechol-O-methyltransferase (COMT) inhibitors were synthesized. Physicochemical properties, including ligand lipophilic efficiency (LLE) and clogP, were used to guide compound design and attempt to improve inhibitor pharmacokinetics. Incorporation of heterocyclic central rings provided improvements in physicochemical parameters but did not significantly reduce in vitro or in vivo clearance. Nevertheless, compound 11 was identified as a potent inhibitor with sufficient in vivo exposure to significantly affect the dopamine metabolites homovanillic acid (HVA) and dihydroxyphenylacetic acid (DOPAC), and indicate central COMT inhibition. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. DNA Methyltransferase Protein Synthesis Is Reduced in CXXC Finger Protein 1–Deficient Embryonic Stem Cells

    OpenAIRE

    Butler, Jill S; Palam, Lakshmi R.; Tate, Courtney M.; Sanford, Jeremy R; Wek, Ronald C.; Skalnik, David G.

    2009-01-01

    CXXC finger protein 1 (CFP1) binds to unmethylated CpG dinucleotides and is required for embryogenesis. CFP1 is also a component of the Setd1A and Setd1B histone H3K4 methyltransferase complexes. Murine embryonic stem (ES) cells lacking CFP1 fail to differentiate, and exhibit a 70% reduction in global genomic cytosine methylation and a 50% reduction in DNA methyltransferase (DNMT1) protein and activity. This study investigated the underlying mechanism for reduced DNMT1 expression in CFP1-defi...

  5. Identification of the methyltransferase targeting C2499 in Deinococcus radiodurans 23S ribosomal RNA

    DEFF Research Database (Denmark)

    Nielsen, Julie Mundus; Flyvbjerg, Karen Freund; Kirpekar, Finn

    2016-01-01

    and in vivo. We also inactivated the DR_0049 gene in D. radiodurans through insertion of a chloramphenicol resistance cassette. This resulted in complete absence of the cytidine 2499 methylation, which all together demonstrates that DR_0049 encodes the methyltransferase producing m(5)C2499 in D. radiodurans...... 2499 reported. Using homology search, we identified the open reading frame DR_0049 as the primary candidate gene for the methyltransferase that modifies cytidine 2499. Mass spectrometric analysis demonstrated that recombinantly expressed DR0049 protein methylates E. coli cytidine 2499 both in vitro...

  6. Association between TPMT*3C and decreased thiopurine S-methyltransferase activity in patients with neuromyelitis optica spectrum disorders in China.

    Science.gov (United States)

    Gong, Xiaoqing; Mei, Shenghui; Li, Xindi; Li, Xingang; Zhou, Heng; Liu, Yonghong; Zhou, Anna; Yang, Li; Zhao, Zhigang; Zhang, Xinghu

    2017-12-01

    Thiopurines are effective drugs in treating neuromyelitis optica spectrum disorders and other diseases. Thiopurines' toxicity is mainly imputed to thiopurine S-methyltransferase activity. In Chinese population, the most common and important variation of thiopurine S-methyltransferase is TPMT*3C (rs1142345). This study aims to reveal the association between thiopurine S-methyltransferase activity and genetic polymorphisms of thiopurine S-methyltransferase in patients with neuromyelitis optica spectrum disorders in China. A liquid chromatography tandem mass/mass method was used to evaluate the thiopurine S-methyltransferase activity by using 6-mercapthioprine as the substrate in human erythrocyte haemolysate via 1 h incubation at 37 °C to form its methylated product 6-methylmercaptopurine. The amount of 6-methylmercaptopurine was adjusted by haematocrit and normalized to 8 × 10 8 erythrocytes. The selected polymorphisms of thiopurine S-methyltransferase were identified using MassARRAY system (Sequenom) and multiple SNaPshot technique. In 69 patients with neuromyelitis optica spectrum disorders, thiopurine S-methyltransferase activity was 80.29-154.53 (127.51 ± 16.83) pmol/h/8 × 10 8 erythrocytes. TPMT*3C (rs1142345) was associated with lower thiopurine S-methyltransferase activity (BETA = -25.37, P = 0.011). Other selected variants were not associated with thiopurine S-methyltransferase activity. TPMT*3C affects TPMT activity in Chinese patients with neuromyelitis optica spectrum disorders. Further studies are warranted to confirm the results. TPRs = thiopurines; NMOSD = neuromyelitis optica spectrum disorders; TPMT = thiopurine S-methyltransferase; LC-MS/MS = liquid chromatography tandem mass/mass; 6-MMP = 6-methylmercaptopurine; IS = internal standard; SNP = single nucleotide polymorphism; MAF = minor allele frequency; HWE = Hardy-Weinberg equilibrium; BETA = regression coefficients; UTR-3 = untranslated region 3.

  7. Influence of Oxychlorine Phases During the Pyrolysis of Organic Molecules: Implications for the Quest of Organics on Mars with the SAM Experiment Onboard the Curiosity Rover

    Science.gov (United States)

    Millan, M.; Szopa, C.; Buch, A.; Belmahdi, I.; Glavin, D. P.; Freissinet, C.; Eigenbrode, J. L.; Archer, P. D., Jr,; Sutter, B.; Mahaffy, P.

    2017-01-01

    One among the main objectives of the Sample Analysis at Mars (SAM) experiment is the in situ molecular analysis of gases evolving from solid samples heated up to approximately 850 degrees Centigrade, and collected by Curiosity on Mars surface/sub-surface in Gale crater. With this aim, SAM uses a gas-chromatograph coupled to a quadrupole mass spectrometer (GC-QMS) devoted to separate, detect and identify both volatile inorganic and organic compounds. SAM detected chlorinated organic molecules produced in evolved gas analysis (EGA) experiments. Several of these were also detected by the Viking experiments in 1976. SAM also detected oxychlorine compounds that were present at the Phoenix landing site. The oxychlorines may be prevelant over much of the martian surface. The C1 to C3 aliphatic chlorohydrocarbons (chloromethane and di- and trichloromethane) detected by SAM were attributed to reaction products occurring between the oxychlorines phases and the organic compounds coming from SAM instrument background. But SAM also showed the presence of a large excess of chlorobenzene and C2 to C4 dichloroalkanes among the volatile species released by the Cumberland sample of the Sheepbed mudstone. For the first time in the history of the Mars exploration, this proved the presence of Mars indigenous organic material at the Mars' surface. However, the identification of the precursor organic compounds of these chlorohydrocarbons is difficult due to the complexity of the reactions occurring during the sample pyrolysis. Laboratory pyrolysis experiments have demonstrated that oxychlorines phases such as perchlorates and chlorates, decomposed into dioxygen and volatile chlorine bearing molecules (HCl and/or Cl2) during the pyrolysis. These chemical species can then react with the organic molecules present in the martian solid samples through oxidation, chlorination and oxychlorination processes.

  8. The Investigation of Perchlorate/Iron Phase Mixtures as A Possible Source of Oxygen Detected by the Sample Analysis at Mars (SAM) Instrument in Gale Crater, Mars

    Science.gov (United States)

    Sutter, B.; Heil, E.; Morris, R. V.; Archer, P. D.; Ming, D. W.; Niles, P. B.; Eigenbrode, J. L.; Franz, H.; Freissinet C.; Glavin, D. P.; hide

    2015-01-01

    The Sample Analysis at Mars (SAM) instrument onboard the Curiosity rover detected O2 and HCl gas releases from the Rocknest (RN) eolian bedform and the John Klein (JK) and Cumberland (CB) drill hole materials in Gale Crater. Chlorinated hydrocarbons have also been detected by the SAM quadrupole mass spectrometer (QMS) and gas chromatography/mass spectrometer (GCMS). These detections along with the detection of perchlorate (ClO4-) by the Mars Phoenix Lander's Wet Chemistry Laboratory (WCL) suggesting perchlorate is a possible candidate for evolved O2 and chlorine species. Laboratory thermal analysis of individual per-chlorates has yet to provide an unequivocal temperature match to the SAM O2 and HCl release data. These detections along with the detection of perchlorate (ClO4-) by the Mars Phoenix Lander's Wet Chemistry Laboratory suggested perchlorate is a possible candidate for evolved O2 and chlorine species. Laboratory thermal analysis of pure perchlorates has yet to provide an unequivocal temperature match to the SAM O2 and HCl release data. Analog laboratory analysis of iron mineralogy detected in Gale materials that was physically mixed with Ca- and Mg-perchlorate has been shown to catalyze lower O2 release temperatures and approach some SAM O2 release data. Instead of physical mixtures used in previous work, the work presented here utilized perchlorate solutions added to Fe phases. This technique allowed for perchlorate to come in closer contact with the Fe-phase and may more closely mimic Mars conditions where humidity can increase enough to cause deliquescence of the highly hygroscopic perchlorate phases. The objective of this work is to: 1) Utilize a laboratory SAM analog instrument to evaluate the O2 release temperatures from Mg- and Ca-perchlorates solutions applied to Fephases detetected in Gale Crate; and 2) Determine if perchlorate solutions can provide improved matches with the SAM O2 temperature release profiles.

  9. Tracking the MSL-SAM methane detection source location Through Mars Regional Atmospheric Modeling System (MRAMS)

    Science.gov (United States)

    Pla-García, Jorge

    2016-04-01

    1. Introduction: The putative in situ detection of methane by Sample Analysis at Mars (SAM) instrument suite on Curiosi-ty at Gale crater has garnered significant attention because of the potential implications for the presence of geological methane sources or indigenous Martian organisms [1, 2]. SAM reported detection of back-ground levels of atmospheric methane of mean value 0.69±0.25 parts per billion by volume (ppbv) at the 95% confidence interval (CI). Additionally, in four sequential measurements spanning a 60-sol period, SAM observed elevated levels of methane of 7.2±2.1 ppbv (95% CI), implying that Mars is episodically producing methane from an additional unknown source. There are many major unresolved questions regard-ing this detection: 1) What are the potential sources of the methane release? 2) What causes the rapid decrease in concentration? and 3) Where is the re-lease location? 4) How spatially extensive is the re-lease? 5) For how long is CH4 released? Regarding the first question, the source of methane, is so far not identified. It could be related with geo-logical process like methane release from clathrates [3], serpentinisation [4] and volcanism [5]; or due to biological activity from methanogenesis [6]. To answer the second question, the rapid decrease in concentration, it is important to note that the photo-chemical lifetime of methane is of order 100 years, much longer than the atmospheric mixing time scale, and thus the gas should tend to be well mixed except near a source or shortly after an episodic release. The observed spike of 7 ppb from the background of explain the observed fast destruction of methane [7]. In an effort to better address the potential mixing and remaining questions, atmospheric circulation studies of Gale Crater were performed with the Mars Re-gional Atmospheric Modeling System (MRAMS). The model was focused on rover locations using nested grids with a spacing of 330 meters on the in-nermost grid that is centered

  10. Diver visibility measured with a compact scattering-attenuation meter (SAM) compatible with AUVs and other small deployment platforms

    Science.gov (United States)

    Twardowski, Michael S.; Zaneveld, J. Ronald V.; Moore, Casey M.; Mueller, James; Trees, Charles; Schofield, Oscar; Freeman, Scott; Helble, Tyler; Hong, Gerry

    2005-05-01

    An appropriate determination of water clarity is required by defense and security operations assessing subsurface threats compromising harbor and coastal security. For search and inspection operations involving divers, underwater imaging, and electro-optical identification (EOID) systems such as laser line-scanners, the key environmental parameter needed is the optical attenuation coefficient (directly related to diver visibility). To address this need, a scattering-attenuation meter (SAM) measuring attenuation and diver visibility was developed for integration on new compact surveying platforms such as ROVs and the REMUS and glider AUVs. The sensor is compact (18X8X6 cm3), low power, robust, and hydrodynamic with a flat sensing face. The SAM measures attenuation using a novel dual-scattering approach that solves the paradox of making high-resolution attenuation measurements over the long pathlengths required for natural waters with a compact sensor. Attenuation and visibility data is presented from San Diego harbor in coordination with video images of bottom topography collected with a REMUS vehicle, from around New York harbor with a SAM mounted in an autonomous Slocum glider, and from Narragansett Bay. Results show that 1) visibility and/or attenuation in harbor and coastal regions can change rapidly over small scales (meters), especially near the bottom, 2) turbid bottom nepheloid layers are common, 3) typical visibility and/or attenuation levels fall in a range where knowledge of visibility and/or attenuation can be essential in the decision making process for security operations, and 4) attenuation is a significantly more accurate proxy for diver visibility than backscattering.

  11. Evaluating auditory stream segregation of SAM tone sequences by subjective and objective psychoacoustical tasks, and brain activity

    Directory of Open Access Journals (Sweden)

    Lena-Vanessa eDollezal

    2014-06-01

    Full Text Available Auditory stream segregation refers to a segregated percept of signal streams with different acoustic features. Different approaches have been pursued in studies of stream segregation. In psychoacoustics, stream segregation has mostly been investigated with a subjective task asking the subjects to report their percept. Few studies have applied an objective task in which stream segregation is evaluated indirectly by determining thresholds for a percept that depends on whether auditory streams are segregated or not. Furthermore, both perceptual measures and physiological measures of brain activity have been employed but only little is known about their relation. How the results from different tasks and measures are related is evaluated in the present study using examples relying on the ABA- stimulation paradigm that apply the same stimuli. We presented A and B signals that were sinusoidally amplitude modulated (SAM tones providing purely temporal, spectral or both types of cues to evaluate perceptual stream segregation and its physiological correlate. Which types of cues are most prominent was determined by the choice of carrier and modulation frequencies (fmod of the signals. In the subjective task subjects reported their percept and in the objective task we measured their sensitivity for detecting time-shifts of B signals in an ABA- sequence. As a further measure of processes underlying stream segregation we employed functional magnetic resonance imaging (fMRI. SAM tone parameters were chosen to evoke an integrated (1-stream, a segregated (2-stream or an ambiguous percept by adjusting the fmod difference between A and B tones (∆fmod. The results of both psychoacoustical tasks are significantly correlated. BOLD responses in fMRI depend on ∆fmod between A and B SAM tones. The effect of ∆fmod, however, differs between auditory cortex and frontal regions suggesting differences in representation related to the degree of perceptual ambiguity of

  12. Identification of a methyltransferase catalyzing the final step of methyl anthranilate synthesis in cultivated strawberry.

    Science.gov (United States)

    Pillet, Jeremy; Chambers, Alan H; Barbey, Christopher; Bao, Zhilong; Plotto, Anne; Bai, Jinhe; Schwieterman, Michael; Johnson, Timothy; Harrison, Benjamin; Whitaker, Vance M; Colquhoun, Thomas A; Folta, Kevin M

    2017-08-31

    Methyl anthranilate (MA) contributes an attractive fruity note to the complex flavor and aroma of strawberry (Fragaria spp.), yet it is rare in modern cultivars. The genetic basis for its biosynthesis has not been elucidated. Understanding the specific genes required for its synthesis could allow  the development of gene/allele-specific molecular markers to speed breeding of flavorful strawberries. Ripe fruits from individuals in an F1 population resulting from a cross between a MA producer and a non-producer were examined using a bulk-segregant transcriptome approach. MA producer and non-producer transcriptomes were compared, revealing five candidate transcripts that strictly co-segregated with MA production. One candidate encodes an annotated methyltransferase. MA levels are lower when this transcript is suppressed with RNAi, and bacterial cultures expressing the protein produced MA in the presence of anthranilic acid. Frozen fruit powders reconstituted with anthranilic acid and a methyl donor produced MA only if the transcript was detected in the fruit powder. A DNA-based molecular marker was developed that segregates with the MA-producing gene variant. These analyses indicate that the methyltransferase, now noted ANTHRANILIC ACID METHYL TRANSFERASE (FanAAMT), mediates the ultimate step of MA production in cultivated strawberry. Identification of this gene and its associated molecular marker may hasten breeding efforts to introduce this important volatile into modern cultivars.

  13. Characterization of a Bvg-regulated fatty acid methyl-transferase in Bordetella pertussis.

    Science.gov (United States)

    Rivera-Millot, Alex; Lesne, Elodie; Solans, Luis; Coutte, Loic; Bertrand-Michel, Justine; Froguel, Philippe; Dhennin, Véronique; Hot, David; Locht, Camille; Antoine, Rudy; Jacob-Dubuisson, Françoise

    2017-01-01

    The whooping cough agent Bordetella pertussis controls the expression of its large virulence regulon in a coordinated manner through the two-component signal transduction system BvgAS. In addition to the genes coding for bona fide virulence factors, the Bvg regulon comprises genes of unknown function. In this work, we characterized a new Bvg-activated gene called BP2936. Homologs of BP2936 are found in other pathogenic Bordetellae and in several other species, including plant pathogens and environmental bacteria. We showed that the gene product of BP2936 is a membrane-associated methyl-transferase of free fatty acids. We thus propose to name it FmtB, for fatty acid methyl-transferase of Bordetella. The role of this protein was tested in cellular and animal models of infection, but the loss of BP2936 did not appear to affect host-pathogen interactions in those assays. The high level of conservation of BP2936 among B. pertussis isolates nevertheless argues that it probably plays a role in the life cycle of this pathogen.

  14. Identification of the methyltransferase targeting C2499 in Deinococcus radiodurans 23S ribosomal RNA.

    Science.gov (United States)

    Mundus, Julie; Flyvbjerg, Karen Freund; Kirpekar, Finn

    2016-01-01

    The bacterium Deinococcus radiodurans-like all other organisms-introduces nucleotide modifications into its ribosomal RNA. We have previously found that the bacterium contains a Carbon-5 methylation on cytidine 2499 of its 23S ribosomal RNA, which is so far the only modified version of cytidine 2499 reported. Using homology search, we identified the open reading frame DR_0049 as the primary candidate gene for the methyltransferase that modifies cytidine 2499. Mass spectrometric analysis demonstrated that recombinantly expressed DR0049 protein methylates E. coli cytidine 2499 both in vitro and in vivo. We also inactivated the DR_0049 gene in D. radiodurans through insertion of a chloramphenicol resistance cassette. This resulted in complete absence of the cytidine 2499 methylation, which all together demonstrates that DR_0049 encodes the methyltransferase producing m(5)C2499 in D. radiodurans 23S rRNA. Growth experiments disclosed that inactivation of DR_0049 is associated with a severe growth defect, but available ribosome structures show that cytidine 2499 is positioned very similar in D. radiodurans harbouring the modification and E. coli without the modification. Hence there is no obvious structure-based explanation for the requirement for the C2499 posttranscriptional modification in D. radiodurans.

  15. DNA methyltransferase 1 (Dnmt1) mutation affects Snrpn imprinting in the mouse male germ line.

    Science.gov (United States)

    Saferali, Aabida; Moussette, Sanny; Chan, Donovan; Trasler, Jacquetta; Chen, Taiping; Rozen, Rima; Naumova, Anna K

    2012-09-01

    DNA methylation and DNA methyltransferases are essential for spermatogenesis. Mutations in the DNA methyltransferase Dnmt1 gene exert a paternal effect on epigenetic states and phenotypes of offspring, suggesting that DNMT1 is important for the epigenetic remodeling of the genome that takes place during spermatogenesis. However, the specific role of DNMT1 in spermatogenesis and the establishment of genomic imprints in the male germ line remains elusive. To further characterize the effect of DNMT1 deficiency on the resetting of methylation imprints during spermatogenesis, we analyzed the methylation profiles of imprinted regions in the spermatozoa of mice that were heterozygous for a Dnmt1 loss-of-function mutation. The mutation did not affect the H19 or IG differentially methylated regions (DMRs) that are usually highly methylated but led to a partial hypermethylation of the Snrpn DMR, a region that should normally be unmethylated in mature spermatozoa. This defect does not appear in mouse models with mutations in Dnmt3a and Mthfr genes and, therefore, it is specific for the Dnmt1 gene and is suggestive of a role of DNMT1 in imprint resetting or maintenance in the male germ line.

  16. The murine catecholamine methyltransferase mTOMT is essential for mechanotransduction by cochlear hair cells

    Science.gov (United States)

    Cunningham, Christopher L; Wu, Zizhen; Jafari, Aria; Zhao, Bo; Schrode, Kat; Harkins-Perry, Sarah; Lauer, Amanda; Müller, Ulrich

    2017-01-01

    Hair cells of the cochlea are mechanosensors for the perception of sound. Mutations in the LRTOMT gene, which encodes a protein with homology to the catecholamine methyltransferase COMT that is linked to schizophrenia, cause deafness. Here, we show that Tomt/Comt2, the murine ortholog of LRTOMT, has an unexpected function in the regulation of mechanotransduction by hair cells. The role of mTOMT in hair cells is independent of mTOMT methyltransferase function and mCOMT cannot substitute for mTOMT function. Instead, mTOMT binds to putative components of the mechanotransduction channel in hair cells and is essential for the transport of some of these components into the mechanically sensitive stereocilia of hair cells. Our studies thus suggest functional diversification between mCOMT and mTOMT, where mTOMT is critical for the assembly of the mechanotransduction machinery of hair cells. Defects in this process are likely mechanistically linked to deafness caused by mutations in LRTOMT/Tomt. DOI: http://dx.doi.org/10.7554/eLife.24318.001 PMID:28504928

  17. The motility of a human parasite, Toxoplasma gondii, is regulated by a novel lysine methyltransferase.

    Directory of Open Access Journals (Sweden)

    Aoife T Heaslip

    2011-09-01

    Full Text Available Protozoa in the phylum Apicomplexa are a large group of obligate intracellular parasites. Toxoplasma gondii and other apicomplexan parasites, such as Plasmodium falciparum, cause diseases by reiterating their lytic cycle, comprising host cell invasion, parasite replication, and parasite egress. The successful completion of the lytic cycle requires that the parasite senses changes in its environment and switches between the non-motile (for intracellular replication and motile (for invasion and egress states appropriately. Although the signaling pathway that regulates the motile state switch is critical to the pathogenesis of the diseases caused by these parasites, it is not well understood. Here we report a previously unknown mechanism of regulating the motility activation in Toxoplasma, mediated by a protein lysine methyltransferase, AKMT (for Apical complex lysine (K methyltransferase. AKMT depletion greatly inhibits activation of motility, compromises parasite invasion and egress, and thus severely impairs the lytic cycle. Interestingly, AKMT redistributes from the apical complex to the parasite body rapidly in the presence of egress-stimulating signals that increase [Ca²⁺] in the parasite cytoplasm, suggesting that AKMT regulation of parasite motility might be accomplished by the precise temporal control of its localization in response to environmental changes.

  18. Histone methyltransferase 1 regulates the encystation process in the parasite Giardia lamblia.

    Science.gov (United States)

    Salusso, Agostina; Zlocowski, Natacha; Mayol, Gonzalo F; Zamponi, Nahuel; Rópolo, Andrea S

    2017-08-01

    In eukaryotes, histone lysine methylation is associated with either active or repressed chromatin states, depending on the status of methylation. Even when the amino-terminus of Giardia lamblia histones diverges from other organisms, these regions contain lysine residues that are potential targets for methylation. When we examined the role of the histone methyltransferase 1 (HMT1) in the regulation of the encystation process by giardial histone methyltransferase 1 (GlHMT1) overexpression or downregulation, we observed an increase or a decrease in cyst production, respectively, compared to wild-type trophozoites. A time-lapse analysis of encystation showed that overexpression of GlHMT1 induced an earlier and faster process than in wild-type cells together with an upregulation of mRNA expression of cyst wall proteins. Subcellular localization studies indicated that GlHMT1-hemaglutinin was mainly associated with the nuclear and perinuclear region in both growing and encysting parasites, in agreement with bioinformatics analyses showing that GlHMT-1 possesses nuclear localization signals in addition to the classical SU(var)3-9, Enhancer-of-Zeste, Trithorax (SET), and post-SET domains. Altogether, these findings suggest that the function of HMT1 is critical for the success and timing of the encystation process, and reinforce the idea that epigenetic marks are critical for cyst formation in G. lamblia. © 2017 Federation of European Biochemical Societies.

  19. The murine catecholamine methyltransferase mTOMT is essential for mechanotransduction by cochlear hair cells.

    Science.gov (United States)

    Cunningham, Christopher L; Wu, Zizhen; Jafari, Aria; Zhao, Bo; Schrode, Kat; Harkins-Perry, Sarah; Lauer, Amanda; Müller, Ulrich

    2017-05-15

    Hair cells of the cochlea are mechanosensors for the perception of sound. Mutations in the LRTOMT gene, which encodes a protein with homology to the catecholamine methyltransferase COMT that is linked to schizophrenia, cause deafness. Here, we show that Tomt/Comt2, the murine ortholog of LRTOMT, has an unexpected function in the regulation of mechanotransduction by hair cells. The role of mTOMT in hair cells is independent of mTOMT methyltransferase function and mCOMT cannot substitute for mTOMT function. Instead, mTOMT binds to putative components of the mechanotransduction channel in hair cells and is essential for the transport of some of these components into the mechanically sensitive stereocilia of hair cells. Our studies thus suggest functional diversification between mCOMT and mTOMT, where mTOMT is critical for the assembly of the mechanotransduction machinery of hair cells. Defects in this process are likely mechanistically linked to deafness caused by mutations in LRTOMT/Tomt.

  20. Identification and expression profiling of DNA methyltransferases during development and stress conditions in Solanaceae.

    Science.gov (United States)

    Kumar, Rahul; Chauhan, Pankaj Kumar; Khurana, Ashima

    2016-09-01

    DNA methyltransferase (DMTase) enzymes contribute to plant development and stress responses by de novo establishment and subsequent maintenance of DNA methylation during replication. However, the molecular mechanism underlying this activity remains obscure, especially in crop species. Using DMTase homolog complement in six Solanaceae species, we demonstrated here that their number remained conserved in Solanum lineage, whereas it was expanded in both pepper and Nicotiana benthamiana. Non-synonymous vs synonymous (Ka/Ks) substitution ratio revealed that most of the Solanaceous DMTase homologs undergo purifying selection. The genomic sequences of tomato DMT homologs in its wild relative, Solanum pennellii, remained highly conserved in their exons and methyltransferase domains. Structure analysis further revealed highly similar folding of DMTase homologs and conservation in the residues participating in protein-protein interaction in Solanum lineage, whereas a considerable diversification was observed of pepper homologs. Transcript profiling of DMTases highlighted both similar and distinct expression patterns of tomato homologs in other species during fruit development and stress responses. Overall, our analysis provides a strong basis for in-depth exploration of both conserved as well as distinct functions of tomato DMTase homologs in other economically important Solanaceae species.

  1. Regulation of DNA replication and chromosomal polyploidy by the MLL-WDR5-RBBP5 methyltransferases

    Directory of Open Access Journals (Sweden)

    Fei Lu

    2016-10-01

    Full Text Available DNA replication licensing occurs on chromatin, but how the chromatin template is regulated for replication remains mostly unclear. Here, we have analyzed the requirement of histone methyltransferases for a specific type of replication: the DNA re-replication induced by the downregulation of either Geminin, an inhibitor of replication licensing protein CDT1, or the CRL4CDT2 ubiquitin E3 ligase. We found that siRNA-mediated reduction of essential components of the MLL-WDR5-RBBP5 methyltransferase complexes including WDR5 or RBBP5, which transfer methyl groups to histone H3 at K4 (H3K4, suppressed DNA re-replication and chromosomal polyploidy. Reduction of WDR5/RBBP5 also prevented the activation of H2AX checkpoint caused by re-replication, but not by ultraviolet or X-ray irradiation; and the components of MLL complexes co-localized with the origin recognition complex (ORC and MCM2-7 replicative helicase complexes at replication origins to control the levels of methylated H3K4. Downregulation of WDR5 or RBBP5 reduced the methylated H3K4 and suppressed the recruitment of MCM2-7 complexes onto replication origins. Our studies indicate that the MLL complexes and H3K4 methylation are required for DNA replication but not for DNA damage repair.

  2. Structure of the arginine methyltransferase PRMT5-MEP50 reveals a mechanism for substrate specificity.

    Directory of Open Access Journals (Sweden)

    Meng-Chiao Ho

    Full Text Available The arginine methyltransferase PRMT5-MEP50 is required for embryogenesis and is misregulated in many cancers. PRMT5 targets a wide variety of substrates, including histone proteins involved in specifying an epigenetic code. However, the mechanism by which PRMT5 utilizes MEP50 to discriminate substrates and to specifically methylate target arginines is unclear. To test a model in which MEP50 is critical for substrate recognition and orientation, we determined the crystal structure of Xenopus laevis PRMT5-MEP50 complexed with S-adenosylhomocysteine (SAH. PRMT5-MEP50 forms an unusual tetramer of heterodimers with substantial surface negative charge. MEP50 is required for PRMT5-catalyzed histone H2A and H4 methyltransferase activity and binds substrates independently. The PRMT5 catalytic site is oriented towards the cross-dimer paired MEP50. Histone peptide arrays and solution assays demonstrate that PRMT5-MEP50 activity is inhibited by substrate phosphorylation and enhanced by substrate acetylation. Electron microscopy and reconstruction showed substrate centered on MEP50. These data support a mechanism in which MEP50 binds substrate and stimulates PRMT5 activity modulated by substrate post-translational modifications.

  3. The Histone Methyltransferase Activity of MLL1 Is Dispensable for Hematopoiesis and Leukemogenesis

    Directory of Open Access Journals (Sweden)

    Bibhu P. Mishra

    2014-05-01

    Full Text Available Despite correlations between histone methyltransferase (HMT activity and gene regulation, direct evidence that HMT activity is responsible for gene activation is sparse. We address the role of the HMT activity for MLL1, a histone H3 lysine 4 (H3K4 methyltransferase critical for maintaining hematopoietic stem cells (HSCs. Here, we show that the SET domain, and thus HMT activity of MLL1, is dispensable for maintaining HSCs and supporting leukemogenesis driven by the MLL-AF9 fusion oncoprotein. Upon Mll1 deletion, histone H4 lysine 16 (H4K16 acetylation is selectively depleted at MLL1 target genes in conjunction with reduced transcription. Surprisingly, inhibition of SIRT1 is sufficient to prevent the loss of H4K16 acetylation and the reduction in MLL1 target gene expression. Thus, recruited MOF activity, and not the intrinsic HMT activity of MLL1, is central for the maintenance of HSC target genes. In addition, this work reveals a role for SIRT1 in opposing MLL1 function.

  4. Comparative analysis of DNA methyltransferase gene family in fungi: a focus on Basidiomycota

    Directory of Open Access Journals (Sweden)

    Ruirui Huang

    2016-10-01

    Full Text Available DNA methylation plays a crucial role in the regulation of gene expression in eukaryotes. Mushrooms belonging to the phylum Basidiomycota are highly valued for both nutritional and pharmaceutical uses. A growing number of studies have demonstrated the significance of DNA methylation in the development of plants and animals. However, our understanding of DNA methylation in mushrooms is limited. In this study, we identified and conducted comprehensive analyses on DNA methyltransferases (DNMtases in representative species from Basidiomycota and Ascomycota, and obtained new insights into their classification and characterization in fungi. Our results revealed that DNMtases in basidiomycetes can be divided into two classes, the Dnmt1 class and the newly defined Rad8 class. We also demonstrated that the fusion event between the characteristic domains of the DNMtases family and Snf2 family in the Rad8 class is fungi-specific, possibly indicating a functional novelty of Rad8 DNMtases in fungi. Additionally, expression profiles of DNMtases in the edible mushroom Pleurotus ostreatus revealed diverse expression patterns in various organs and developmental stages. For example, DNMtase genes displayed higher expression levels in dikaryons than in monokaryons. Consistent with the expression profiles, we found that dikaryons are more susceptible to the DNA methyltransferase inhibitor 5-azacytidine. Taken together, our findings pinpoint an important role of DNA methylation during the growth of mushrooms and provide a foundation for understanding of DNMtases in basidiomycetes.

  5. Molecular cloning and functional identification of sterol C24-methyltransferase gene from Tripterygium wilfordii

    Directory of Open Access Journals (Sweden)

    Hongyu Guan

    2017-09-01

    Full Text Available Sterol C24-methyltransferase (SMT plays multiple important roles in plant growth and development. SMT1, which belongs to the family of transferases and transforms cycloartenol into 24-methylene cycloartenol, is involved in the biosynthesis of 24-methyl sterols. Here, we report the cloning and characterization of a cDNA encoding a sterol C24-methyltransferase from Tripterygium wilfordii (TwSMT1. TwSMT1 (GenBank access number KU885950 is a 1530 bp cDNA with a 1041 bp open reading frame predicted to encode a 346-amino acid, 38.62 kDa protein. The polypeptide encoded by the SMT1 cDNA was expressed and purified as a recombinant protein from Escherichia coli (E. coli and showed SMT activity. The expression of TwSMT1 was highly up-regulated in T. wilfordii cell suspension cultures treated with methyl jasmonate (MeJA. Tissue expression pattern analysis showed higher expression in the phellem layer compared to the other four organs (leaf, stem, xylem and phloem, which is about ten times that of the lowest expression in leaf. The results are meaningful for the study of sterol biosynthesis of T. wilfordii and will further lay the foundations for the research in regulating both the content of other main compounds and growth and development of T. wilfordii.

  6. Structural insights into methyltransferase KsgA function in 30S ribosomal subunit biogenesis.

    Science.gov (United States)

    Boehringer, Daniel; O'Farrell, Heather C; Rife, Jason P; Ban, Nenad

    2012-03-23

    The assembly of the ribosomal subunits is facilitated by ribosome biogenesis factors. The universally conserved methyltransferase KsgA modifies two adjacent adenosine residues in the 3'-terminal helix 45 of the 16 S ribosomal RNA (rRNA). KsgA recognizes its substrate adenosine residues only in the context of a near mature 30S subunit and is required for the efficient processing of the rRNA termini during ribosome biogenesis. Here, we present the cryo-EM structure of KsgA bound to a nonmethylated 30S ribosomal subunit. The structure reveals that KsgA binds to the 30S platform with the catalytic N-terminal domain interacting with substrate adenosine residues in helix 45 and the C-terminal domain making extensive contacts to helix 27 and helix 24. KsgA excludes the penultimate rRNA helix 44 from adopting its position in the mature 30S subunit, blocking the formation of the decoding site and subunit joining. We suggest that the activation of methyltransferase activity and subsequent dissociation of KsgA control conformational changes in helix 44 required for final rRNA processing and translation initiation.

  7. Structural Insights into Methyltransferase KsgA Function in 30S Ribosomal Subunit Biogenesis*

    Science.gov (United States)

    Boehringer, Daniel; O'Farrell, Heather C.; Rife, Jason P.; Ban, Nenad

    2012-01-01

    The assembly of the ribosomal subunits is facilitated by ribosome biogenesis factors. The universally conserved methyltransferase KsgA modifies two adjacent adenosine residues in the 3′-terminal helix 45 of the 16 S ribosomal RNA (rRNA). KsgA recognizes its substrate adenosine residues only in the context of a near mature 30S subunit and is required for the efficient processing of the rRNA termini during ribosome biogenesis. Here, we present the cryo-EM structure of KsgA bound to a nonmethylated 30S ribosomal subunit. The structure reveals that KsgA binds to the 30S platform with the catalytic N-terminal domain interacting with substrate adenosine residues in helix 45 and the C-terminal domain making extensive contacts to helix 27 and helix 24. KsgA excludes the penultimate rRNA helix 44 from adopting its position in the mature 30S subunit, blocking the formation of the decoding site and subunit joining. We suggest that the activation of methyltransferase activity and subsequent dissociation of KsgA control conformational changes in helix 44 required for final rRNA processing and translation initiation. PMID:22308031

  8. Inhibition of histone methyltransferase EZH2 ameliorates early acute renal allograft rejection in rats.

    Science.gov (United States)

    Li, Long; Zhang, Yi; Xu, Ming; Rong, Ruiming; Wang, Jina; Zhu, Tongyu

    2016-10-26

    Although histone methyltransferases EZH2 has been proved to have significant regulatory effect on the immune rejection after hematopoietic stem cell transplantation, its role in solid-organ transplantation remains uncovered. In this study, we investigate whether histone methylation regulation can impact renal allograft rejection in rat models. Allogeneic rat renal transplantation model (Wistar to Lewis) was established, and the recipients were administrated with EZH2 inhibitor DZNep after transplantation. Renal allografts and peripheral blood were collected on day 5 after transplantation for histological examination and mechanism investigation. We found that inhibition of EZH2 by DZNep after transplantation significantly ameliorated acute rejection (AR), with decreased histological injury and reduced inflammatory infiltration in renal allografts. Attenuation of AR was due to the prohibited activation of alloreactive T cells, the subsequent impaired production of inflammatory cytokines, and also the elevated apoptosis of alloreactive T cells in both renal allografts and periphery. However, inhibition of EZH2 did not increase the regulatory T cells during the AR. Disruption of EZH2 by DZNep suppressed the immune responses of alloreactive T cells and ameliorated AR of renal allografts. This suggests a therapeutic potential of targeting histone methyltransferases EZH2 in treating allograft rejection after solid organ transplantation.

  9. Folate deficiency and aberrant expression of DNA methyltransferase 1 were associated with cervical cancerization.

    Science.gov (United States)

    Wang, Jin-tao; Ding, Ling; Jiang, Shi-Wen; Hao, Junxia; Zhao, Wei-min; Zhou, Quin; Yang, Zuo-kai; Zhang, Li

    2014-01-01

    DNA methyltransferase 1 (DNMT1) plays a significant role in maintaining DNA methylation. Aberrant DNA methylation is a recognized feature of human cancers and folate is directly involved in DNA methylation via one-carbon metabolism. Previous reports also have suggested that folate deficiency was associated with many cancers. The aim of the present study was to evaluate the effect of folate deficiency and aberrant expression of DNA methyltransferase 1 (DNMT1) on cervical cancerization. The expression of DNMT1 protein and mRNA and levels of serum folate were detected in 238 women with a diagnosis of normal cervix (NC,n = 53), cervical intraepithelial neoplasia (CIN I, n = 52; CIN II/III, n = 53), and squamous cell carcinoma of the cervix (SCC; n = 80). In addition, the expression of DNMT1 protein and mRNA was measured in cervical cancer cells (Caski and C33A) treated by different concentration of folate. Serum folate levels decreased and expression levels of DNMT1 protein and mRNA increased gradually with progressive severity of the cervix lesions (Pcancerization. Folate supplement and recovery of aberrant DNA methylation status may offer a new strategy for prevention and therapy of cancers.

  10. Recovery of biological active catechol-O-methyltransferase isoforms from Q-sepharose.

    Science.gov (United States)

    Correia, F F; Santos, F M; Pedro, A Q; Bonifácio, M J; Queiroz, J A; Passarinha, L A

    2014-01-01

    The development of new catechol-O-methyltransferase inhibitors has led to an improvement in the treatment of Parkinson's disease. However, despite the fact that the soluble isoform has been extensively investigated, few studies have been published concerning membrane isoform chromatographic recovery and bioactivity levels. In this work, chromatographic profiles of both catechol-O-methyltransferase isoforms were compared using quaternary amine as a ligand to evaluate its activity levels and recovery rates. Results show that both proteins required different conditions for adsorption; the soluble isoform adsorption was performed at low ionic strength, while the membrane isoform required increasing linear salt gradient. However, the application of 0.5% Triton X-100 promoted membrane isoform adsorption even at low ionic strength. Indeed, chromatographic conditions of both isoforms became similar when detergents were applied. The developed methods also appear to be highly effective in bioactivity recovery, presenting rates of 107% for soluble protein and 67 and 91% for membrane isoform without and with detergents, respectively. The chromatographic strategies with and without detergents resulted in a 4.3- and sevenfold purification, respectively, corresponding to specific activity values of 331 and 496 nmol/h/mg. Thus, the use of Q-sepharose as anion exchanger was effective in the recovery of both enzymes, which is a requirement for further kinetic and pharmacological trials. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Human catechol-O-methyltransferase: Cloning and expression of the membrane-associated form

    Energy Technology Data Exchange (ETDEWEB)

    Bertocci, B.; Miggiano, V.; Da Prada, M.; Dembic, Z.; Lahm, H.W.; Malherbe, P. (F. Hoffmann-La Roche Ltd., Basel (Switzerland))

    1991-02-15

    A cDNA clone for human catechol-O-methyltransferase was isolated from a human hepatoma cell line (Hep G2) cDNA library by hybridization screening with a porcine cDNA probe. The cDNA clone was sequenced and found to have an insert of 1226 nucleotides. The deduced primary structure of hCOMT is composed of 271 amino acid residues with the predicted molecular mass of 30 kDa. At its N terminus it has a hydrophobic segment of 21 amino acid residues that may be responsible for insertion of hCOMT into the endoplasmic reticulum membrane. The primary structure of hCOMT exhibits high homology to the porcine partial cDNA sequence (93%). The deduced amino acid sequence contains two tryptic peptide sequences (T-22, T-33) found in porcine liver catechol-O-methyltransferase (CEMT). The coding region of hCOMT cDNA was placed under the control of the cytomegalovirus promoter to transfect human kidney 293 cells. The recombinant hCOMT was shown by immunoblot analysis to be mainly associated with the membrane fraction. RNA blot analysis revealed one COMT mRNA transcript of 1.4 kilobases in Hep G2 poly(A){sup +} RNA.

  12. Improved radioenzymatic assay for plasma norepinephrine using purified phenylethanolamine n-methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Bowsher, R.R.; Henry, D.P.

    1986-03-01

    Radioenzymatic assays have been developed for catecholamines using either catechol O-methyltransferase (COMT) or phenylethanolamine N-methyltransferase (PNMT). Assays using PNMT are specific for norepinephrine (NE) and require minimal manipulative effort but until now have been less sensitive than the more complex procedures using COMT. The authors report an improved purification scheme for bovine PNMT which has permitted development of an NE assay with dramatically improved sensitivity (0.5 pg), specificity and reproducibility (C.V. < 5%). PNMT was purified by sequential pH 5.0 treatment and dialysis and by column chromatographic procedures using DEAE-Sephacel, Sepharcryl S-200 and Phenyl-Boronate Agarose. Recovery of PNMT through the purification scheme was 50%, while blank recovery was <.001%. NE can be directly quantified in 25 ul of human plasma and an 80 tube assay can be completed within 4 h. The capillary to venous plasma NE gradient was examined in 8 normotensive male subjects. Capillary plasma (NE (211.2 +/- 61.3 pg/ml)) was lower than venous plasma NE (366.6 +/- 92.5 pg/ml) in all subjects (p < 0.005). This difference suggests that capillary (NE) may be a unique indicator of sympathetic nervous system activity in vivo. In conclusion, purification of PNMT has facilitated development of an improved radioenzymatic for NE with significantly improved sensitivity.

  13. Targeting epigenetic regulators for cancer therapy: modulation of bromodomain proteins, methyltransferases, demethylases, and microRNAs.

    Science.gov (United States)

    Gelato, Kathy A; Shaikhibrahim, Zaki; Ocker, Matthias; Haendler, Bernard

    2016-07-01

    Histone deacetylases (HDACs) and DNA methyltransferases (DNMTs) were the first epigenetic targets to be successfully addressed for cancer treatment, but more recently additional families of epigenetic modulators have been the subject of intense research. Potent inhibitors have been identified in several instances and have proven to be invaluable tools for studying these proteins in normal physiology and in disease. Some have now progressed to clinical studies in hematological and solid tumors, and encouraging early results have been reported. This article reviews recent advances regarding the roles of new epigenetic players beyond HDACs and DNMTs in cancer, and discusses the impact of selective chemical probes on unravelling their function. The emerging field of non-coding RNAs (ncRNAs) and ongoing clinical studies with epigenetic drugs and microRNAs (miRNAs) are also addressed. The roles of different epigenetic factors in numerous cancers have been unraveled recently, leading to the initiation of clinical studies. With inhibitors of BET bromodomain proteins, the histone methyltransferases EZH2 and DOT1L, and the histone demethylase LSD1 progressing through clinical trials, and the recognition of the importance of ncRNAs as potential biomarkers and therapeutics, this bears the hope that novel epigenetic therapies will be approved soon.

  14. Structure and Mechanism of the Rebeccamycin Sugar 4'-O-Methyltransferase RebM

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Shanteri; McCoy, Jason G.; Zhang, Changsheng; Bingman, Craig A.; Phillips, Jr., George N.; Thorson, Jon S. (UW)

    2008-12-12

    The 2.65-{angstrom} crystal structure of the rebeccamycin 4'-O-methyltransferase RebM in complex with S-adenosyl-l-homocysteine revealed RebM to adopt a typical S-adenosylmethionine-binding fold of small molecule O-methyltransferases (O-MTases) and display a weak dimerization domain unique to MTases. Using this structure as a basis, the RebM substrate binding model implicated a predominance of nonspecific hydrophobic interactions consistent with the reported ability of RebM to methylate a wide range of indolocarbazole surrogates. This model also illuminated the three putative RebM catalytic residues (His{sup 140/141} and Asp{sup 166}) subsequently found to be highly conserved among sequence-related natural product O-MTases from GC-rich bacteria. Interrogation of these residues via site-directed mutagenesis in RebM demonstrated His{sup 140} and Asp{sup 166} to be most important for catalysis. This study reveals RebM to be a member of the general acid/base-dependent O-MTases and, as the first crystal structure for a sugar O-MTase, may also present a template toward the future engineering of natural product MTases for combinatorial applications.

  15. Protein substrates of the arginine methyltransferase Hmt1 identified by proteome arrays.

    Science.gov (United States)

    Low, Jason K K; Im, Hogune; Erce, Melissa A; Hart-Smith, Gene; Snyder, Michael P; Wilkins, Marc R

    2016-02-01

    Arginine methylation on nonhistone proteins is associated with a number of cellular processes including RNA splicing, protein localization, and the formation of protein complexes. In this manuscript, Saccharomyces cerevisiae proteome arrays carrying 4228 proteins were used with an antimethylarginine antibody to first identify 88 putatively arginine-methylated proteins. By treating the arrays with recombinant arginine methyltransferase Hmt1, 42 proteins were found to be possible substrates of this enzyme. Analysis of the putative arginine-methylated proteins revealed that they were predominantly nuclear or nucleolar in localization, consistent with the localization of Hmt1. Many are involved in known methylarginine-associated functions, such as RNA processing and ribonucleoprotein complex biogenesis, yet others are of newer classes, namely RNA/DNA helicases and tRNA-associated proteins. Using ex vivo methylation and MS/MS, a set of 12 proteins (Brr1, Dia4, Hts1, Mpp10, Mrd1, Nug1, Prp43, Rpa43, Rrp43, Spp381, Utp4, and Npl3), including the RNA helicase Prp43 and tRNA ligases Dia4 and Hts1, were all validated as Hmt1 substrates. Interestingly, the majority of these also had human orthologs, or family members, that have been documented elsewhere to carry arginine methylation. These results confirm arginine methylation as a widespread modification and Hmt1 as the major arginine methyltransferase in the S. cerevisiae cell. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Novel Function of Lysine Methyltransferase G9a in the Regulation of Sox2 Protein Stability.

    Directory of Open Access Journals (Sweden)

    Jae-Young Lee

    Full Text Available G9a is a lysine methyltransferase (KMTase for histone H3 lysine 9 that plays critical roles in a number of biological processes. Emerging evidence suggests that aberrant expression of G9a contributes to tumor metastasis and maintenance of a malignant phenotype in cancer by inducing epigenetic silencing of tumor suppressor genes. Here, we show that G9a regulates Sox2 protein stability in breast cancer cells. When G9a lysine methyltransferase activity was chemically inhibited in the ER(+ breast cancer cell line MCF7, Sox2 protein levels were decreased. In addition, ectopic overexpression of G9a induced accumulation of Sox2. Changes in cell migration, invasion, and mammosphere formation by MCF7 cells were correlated with the activity or expression level of G9a. Ectopic expression of G9a also increased Sox2 protein levels in another ER(+ breast cancer cell line, ZR-75-1, whereas it did not affect Sox2 expression in MDA-MB-231 cells, an ER(- breast cancer cell line, or in glioblastoma cell lines. Furthermore, treatment of mouse embryonic stem cells with a KMT inhibitor, BIX-01294, resulted in a rapid reduction in Sox2 protein expression despite increased Sox2 transcript levels. This finding suggests that G9a has a novel function in the regulation of Sox2 protein stability in a cell type-dependent manner.

  17. Discovery of Potent and Selective Inhibitors for G9a-Like Protein (GLP) Lysine Methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Xiong, Yan; Li, Fengling; Babault, Nicolas; Dong, Aiping; Zeng, Hong; Wu, Hong; Chen, Xin; Arrowsmith, Cheryl H.; Brown, Peter J.; Liu, Jing; Vedadi, Masoud; Jin, Jian

    2017-02-14

    G9a-like protein (GLP) and G9a are highly homologous protein lysine methyltransferases (PKMTs) sharing approximately 80% sequence identity in their catalytic domains. GLP and G9a form a heterodimer complex and catalyze mono- and dimethylation of histone H3 lysine 9 and nonhistone substrates. Although they are closely related, GLP and G9a possess distinct physiological and pathophysiological functions. Thus, GLP or G9a selective small-molecule inhibitors are useful tools to dissect their distinct biological functions. We previously reported potent and selective G9a/GLP dual inhibitors including UNC0638 and UNC0642. Here we report the discovery of potent and selective GLP inhibitors including 4 (MS0124) and 18 (MS012), which are >30-fold and 140-fold selective for GLP over G9a and other methyltransferases, respectively. The cocrystal structures of GLP and G9a in the complex with either 4 or 18 displayed virtually identical binding modes and interactions, highlighting the challenges in structure-based design of selective inhibitors for either enzyme.

  18. A molecular dynamics study on the adsorption of a mussel protein on two different films: Polymer film and a SAM

    Science.gov (United States)

    He, Cunxue; Zhang, Heng; Lin, Cunguo; Wang, Li; Yuan, Shiling

    2017-05-01

    The adhesion of marine life would produce a certain degree of corrosion effect on the hull surface. Shellfish organisms, such as barnacles and mussels, were always used to research the impediment of coating material to protein adsorption. In this work, the adsorbed behaviors of mussel protein on the PDMS and C7F16-SAM surfaces were explored by molecular dynamics (MD) simulations. Simulation results showed that protein was strongly adsorbed onto the hydrophobic surface, as reflected by the large interaction energy; while the adsorption onto the hydrophilic PDMS surface was weak due to two strongly adhered water layers.

  19. Progress Report on SAM Reduced-Order Model Development for Thermal Stratification and Mixing during Reactor Transients

    Energy Technology Data Exchange (ETDEWEB)

    Hu, R. [Argonne National Lab. (ANL), Argonne, IL (United States)

    2017-09-01

    This report documents the initial progress on the reduced-order flow model developments in SAM for thermal stratification and mixing modeling. Two different modeling approaches are pursued. The first one is based on one-dimensional fluid equations with additional terms accounting for the thermal mixing from both flow circulations and turbulent mixing. The second approach is based on three-dimensional coarse-grid CFD approach, in which the full three-dimensional fluid conservation equations are modeled with closure models to account for the effects of turbulence.

  20. Structural Basis for the Methylation of G1405 in 16S rRNA by Aminoglycoside Resistance Methyltransferase Sgm from an Antibiotic Producer: a Diversity of Active Sites in m7G Methyltransferases

    Energy Technology Data Exchange (ETDEWEB)

    Husain, N.; Tkaczuk, K; Tulsidas, S; Kaminska, K; Cubrilo, S; Maravic -Vlahovicek, G; Bujnicki, J; Sivaraman, J

    2010-01-01

    Sgm (Sisomicin-gentamicin methyltransferase) from antibiotic-producing bacterium Micromonospora zionensis is an enzyme that confers resistance to aminoglycosides like gentamicin and sisomicin by specifically methylating G1405 in bacterial 16S rRNA. Sgm belongs to the aminoglycoside resistance methyltransferase (Arm) family of enzymes that have been recently found to spread by horizontal gene transfer among disease-causing bacteria. Structural characterization of Arm enzymes is the key to understand their mechanism of action and to develop inhibitors that would block their activity. Here we report the structure of Sgm in complex with cofactors S-adenosylmethionine (AdoMet) and S-adenosylhomocysteine (AdoHcy) at 2.0 and 2.1 {angstrom} resolution, respectively, and results of mutagenesis and rRNA footprinting, and protein-substrate docking. We propose the mechanism of methylation of G1405 by Sgm and compare it with other m{sup 7}G methyltransferases, revealing a surprising diversity of active sites and binding modes for the same basic reaction of RNA modification. This analysis can serve as a stepping stone towards developing drugs that would specifically block the activity of Arm methyltransferases and thereby re-sensitize pathogenic bacteria to aminoglycoside antibiotics.

  1. Preliminary investigation on the combined effect of S-adenosyl-L-methionine (SAM and oseltamivir on experimental influenza А virus infection in mice

    Directory of Open Access Journals (Sweden)

    Milka M. Mileva

    2016-12-01

    Full Text Available nfluenza is one of the most contageous viral diseases, caused by influenza virus and affects thousands of people every year. The infection causes changes in the intracellular redox balance, increased production of reactive oxygen species, development of antioxidant deficiency and conditions of oxidative stress. Decreased level of gluthatione during flu is responsible for the severe pathology and complications. The purpose of our studies was to follow the effect of the combination S-adenosyl-L-methionine (SAM as a precursor of glutathione and the specific neuraminidase inhibitor oseltamivir in influenza infected mice. SAM was given as a single daily dose of 50,100 and 150 mg/kg, starting from 5 days before infection until day 4th after viral inoculation. Oseltamivir was given in a daily dose of 2.5 mg/kg in two intakes for 5 days, starting from 4th hour before infection. End-point evaluation was 14 day survival rate, avarege survival time, index of protection, and virus titer in lungs. The results showed that application of SAM alone did not have any antiviral prevention. In mice supplemented with oseltamivir only survival rate was 70%, but combination of oseltamivir and SAM in lower doses led to rising of 90% of protection. The present findings suggest that combined therapy of SAM as a precursor of glutathione and the specific inhibitor of inflienza virus replication oseltamivir could be effective on modulation of host defense mechanism(s in low therapeutic doses.

  2. A Standard Analysis Method (SAM) for the automated analysis of polychlorinated biphenyls (PCBs) in soils using the chemical analysis automation (CAA) paradigm: Validation and performance

    Energy Technology Data Exchange (ETDEWEB)

    Rzeszutko, C.; Johnson, C.R.; Monagle, M. [Los Alamos National Lab., NM (United States); Klatt, L.N. [Oak Ridge National Lab., TN (United States)

    1997-11-01

    The Chemical Analysis Automation (CAA) program is developing a standardized modular automation strategy for chemical analysis. In this automation concept, analytical chemistry is performed with modular building blocks that correspond to individual elements of the steps in the analytical process. With a standardized set of behaviors and interactions, these blocks can be assembled in a plug-and-play manner into a complete analysis system. These building blocks, which are referred to as Standard laboratory Modules (SLM), interface to a host control system that orchestrates the entire analytical process, from sample preparation through data interpretation. The integrated system is called a Standard Analysis Method (SAM). A SAM for the automated determination of polychlorinated biphenyls (PCBs) in soils, assembled in a mobile laboratory, is undergoing extensive testing and validation. The SAM consists of the following SLMs: a four-channel Soxhlet extractor, a high-volume concentration, a column clean-up, a gas chromatography, a PCB data-interpretation module, a robot, and a human-computer interface. The SAM is configured to meet the requirements specified in the US Environmental Protection Agency`s (EPA) SW-846 methods 3541/3620A/8082 for the analysis of PCBs in soils. The PCB SAM will be described along with the developmental test plan. Performance data obtained during developmental testing will also be discussed.

  3. Isotopic Composition of Carbon Dioxide Released from Confidence Hills Sediment as Measured by the Sample Analysis at Mars (SAM) Quadrupole Mass Spectrometer

    Science.gov (United States)

    Franz, H. B.; Mahaffy, P. R.; Stern, J.; Archer, P., Jr.; Conrad, P.; Eigenbrode, J.; Freissinet, C.; Glavin, D.; Grotzinger, J. P.; Jones, J.; hide

    2015-01-01

    In October 2014, the Mars Science Laboratory (MSL) "Curiosity" rover drilled into the sediment at the base of Mount Sharp in a location namsed Cionfidence Hills (CH). CH marked the fifth sample pocessed by the Sample Analysis at Mars (SAM) instrument suite since Curiosity arrived in Gale Crater, with previous analyses performed at Rocknest (RN), John Klein (JK), Cumberland (CB), and Windjana (WJ). Evolved gas analysis (EGA) of all samples has indicated H2O as well as O-, C- and S-bearing phases in the samples, often at abundances that would be below the detection limit of the CheMin instrument. By examining the temperatures at which gases are evolved from samples, SAM EGA data can help provide clues to the mineralogy of volatile-bearing phases when their identities are unclear to CheMin. SAM may also detect gases evolved from amorphous material in solid samples, which is not suitable for analysis by CheMin. Finally, the isotopic composition of these gases may suggest possible formation scenarios and relationships between phases. We will discuss C isotope ratios of CO2 evolved from the CH sample as measured with SAM's quadrupole mass spectrometer (QMS) and draw comparisons to samples previously analyzed by SAM.

  4. Search for nitrates on Mars by the Sample Analysis at Mars (SAM) Instrument

    Science.gov (United States)

    Navarro-Gonzalez, Rafael; Stern, Jennifer; Freissinet, Caroline; Franz, Heather; McKay, Christopher; Coll, Patrice; Sutter, Brad; Archer, Doug; McAdam, Amy; Cabane, Michel; Ming, Douglas; Glavin, Daniel; Eigenbrode, Jennifer; Leshin, Laurie; Wong, Michael; Atreya, Sushil; Wray, James; Steele, Andrew; Buch, Arnaud; Prats, Benito

    2014-05-01

    One of the main goals of the Mars Science Laboratory is to determine whether the planet ever had environmental conditions capable of supporting microbial life. Nitrogen is a fundamental element for life, and is present in structural (e.g., proteins), catalytic (e.g., enzymes and ribozymes), energy transfer (e.g., ATP) and information storage (RNA and DNA) bio-molecules. Planetary models suggest that nitrogen was abundant in the early Martian atmosphere as dinitrogen (N2). However, a fraction of N2 has been lost to space by sputtering and photochemical processes [1, 2], impact erosion [3], and chemical oxidation to nitrates [4, 5]. Nitrates produced early in Mars' history by photochemistry may later decompose back into N2 by the current impact flux [6]. It is estimated that the Martian surface could contain soil nitrates at levels of 0.3 wt.% N, if mixed homogenously [6], or a layer of pure NaNO3 of about 3 m thickness [5] distributed globally. Nitrates are a fundamental source for nitrogen for terrestrial microorganisms. Therefore, the detection of soil nitrates is important to assess habitability in the Martian environment. The only previous attempt to search for soil nitrates was by TEGA and the MECA WCL on the Phoenix mission but no evolved N-containing species were detected [7]. Nitrates have been tentatively identified in two Martian meteorites: Nakhla [8] and EETA79001 [9]. SAM is capable of detecting nitrates by their thermal decomposition into nitric oxide, NO. SAM analyzed samples from Rocknest soil and two drill holes located at John Klein (JK) and Cumberland (CB) mudstones in the Sheepbed member of the Yellowknife Bay formation in Gale Crater. There appear to be several peaks associated with the release of m/z 30 in the temperature range from 150° C to 600° C. m/z 30 can be attributed to nitric oxide; however, other possible chemical interferences may be present and are assessed. The origin of nitric oxide is discussed and its thermal evolution is

  5. Conversion of nicotinic acid to trigonelline is catalyzed by N-methyltransferase belonged to motif B′ methyltransferase family in Coffea arabica

    Energy Technology Data Exchange (ETDEWEB)

    Mizuno, Kouichi, E-mail: koumno@akita-pu.ac.jp [Faculty of Bioresource Sciences, Akita Prefectural University, Akita City, Akita 010-0195 (Japan); Matsuzaki, Masahiro [Faculty of Bioresource Sciences, Akita Prefectural University, Akita City, Akita 010-0195 (Japan); Kanazawa, Shiho [Graduate School of Humanities and Sciences, Ochanomizu University, Otsuka, Bunkyo-ku, Tokyo 112-8610 (Japan); Tokiwano, Tetsuo; Yoshizawa, Yuko [Faculty of Bioresource Sciences, Akita Prefectural University, Akita City, Akita 010-0195 (Japan); Kato, Misako [Graduate School of Humanities and Sciences, Ochanomizu University, Otsuka, Bunkyo-ku, Tokyo 112-8610 (Japan)

    2014-10-03

    Graphical abstract: Trigonelline synthase catalyzes the conversion of nicotinic acid to trigonelline. We isolated and characterized trigonelline synthase gene(s) from Coffea arabica. - Highlights: • Trigonelline is a major compound in coffee been same as caffeine is. • We isolated and characterized trigonelline synthase gene. • Coffee trigonelline synthases are highly homologous with coffee caffeine synthases. • This study contributes the fully understanding of pyridine alkaloid metabolism. - Abstract: Trigonelline (N-methylnicotinate), a member of the pyridine alkaloids, accumulates in coffee beans along with caffeine. The biosynthetic pathway of trigonelline is not fully elucidated. While it is quite likely that the production of trigonelline from nicotinate is catalyzed by N-methyltransferase, as is caffeine synthase (CS), the enzyme(s) and gene(s) involved in N-methylation have not yet been characterized. It should be noted that, similar to caffeine, trigonelline accumulation is initiated during the development of coffee fruits. Interestingly, the expression profiles for two genes homologous to caffeine synthases were similar to the accumulation profile of trigonelline. We presumed that these two CS-homologous genes encoded trigonelline synthases. These genes were then expressed in Escherichiacoli, and the resulting recombinant enzymes that were obtained were characterized. Consequently, using the N-methyltransferase assay with S-adenosyl[methyl-{sup 14}C]methionine, it was confirmed that these recombinant enzymes catalyzed the conversion of nicotinate to trigonelline, coffee trigonelline synthases (termed CTgS1 and CTgS2) were highly identical (over 95% identity) to each other. The sequence homology between the CTgSs and coffee CCS1 was 82%. The pH-dependent activity curve of CTgS1 and CTgS2 revealed optimum activity at pH 7.5. Nicotinate was the specific methyl acceptor for CTgSs, and no activity was detected with any other nicotinate derivatives, or

  6. Correlation Does Not Imply Causation: Histone Methyltransferases, but Not Histone Methylation, SET the Stage for Enhancer Activation.

    Science.gov (United States)

    Pollex, Tim; Furlong, Eileen E M

    2017-05-18

    Although H3K4me1 is a pervasive "mark" of enhancers, its functional requirement for enhancer activity remains unclear. In this issue of Molecular Cell, Dorighi et al. (2017) show that in some contexts, the methyltransferase complex, rather than the H3K4me1 mark, is required for gene expression. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. Characterization of cytosine methylated regions and 5-cytosine DNA methyltransferase (Ehmeth) in the protozoan parasite Entamoeba histolytica

    Science.gov (United States)

    Fisher, Ohad; Siman-Tov, Rama; Ankri, Serge

    2004-01-01

    The DNA methylation status of the protozoan parasite Entamoeba histolytica was heretofore unknown. In the present study, we developed a new technique, based on the affinity of methylated DNA to 5-methylcytosine antibodies, to identify methylated DNA in this parasite. Ribosomal DNA and ribosomal DNA circles were isolated by this method and we confirmed the validity of our approach by sodium bisulfite sequencing. We also report the identification and the characterization of a gene, Ehmeth, encoding a DNA methyltransferase strongly homologous to the human DNA methyltransferase 2 (Dnmt2). Immunofluorescence microscopy using an antibody raised against a recombinant Ehmeth showed that Ehmeth is concentrated in the nuclei of trophozoites. The recombinant Ehmeth has a weak but significant methyltransferase activity when E.histolytica genomic DNA is used as substrate. 5-Azacytidine (5-AzaC), an inhibitor of DNA methyltransferase, was used to study in vivo the role of DNA methylation in E.histolytica. Genomic DNA of trophozoites grown with 5-AzaC (23 µM) was undermethylated and the ability of 5-AzaC-treated trophozoites to kill mammalian cells or to cause liver abscess in hamsters was strongly impaired. PMID:14715927

  8. Downregulation of DNA (cytosine-5-)methyltransferase is a late event in NGF-induced PC12 cell differentiation.

    Science.gov (United States)

    Deng, J; Szyf, M

    1999-07-23

    DNA methylation patterns are a critical component of the epigenetic machinery that controls the expression of genetic programs in vertebrates. DNA methyltransferase gene (dnmt1) encodes the enzyme catalyzing the methylation of DNA during replication. We tested the hypothesis that the expression of dnmt1 is regulated with the developmental state of neuronal cells. We show that DNA methyltransferase (Dnmt1) activity is sharply reduced 4 days after induction of differentiation of PC12 cells with NGF. Similarly, the adult brain expresses reduced levels of Dnmt1 activity. We propose that the level of Dnmt1 is downregulated to adjust the activity of the DNA methyltransferase to a different role in mature post-mitotic neurons. Both the abundance of dnmt1 mRNA as well as the Dnmt1 polypeptide are downregulated. Downregulation of dnmt1 parallels other indicators of withdrawal from the cell cycle such as induction of p21, and downregulation of the S phase maker PCNA (proliferating cell nuclear antigen). The temporal pattern of downregulation of dnmt1 in nerve growth factor (NGF)-induced PC12 cells is different from myotube differentiation where downregulation of DNA methyltransferase and demethylation is an early event and was proposed to play a causal role in differentiation. We propose that NGF differentiation of PC12 cells represents a different paradigm of involvement of DNA methylation in terminal differentiation. Copyright 1999 Elsevier Science B.V.

  9. Polymorphisms in O-methyltransferase genes are associated with stover cell wall digestibility in European maize (Zea mays L.)

    DEFF Research Database (Denmark)

    Brenner, Everton A; Zein, Imad; Chen, Yongsheng

    2010-01-01

    Background OMT (O-methyltransferase) genes are involved in lignin biosynthesis, which relates to stover cell wall digestibility. Reduced lignin content is an important determinant of both forage quality and ethanol conversion efficiency of maize stover. Results Variation in genomic sequences codi...

  10. Identification of the methyltransferase targeting C2499 in Deinococcus radiodurans 23S ribosomal RNA

    DEFF Research Database (Denmark)

    Nielsen, Julie Mundus; Flyvbjerg, Karen Freund; Kirpekar, Finn

    2016-01-01

    2499 reported. Using homology search, we identified the open reading frame DR_0049 as the primary candidate gene for the methyltransferase that modifies cytidine 2499. Mass spectrometric analysis demonstrated that recombinantly expressed DR0049 protein methylates E. coli cytidine 2499 both in vitro...

  11. Thiopurine Methyltransferase Enzyme Activity Determination before Treatment of Inflammatory Bowel Disease with Azathioprine: Effect on Cost and Adverse Events

    Directory of Open Access Journals (Sweden)

    Farzana A Sayani

    2005-01-01

    Full Text Available BACKGROUND: Azathioprine (AZA, used to treat inflammatory bowel disease (IBD, is metabolized by thiopurine methyltransferase (TPMT. The accumulation of individual metabolites varies because humans display genetic polymorphism for TPMT expression. Deficiencies in TPMT result in accumulation of toxic metabolites, followed by neutropenia and hepatic inflammation. Concern over acute toxicity frequently leads to under dosing and frequent monitoring tests and visits.

  12. Vagus nerve contributes to the development of steatohepatitis and obesity in phosphatidylethanolamine N-methyltransferase deficient mice

    NARCIS (Netherlands)

    Gao, Xia; van der Veen, Jelske N.; Zhu, Linfu; Chaba, Todd; Ordonez, Marta; Lingrell, Susanne; Koonen, Debby P. Y.; Dyck, Jason R. B.; Gomez-Munoz, Antonio; Vance, Dennis E.; Jacobs, Rene L.

    BACKGROUND & AIMS: Phosphatidylethanolamine N-methyltransferase (PEMT), a liver enriched enzyme, is responsible for approximately one third of hepatic phosphatidylcholine biosynthesis. When fed a high-fat diet (HFD), Pemt(-/-) mice are protected from HF-induced obesity; however, they develop

  13. Thirteen New Patients with Guanidinoacetate Methyltransferase Deficiency and Functional Characterization of Nineteen Novel Missense Variants in the GAMT Gene

    NARCIS (Netherlands)

    Mahmutoglu, S.; Ndika, J.D.T.; Kanhai, W.; de Villemeur, T.B.; Cheillan, D.; Christensen, E.; Dorison, N.; Hannig, V.; Hendriks, Y.M.C.; Hofstede, F.C.; Lion-Francois, L.; Lund, A.M.; Mundy, H.; Pitelet, G.; Raspall-Chaure, M.; Scott-Schwoerer, J.A.; Szakszon, K.; Valayannopoulos, V.; Williams, M.; Salomons, G.S.

    2014-01-01

    Guanidinoacetate methyltransferase deficiency (GAMT-D) is an autosomal recessively inherited disorder of creatine biosynthesis. Creatine deficiency on cranial proton magnetic resonance spectroscopy, and elevated guanidinoacetate levels in body fluids are the biomarkers of GAMT-D. In 74 patients, 50

  14. Catechol-O-methyltransferase gene and obsessive-compulsive symptoms in patients with recent-onset schizophrenia: Preliminary results

    NARCIS (Netherlands)

    Zinkstok, Janneke; van Nimwegen, Lonneke; van Amelsvoort, Therese; de Haan, Lieuwe; Yusuf, Maryan Abdulkadir; Baas, Frank; Linszen, Don

    2008-01-01

    The catechol-O-methyltransferase (COMT) gene is a candidate gene for schizophrenia because of its role in the breakdown of dopamine in the prefrontal cortex. The COMT gene contains a functional polymorphism changing enzyme activity that has been associated with some neuropsychiatric

  15. Properties of the Membrane Binding Component of Catechol-O-methyltransferase Revealed by Atomistic Molecular Dynamics Simulations

    DEFF Research Database (Denmark)

    Orlowski, A.; St-Pierre, J. F.; Magarkar, A.

    2011-01-01

    We used atomistic simulations to study the membrane-bound form of catechol-O-methyltransferase (MB-COMT). In particular we investigated the 26-residue transmembrane a-helical segment of MB-COMT together with the 24-residue fragment that links the transmembrane component to the main protein unit...

  16. No up-regulation of the phosphatidylethanolamine N-methyltransferase pathway and choline production by sex hormones in cats

    NARCIS (Netherlands)

    Valtolina, Chiara; Vaandrager, Arie B; Favier, Robert P; Robben, Joris H; Tuohetahuntila, Maidina; Kummeling, Anne; Jeusette, Isabelle; Rothuizen, Jan

    2015-01-01

    BACKGROUND: Feline hepatic lipidosis (FHL) is a common cholestatic disease affecting cats of any breed, age and sex. Both choline deficiency and low hepatic phosphatidylethanolamine N-methyltransferase (PEMT) activity are associated with hepatic lipidosis (HL) in humans, mice and rats. The PEMT

  17. Thirteen new patients with guanidinoacetate methyltransferase deficiency and functional characterization of nineteen novel missense variants in the GAMT gene

    DEFF Research Database (Denmark)

    Mercimek-Mahmutoglu, Saadet; Ndika, Joseph; Kanhai, Warsha

    2014-01-01

    Guanidinoacetate methyltransferase deficiency (GAMT-D) is an autosomal recessively inherited disorder of creatine biosynthesis. Creatine deficiency on cranial proton magnetic resonance spectroscopy, and elevated guanidinoacetate levels in body fluids are the biomarkers of GAMT-D. In 74 patients 5...

  18. YccW is the m5C methyltransferase specific for 23S rRNA nucleotide 1962

    DEFF Research Database (Denmark)

    Purta, Elzbieta; O'Connor, Michelle; Bujnicki, Janusz M

    2008-01-01

    Methylation at the 5-position of cytosine [m(5)C (5-methylcytidine)] occurs at three RNA nucleotides in Escherichia coli. All these modifications are at highly conserved nucleotides in the rRNAs, and each is catalyzed by its own m(5)C methyltransferase enzyme. Two of the enzymes, RsmB and Rsm......F, are already known and methylate 16S rRNA at nucleotides C967 and C1407, respectively. Here, we report the identity of the third E. coli m(5)C methyltransferase. Analysis of rRNAs by matrix-assisted laser desorption/ionization mass spectrometry showed that inactivation of the yccW gene leads to loss of m(5)C....... coli marginally reduces its growth rate. YccW had previously eluded identification because it displays only limited sequence similarity to the m(5)C methyltransferases RsmB and RsmF and is in fact more similar to known m(5)U (5-methyluridine) RNA methyltransferases. In keeping with the previously...

  19. 5-(4-pyridinyl)-1,3,4-oxadiazole-2-thiol on gold: SAM formation and electroactivity

    Energy Technology Data Exchange (ETDEWEB)

    Paulo, Tercio de F.; Silva, Maria A.S. da; Pinheiro, Solange de O.; Meyer, Emerson; Moreira, Icaro de S.; Diogenes, Izaura C.N. [Universidade Federal do Ceara (UFC), Fortaleza, CE (Brazil). Dept. de Quimica Organica e Inorganica]. E-mail: izaura@dqoi.ufc.br; Pinheiro, Lucidalva S.; Freire, Jose A. [Universidade Federal do Ceara (UFC), Fortaleza, CE (Brazil). Dept. de Fisica; Tanaka, Auro A. [Universidade Federal do Maranhao (UFMA), Sao Luis, MA (Brazil). Dept. de Quimica; Lima Neto, Pedro de [Universidade Federal do Ceara (UFC), Fortaleza, CE (Brazil). Dept. de Quimica Analitica e Fisico Quimica

    2008-07-01

    5-(4-pyridinyl)-1,3,4-oxadiazole-2-thiol (Hpyt) spontaneously adsorbs on gold forming SAMs (self-assembled monolayers) that, based on STM (Scanning Tunneling Microscopy) and electrochemical data, contain pinholes through which [Fe(CN){sub 6}]{sup 4-} and [Ru(NH{sub 3}){sub 6}]{sup 3+} probe molecules access the underlying gold electrode. For the former molecule, the dependence of the faradaic current on the electrolyte solution pH value allowed the evaluation of the surface pKa as 4.2. The thermodynamic parameters {delta}H{sub ads} and {delta}G{sub ads} for the Hpyt adsorption process could be described by the Langmuir model and were calculated as -20.01 and -39.39 kJ mol{sup -1}, respectively. Electrodic redox reaction of cytochrome c metalloprotein was accessed by using the Hpyt SAM with a heterogeneous electron transfer rate constant of 2.29 x 10{sup -3} cm s{sup -1}. (author)

  20. Mars Atmospheric Composition, Isotope Ratios and Seasonal Variations: Overview and Updates of the SAM Measurements at Gale Crater

    Science.gov (United States)

    Niles, Paul

    2014-01-01

    We will summarize the in situ measurements of atmospheric composition and the isotopic ratios of D/H in water, C-13/C-12, O-18/O-16, O-17 / O-16, and C-13 O-18 / C-12 O-16 in carbon dioxide, and Ar-38 / Ar-36, Kr-x / Kr-84, and N-15 / N-14 made in the martian atmosphere at Gale Crater from the Curiosity Rover using the Sample Analysis at Mars (SAM)'s Quadrupole Mass Spectrometer (QMS) and Tunable Laser Spectrometer (TLS). With data over 700 sols since the Curiosity landing, we will discuss evidence and implications for changes on seasonal and other timescales. We will also present results for continued methane and methane enrichment experiments over this time period. Comparison between our measurements in the modern atmosphere and those of martian meteorites like ALH 84001 implies that the martian reservoirs of CO2 and H2O were largely established approximately 4 billion years ago, but that atmospheric loss or surface interaction may be still ongoing.

  1. Functional Analysis of CP2-Like Domain and SAM-Like Domain in TFCP2L1, Novel Pluripotency Factor of Embryonic Stem Cells.

    Science.gov (United States)

    Kim, Chang Min; Jang, Tae-Ho; Park, Hyun Ho

    2016-06-01

    TFCP2L1 is a transcription factor that facilitates establishment and maintenance of pluripotency in embryonic stem cells by forming a complex transcriptional network with other transcription factors (OCT4, SOX2, and NANOG). TFCP2L1 contains two distinct domains, the CP2-like domain at the N-terminus and the SAM-like domain at the C-terminus. In this study, we found that TFCP2L1 is hexamerized in solution via the C-terminal SAM-like domain. We also found that homo-oligomerization of SAM-like domain is dependent on the concentration of the proteins. Finally, we found that TFCP2L1 binds directly to DNA via the N-terminal CP2-like domain.

  2. The Au-S bond and SAM-protein contact in long-range electron transfer of pure and biomimetic metalloproteins via functionalized alkanethiol linkers

    DEFF Research Database (Denmark)

    Chi, Qijin; Ford, Michael J.; Halder, Arnab

    functionalized alkanethiols have emerged as core linkers. We have studied molecular linking in the long-range ET (LRET) processes in detail using electrochemistry, in situ STM and AFM, and electronic structure computations. A focus is the electronic structure of the Au-S link and the SAM packing. We have...... illustrate this by high-resolution in situ STM of straight, branched, and chiral alkanethiols on Au(111)-electrode surfaces. We discuss next LRET of two SAM immobilized multi-copper enzymes, nitrite reductase and laccase, mapped to single-molecule resolution by in situ STM and AFM. The voltammetry...... degree of electronic structure sophistication. At the molecular scale this requirement is met by NPs of the renownedmixed-valence Prussian Blue (PB) assembled on Au(111)-electrode surfaces via functionalized alkanethiols. PBNP SAMs show LRET comparable to metalloproteins. Alkanethiols with different...

  3. The Detection of Evolved Oxygen from the Rocknest Eolian Bedform Material by the Sample Analysis at Mars(SAM) instrument at the Mars Curiosity Landing Site

    Science.gov (United States)

    Sutter, B.; Archer, D.; Ming, D.; Eigenbrode, J. L.; Franz, H.; Glavin, D. P.; McAdam, A.; Mahaffy, P.; Stern, J.; Navarro-Gonzalex, R.; hide

    2013-01-01

    The Sample Analysis at Mars (SAM) instrument onboard the Curiosity rover detected an O2 gas release from the Rocknest eolain bedform (Fig. 1). The detection of perchlorate (ClO4-) by the Mars Phoenix Lander s Wet Chemistry Laboratory (WCL) [1] suggests that perchlorate is a possible candidate for evolved O2 release detected by SAM. The perchlorate would also serve as a source of chlorine in the chlorinated hydrocarbons detected by the SAM quadrupole mass spectrometer (QMS) and gas chromatography/mass spectrometer (GCMS) [2,3]. Chlorates (ClO3-) [4,5] and/or superoxides [6] may also be sources of evolved O2 from the Rocknest materials. The work objectives are to 1) evaluate the O2 release temperatures from Rocknest materials, 2) compare these O2 release temperatures with a series of perchlorates and chlorates, and 3) evaluate superoxide O2- sources and possible perchlorate interactions with other Rocknest phases during QMS analysis.

  4. Methylation of sulfhydryl groups: a new function for a family of small molecule plant O-methyltransferases

    Science.gov (United States)

    Coiner, Heather; Schröder, Gudrun; Wehinger, Elke; Liu, Chang-Jun; Noel, Joseph P.; Schwab, Wilfried; Schröder, Joachim

    2010-01-01

    Summary In plants, type I and II S-adenosyl-L-methionine-dependent O-methyltransferases (OMTs) catalyze most hydroxyl group methylations of small molecules. A homology-based RT-PCR strategy using Catharanthus roseus (Madagascar periwinkle) RNA previously identified six new type I plant OMT family members. We now describe the molecular and biochemical characterization of a seventh protein. It shares 56–58% identity with caffeic acid OMTs (COMTs), but it failed to methylate COMT substrates, and had no activity with flavonoids. However, the in vitro incubations revealed unusually high background levels without added substrates. A search for the responsible component revealed that the enzyme methylated dithiothreitol (DTT), the reducing agent added for enzyme stabilization. Unexpectedly, product analysis revealed that the methylation occurred on a sulfhydryl moiety, not on a hydroxyl group. Analysis of 34 compounds indicated a broad substrate range, with a preference for small hydrophobic molecules. Benzene thiol (Km 220 μM) and furfuryl thiol (Km 60 μM) were the best substrates (6–7-fold better than DTT). Small isosteric hydrophobic substrates with hydroxyl groups, like phenol and guaiacol, were also methylated, but the activities were at least 5-fold lower than with thiols. The enzyme was named C. roseus S-methyltransferase 1 (CrSMT1). Models based on the COMT crystal structure suggest that S-methylation is mechanistically identical to O-methylation. CrSMT1 so far is the only recognized example of an S-methyltransferase in this protein family. Its properties indicate that a few changes in key residues are sufficient to convert an OMT into a S-methyltransferase (SMT). Future functional investigations of plant methyltransferases should consider the possibility that the enzymes may direct methylation at sulfhydryl groups. PMID:16623883

  5. MicroRNA-29a Alleviates Bile Duct Ligation Exacerbation of Hepatic Fibrosis in Mice through Epigenetic Control of Methyltransferases

    Directory of Open Access Journals (Sweden)

    Ya-Ling Yang

    2017-01-01

    Full Text Available MicroRNA-29 (miR-29 is found to modulate hepatic stellate cells’ (HSCs activation and, thereby, reduces liver fibrosis pathogenesis. Histone methyltransferase regulation of epigenetic reactions reportedly participates in hepatic fibrosis. This study is undertaken to investigate the miR-29a regulation of the methyltransferase signaling and epigenetic program in hepatic fibrosis progression. miR-29a transgenic mice (miR-29aTg mice and wild-type littermates were subjected to bile duct-ligation (BDL to develop cholestatic liver fibrosis. Primary HSCs were transfected with a miR-29a mimic and antisense inhibitor. Profibrogenic gene expression, histone methyltransferases and global genetic methylation were probed with real-time quantitative RT-PCR, immunohistochemical stain, Western blot and ELISA. Hepatic tissue in miR-29aTg mice displayed weak fibrotic matrix as evidenced by Sirius Red staining concomitant with low fibrotic matrix collagen 1α1 expression within affected tissues compared to the wild-type mice. miR-29a overexpression reduced the BDL exaggeration of methyltransferases, DNMT1, DNMT3b and SET domain containing 1A (SET1A expression. It also elevated phosphatase and tensin homolog deleted on chromosome 10 (PTEN signaling within liver tissue. In vitro, miR-29a mimic transfection lowered collagen 1α1, DNMT1, DNMT3b and SET1A expression in HSCs. Gain of miR-29a signaling resulted in DNA hypomethylation and high PTEN expression. This study shines a new light on miR-29a inhibition of methyltransferase, a protective effect to maintain the DNA hypomethylation state that decreases fibrogenic activities in HSC. These robust analyses also highlight the miR-29a regulation of epigenetic actions to ameliorate excessive fibrosis during cholestatic liver fibrosis development.

  6. Targeted DNA methylation in pericentromeres with genome editing-based artificial DNA methyltransferase.

    Science.gov (United States)

    Yamazaki, Taiga; Hatano, Yu; Handa, Tetsuya; Kato, Sakiko; Hoida, Kensuke; Yamamura, Rui; Fukuyama, Takashi; Uematsu, Takayuki; Kobayashi, Noritada; Kimura, Hiroshi; Yamagata, Kazuo

    2017-01-01

    To study the impact of epigenetic changes on biological functions, the ability to manipulate the epigenetic status of certain genomic regions artificially could be an indispensable technology. "Epigenome editing" techniques have gradually emerged that apply TALE or CRISPR/Cas9 technologies with various effector domains isolated from epigenetic code writers or erasers such as DNA methyltransferase, 5-methylcytosine oxidase, and histone modification enzymes. Here we demonstrate that a TALE recognizing a major satellite, consisting of a repeated sequence in pericentromeres, could be fused with the bacterial CpG methyltransferase, SssI. ChIP-qPCR assays demonstrated that the fusion protein TALMaj-SssI preferentially bound to major chromosomal satellites in cultured cell lines. Then, TALMaj-SssI was expressed in fertilized mouse oocytes with hypomethylated major satellites (10-20% CpG islands). Bisulfite sequencing revealed that the DNA methylation status was increased specifically in major satellites (50-60%), but not in minor satellites or other repeat elements, such as Intracisternal A-particle (IAP) or long interspersed nuclear elements-1 (Line1) when the expression level of TALMaj-SssI is optimized in the cell. At a microscopic level, distal ends of chromosomes at the first mitotic stage were dramatically highlighted by the mCherry-tagged methyl CpG binding domain of human MBD1 (mCherry-MBD-NLS). Moreover, targeted DNA methylation to major satellites did not interfere with kinetochore function during early embryonic cleavages. Co-injection of dCas9 fused with SssI and guide RNA (gRNA) recognizing major satellite sequences enabled increment of the DNA methylation in the satellites, but a few off-target effects were also observed in minor satellites and retrotransposons. Although CRISPR can be applied instead of the TALE system, technical improvements to reduce off-target effects are required. We have demonstrated a new method of introducing DNA methylation without

  7. Targeted DNA methylation in pericentromeres with genome editing-based artificial DNA methyltransferase.

    Directory of Open Access Journals (Sweden)

    Taiga Yamazaki

    Full Text Available To study the impact of epigenetic changes on biological functions, the ability to manipulate the epigenetic status of certain genomic regions artificially could be an indispensable technology. "Epigenome editing" techniques have gradually emerged that apply TALE or CRISPR/Cas9 technologies with various effector domains isolated from epigenetic code writers or erasers such as DNA methyltransferase, 5-methylcytosine oxidase, and histone modification enzymes. Here we demonstrate that a TALE recognizing a major satellite, consisting of a repeated sequence in pericentromeres, could be fused with the bacterial CpG methyltransferase, SssI. ChIP-qPCR assays demonstrated that the fusion protein TALMaj-SssI preferentially bound to major chromosomal satellites in cultured cell lines. Then, TALMaj-SssI was expressed in fertilized mouse oocytes with hypomethylated major satellites (10-20% CpG islands. Bisulfite sequencing revealed that the DNA methylation status was increased specifically in major satellites (50-60%, but not in minor satellites or other repeat elements, such as Intracisternal A-particle (IAP or long interspersed nuclear elements-1 (Line1 when the expression level of TALMaj-SssI is optimized in the cell. At a microscopic level, distal ends of chromosomes at the first mitotic stage were dramatically highlighted by the mCherry-tagged methyl CpG binding domain of human MBD1 (mCherry-MBD-NLS. Moreover, targeted DNA methylation to major satellites did not interfere with kinetochore function during early embryonic cleavages. Co-injection of dCas9 fused with SssI and guide RNA (gRNA recognizing major satellite sequences enabled increment of the DNA methylation in the satellites, but a few off-target effects were also observed in minor satellites and retrotransposons. Although CRISPR can be applied instead of the TALE system, technical improvements to reduce off-target effects are required. We have demonstrated a new method of introducing DNA

  8. Thermal Reactivity of Organic Molecules in the Presence of Chlorates and Perchlorates and the Quest for Organics on Mars with the SAM Experiment Onboard the Curiostiy Rover

    Science.gov (United States)

    Szopa, Cyril; Millan, Maeva; Buch, Arnaud; Belmahdi, Imene; Coll, Patrice; Glavin, Daniel P.; Freissinet, Caroline; Eigenbrode, Jennifer; archer, doug; sutter, brad; Summons, Roger; Navarro-Gonzalez, Rafael; Mahaffy, Paul; cabane, Michel

    2016-10-01

    One of the main objectives of the Sample Analysis at Mars (SAM) experiment is the in situ molecular analysis of gases evolving from solid samples collected by Curiosity when they are heated up to ~850°C. With this aim SAM uses a gas-chromatograph coupled to a mass spectrometer (GC-MS) able to detect and identify both inorganic and organic molecules released by the samples.During the pyrolysis, chemical reactions occur between oxychlorines, probably homogeneously distributed at Mars's surface, and organic compounds SAM seeks for. This was confirmed by the first chlorohydrocarbons (chloromethane and di- and trichloromethane) detected by SAM that were entirely attributed to reaction products occurring between these oxychlorines and organics from instrument background. But SAM also detected in the Sheepbed mudstone of Gale crater, chloroalkanes produced by reaction between oxychlorines and Mars indigenous organics, proving for the first time the presence of organics in the soil of Mars. However, the identification of the molecules at the origin of these chloroalkanes is much more difficult due to the complexity of the reactivity occurring during the sample pyrolysis. If a first study has already been done recently with this aim, it was relatively limited in terms of parameters investigated.This is the reason why, we performed a systematic study in the laboratory to help understanding the influence of oxychlorines on organic matter during pyrolysis. With this aim, different organic compounds from various chemical families (e.g. amino and carboxylic acids) mixed with different perchlorates and chlorates, in concentrations compatible with the Mars soil from estimations done with SAM measurements, were pyrolyzed under SAM like conditions. The products of reaction were analyzed and identified by GC-MS in order to show a possible correlation between them and the parent molecule. Different parameters were tested for the pyrolysis to evaluate their potential influence on the

  9. SAM Technical Review Committee Final Report: Summary and Key Recommendations from the Onsite TRC Meeting Held April 22-23, 2013

    Energy Technology Data Exchange (ETDEWEB)

    Blair, N.; Dobos, S.; Janzou, S.; Gilman, P.; Freeman, J.; Kaffine, L.

    2013-08-01

    The System Advisor Model (SAM) is a broad and robust set of models and frameworks for analyzing both system performance and system financing. It does this across a range of technologies dominated by solar technologies including photovoltaics (PV) and concentrated solar power (CSP). The U.S. Department of Energy (DOE) Solar Energy Technology Program requested the SAM development team to review the photovoltaic performance modeling with the development community and specifically, with the independent engineering community. The report summarizes the major effort for this technical review committee (TRC).

  10. Miniaturization of High-Throughput Epigenetic Methyltransferase Assays with Acoustic Liquid Handling.

    Science.gov (United States)

    Edwards, Bonnie; Lesnick, John; Wang, Jing; Tang, Nga; Peters, Carl

    2016-02-01

    Epigenetics continues to emerge as an important target class for drug discovery and cancer research. As programs scale to evaluate many new targets related to epigenetic expression, new tools and techniques are required to enable efficient and reproducible high-throughput epigenetic screening. Assay miniaturization increases screening throughput and reduces operating costs. Echo liquid handlers can transfer compounds, samples, reagents, and beads in submicroliter volumes to high-density assay formats using only acoustic energy-no contact or tips required. This eliminates tip costs and reduces the risk of reagent carryover. In this study, we demonstrate the miniaturization of a methyltransferase assay using Echo liquid handlers and two different assay technologies: AlphaLISA from PerkinElmer and EPIgeneous HTRF from Cisbio. © 2015 Society for Laboratory Automation and Screening.

  11. Identification of the methyltransferase targeting C2499 in Deinococcus radiodurans 23S ribosomal RNA

    DEFF Research Database (Denmark)

    Nielsen, Julie Mundus; Flyvbjerg, Karen Freund; Kirpekar, Finn

    2016-01-01

    The bacterium Deinococcus radiodurans-like all other organisms-introduces nucleotide modifications into its ribosomal RNA. We have previously found that the bacterium contains a Carbon-5 methylation on cytidine 2499 of its 23S ribosomal RNA, which is so far the only modified version of cytidine...... and in vivo. We also inactivated the DR_0049 gene in D. radiodurans through insertion of a chloramphenicol resistance cassette. This resulted in complete absence of the cytidine 2499 methylation, which all together demonstrates that DR_0049 encodes the methyltransferase producing m(5)C2499 in D. radiodurans...... 23S rRNA. Growth experiments disclosed that inactivation of DR_0049 is associated with a severe growth defect, but available ribosome structures show that cytidine 2499 is positioned very similar in D. radiodurans harbouring the modification and E. coli without the modification. Hence...

  12. Protective immunization against visceral leishmaniasis using Leishmania sterol 24-c-methyltransferase formulated in adjuvant.

    Science.gov (United States)

    Goto, Yasuyuki; Bogatzki, Lisa Y; Bertholet, Sylvie; Coler, Rhea N; Reed, Steven G

    2007-10-16

    We present here the identification and characterization of Leishmania sterol 24-c-methyltransferase (SMT), as well as data on protection of mice immunized with this Ag formulated in MPL-SE. Serological evaluation revealed that SMT is recognized by VL patients. C57BL/6 mice immunized with this vaccine candidate plus MPL-SE showed Ag-specific Th1 immune responses characterized by robust production of IFN-gamma upon specific Ag re-exposure in vitro. Upon challenge with L. infantum, mice immunized with SMT plus MPL-SE showed significant lower parasite burdens in both spleens and livers compared with non-immunized mice or mice injected with adjuvant alone. The results indicate that SMT/MPL-SE can be an effective vaccine candidate for use against VL.

  13. DNA methyltransferase 1 and DNA methylation patterning contribute to germinal center B-cell differentiation

    DEFF Research Database (Denmark)

    Shaknovich, Rita; Cerchietti, Leandro; Tsikitas, Lucas

    2011-01-01

    The phenotype of germinal center (GC) B cells includes the unique ability to tolerate rapid proliferation and the mutagenic actions of activation induced cytosine deaminase (AICDA). Given the importance of epigenetic patterning in determining cellular phenotypes, we examined DNA methylation...... and the role of DNA methyltransferases in the formation of GCs. DNA methylation profiling revealed a marked shift in DNA methylation patterning in GC B cells versus resting/naive B cells. This shift included significant differential methylation of 235 genes, with concordant inverse changes in gene expression...... affecting most notably genes of the NFkB and MAP kinase signaling pathways. GC B cells were predominantly hypomethylated compared with naive B cells and AICDA binding sites were highly overrepresented among hypomethylated loci. GC B cells also exhibited greater DNA methylation heterogeneity than naive B...

  14. Coupled selection of protein solubility in E. coli using uroporphyrinogen III methyltransferase as red fluorescent reporter.

    Science.gov (United States)

    Wang, Zhenzhen; Yan, Hanwei; Li, Si; Zhang, Kuanliang; Cheng, Beijiu; Fan, Jun

    2014-09-30

    Uroporphyrinogen III methyltransferase (UMT) is a novel reporter owing to the catalytic products accumulated in cells emitting red florescence. Overexpression of UMT confers resistance of the Escherichia coli cells to potassium tellurite that inhibits cell growth. In this study, we applied UMT reporter for monitoring protein solubility of MBP or TEV protease variants under different expression conditions, as well as 12 maize proteins with either the designed linker or N-terminal SUMO tag. Effects of five enzymes involved in heme and siroheme biosynthesis on the reporter were also investigated. With increasing concentrations of potassium tellurite, colony numbers of the mixed cells expressing the selected five proteins with different solubility were decreased, but colonies displaying red fluorescence was identified to be produced the protein with relatively high solubility. The developed UMT reporter system is sensitive for monitoring protein solubility based on coupled fluorescence and chemical selection. Copyright © 2014 Elsevier B.V. All rights reserved.

  15. Adding a Lysine Mimic in the Design of Potent Inhibitors of Histone Lysine Methyltransferases

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Yanqi; Ganesh, Thota; Horton, John R.; Spannhoff, Astrid; Liu, Jin; Sun, Aiming; Zhang, Xing; Bedford, Mark T.; Shinkai, Yoichi; Snyder, James P.; Cheng, Xiaodong (Emory); (Kyoto); (Texas)

    2010-07-19

    Dynamic histone lysine methylation involves the activities of modifying enzymes (writers), enzymes removing modifications (erasers), and readers of the histone code. One common feature of these activities is the recognition of lysines in methylated and unmethylated states, whether they are substrates, reaction products, or binding partners. We applied the concept of adding a lysine mimic to an established inhibitor (BIX-01294) of histone H3 lysine 9 methyltransferases G9a and G9a-like protein by including a 5-aminopentyloxy moiety, which is inserted into the target lysine-binding channel and becomes methylated by G9a-like protein, albeit slowly. The compound enhances its potency in vitro and reduces cell toxicity in vivo. We suggest that adding a lysine or methyl-lysine mimic should be considered in the design of small-molecule inhibitors for other methyl-lysine writers, erasers, and readers.

  16. Structural and Functional Analyses of a Conserved Hydrophobic Pocket of Flavivirus Methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    H Dong; L Liu; G Zou; Y Zhao; Z Li; S Lim; P Shi; H Li

    2011-12-31

    The flavivirus methyltransferase (MTase) sequentially methylates the N7 and 2'-O positions of the viral RNA cap (GpppA-RNA {yields} m(7)GpppA-RNA {yields} m(7)GpppAm-RNA), using S-adenosyl-l-methionine (AdoMet) as a methyl donor. We report here that sinefungin (SIN), an AdoMet analog, inhibits several flaviviruses through suppression of viral MTase. The crystal structure of West Nile virus MTase in complex with SIN inhibitor at 2.0-{angstrom} resolution revealed a flavivirus-conserved hydrophobic pocket located next to the AdoMet-binding site. The pocket is functionally critical in the viral replication and cap methylations. In addition, the N7 methylation efficiency was found to correlate with the viral replication ability. Thus, SIN analogs with modifications that interact with the hydrophobic pocket are potential specific inhibitors of flavivirus MTase.

  17. Brain Histamine -Methyltransferase as a Possible Target of Treatment for Methamphetamine Overdose

    Directory of Open Access Journals (Sweden)

    Junichi Kitanaka

    2016-01-01

    Full Text Available Stereotypical behaviors induced by methamphetamine (METH overdose are one of the overt symptoms of METH abuse, which can be easily assessed in animal models. Currently, there is no successful treatment for METH overdose. There is increasing evidence that elevated levels of brain histamine can attenuate METH-induced behavioral abnormalities, which might therefore constitute a novel therapeutic treatment for METH abuse and METH overdose. In mammals, histamine N -methyltransferase (HMT is the sole enzyme responsible for degrading histamine in the brain. Metoprine, one of the most potent HMT inhibitors, can cross the blood-brain barrier and increase brain histamine levels by inhibiting HMT. Consequently, this compound can be a candidate for a prototype of drugs for the treatment of METH overdose.

  18. Suz12 is essential for mouse development and for EZH2 histone methyltransferase activity

    DEFF Research Database (Denmark)

    Pasini, Diego; Bracken, Adrian P; Jensen, Michael R

    2004-01-01

    SUZ12 is a recently identified Polycomb group (PcG) protein, which together with EZH2 and EED forms different Polycomb repressive complexes (PRC2/3). These complexes contain histone H3 lysine (K) 27/9 and histone H1 K26 methyltransferase activity specified by the EZH2 SET domain. Here we show...... that mice lacking Suz12, like Ezh2 and Eed mutant mice, are not viable and die during early postimplantation stages displaying severe developmental and proliferative defects. Consistent with this, we demonstrate that SUZ12 is required for proliferation of cells in tissue culture. Furthermore, we demonstrate...... that SUZ12 is essential for the activity and stability of the PRC2/3 complexes in mouse embryos, in tissue culture cells and in vitro. Strikingly, Suz12-deficient embryos show a specific loss of di- and trimethylated H3K27, demonstrating that Suz12 is indeed essential for EZH2 activity in vivo...

  19. Cancer stem cell overexpression of nicotinamide N-methyltransferase enhances cellular radiation resistance

    DEFF Research Database (Denmark)

    D’Andrea, Filippo P.; Safwat, Akmal; Kassem, Moustapha

    2011-01-01

    validated with q-RT-PCR using TaqMan probes. ResultsThe CE8 clone was more radiation resistant than the BB3 clone. From a pool of 15 validated genes with altered expression in the CE8 clone, we found the enzyme nicotinamide N-methyltransferase (NNMT) more than 5-fold upregulated. In-depth pathway analysis...... found the genes involved in cancer, proliferation, DNA repair and cell death. ConclusionsThe higher radiation resistance in clone CE8 is likely due to NNMT overexpression. The higher levels of NNMT could affect the cellular damage resistance through depletion of the accessible amounts of nicotinamide......, which is a known inhibitor of cellular DNA repair mechanisms....

  20. Structure-based drug design of catechol-O-methyltransferase inhibitors for CNS disorders.

    Science.gov (United States)

    Ma, Zhiguo; Liu, Hongming; Wu, Baojian

    2014-03-01

    Catechol-O-methyltransferase (COMT) is of great importance in pharmacology because it catalyzes the metabolism (methylation) of endogenous and xenobiotic catechols. Moreover, inhibition of COMT is the drug target in the management of central nervous system (CNS) disorders such as Parkinson's disease due to its role in regulation of the dopamine level in the brain. The X-ray crystal structures for COMT have been available since 1994. The active sites for cofactor and substrate/inhibitor binding are well resolved to an atomic level, providing valuable insights into the catalytic mechanisms as well as the role of magnesium ions in catalysis. Determination of how the substrates/inhibitors bind to the protein leads to a structure-based approach that has resulted in potent and selective inhibitors. This review focuses on the design of two types of inhibitors (nitrocatechol-type and bisubstrate inhibitors) for COMT using the protein structures. © 2013 The British Pharmacological Society.

  1. Catechol-O-methyltransferase inhibits colorectal cancer cell proliferation and invasion.

    Science.gov (United States)

    Wu, Wenming; Wu, Qiao; Hong, Xiafei; Xiong, Guangbing; Xiao, Yi; Zhou, Jiaolin; Wang, Wenze; Wu, Huanwen; Zhou, Li; Song, Wei; Dai, Hongmei; Qiu, Huizhong; Zhao, Yupei

    2015-01-01

    Catechol-O-methyltransferase (COMT) has been reported as an important molecule in various types of cancers. The biological function of COMT in colorectal cancer (CRC) has not yet been fully investigated. We constructed a transient transfection of a CRC cell lines to up- and downregulate COMT expression level and tested the proliferative, invasion ability in vitro. We also constructed a stable transduced CRC cell line and conducted tumor-forming capacity experiment in mouse xenograft model in vivo. In vitro experiment showed that COMT inhibited the cell proliferation by regulating p-Akt, PTEN and inhibited G1 to S phase transition by regulating p53, p27, and cyclinD1. COMT inhibited invasion by regulating E-cadherin. In vivo experiment showed decreased tumor growth in COMT overexpressing cell line. COMT has tumor-suppressive functions for CRC cell lines in vitro and in vivo experiments. Copyright © 2015 IMSS. Published by Elsevier Inc. All rights reserved.

  2. Catechol-O-methyltransferase genotype and response to Compensatory Cognitive Training in outpatients with schizophrenia.

    Science.gov (United States)

    Burton, Cynthia Z; Vella, Lea; Kelsoe, John R; Bilder, Robert M; Twamley, Elizabeth W

    2015-06-01

    The catechol-O-methyltransferase (COMT) ValMet polymorphism is associated with cognitive functioning in schizophrenia and may predict cognitive training outcomes. This study aimed to explore the contribution of COMT genotype in predicting improvement following Compensatory Cognitive Training (CCT). We conducted mixed factorial analysis of variance to examine COMT genotype as a predictor of response to CCT (i.e. improved cognitive performance) in 41 participants with schizophrenia-spectrum disorders. We also explored the effect of CCT treatment and COMT genotype on psychiatric symptom severity, functional capacity, and subjective quality of life. Met carrier status did not predict CCT treatment outcomes. COMT genotype may exert only modest effects on cognitive training response. Further research with larger samples is needed to establish genetic predictors of response to cognitive training.

  3. Role of the EZH2 histone methyltransferase as a therapeutic target in cancer.

    Science.gov (United States)

    Italiano, Antoine

    2016-09-01

    Besides being a genetic disease, cancer is also an epigenetic disease. The histone methyltransferase EZH2 is the catalytic subunit of PRC2, a highly conserved protein complex that regulates gene expression by methylating lysine 27 on histone H3. Given its role in tumorigenesis and its prognostic value in several tumor types, this protein appears a relevant therapeutic target. This review focuses on the preclinical and preliminary clinical results of studies investigating EZH2 inhibitors in human malignancies. These emerging data suggest that EZH2 inhibitors represent a very promising class of drugs, which will probably have a major impact on improving outcome and reducing toxicity for patients with indolent and aggressive B-cell lymphomas and other specific solid tumors. Copyright © 2016. Published by Elsevier Inc.

  4. The Sm protein methyltransferase PRMT5 is not required for primordial germ cell specification in mice.

    Science.gov (United States)

    Li, Ziwei; Yu, Juehua; Hosohama, Linzi; Nee, Kevin; Gkountela, Sofia; Chaudhari, Sonal; Cass, Ashley A; Xiao, Xinshu; Clark, Amander T

    2015-03-12

    PRMT5 is a type II protein arginine methyltransferase with roles in stem cell biology, reprograming, cancer and neurogenesis. During embryogenesis in the mouse, it was hypothesized that PRMT5 functions with the master germline determinant BLIMP1 to promote primordial germ cell (PGC) specification. Using a Blimp1-Cre germline conditional knockout, we discovered that Prmt5 has no major role in murine germline specification, or the first global epigenetic reprograming event involving depletion of cytosine methylation from DNA and histone H3 lysine 9 dimethylation from chromatin. Instead, we discovered that PRMT5 functions at the conclusion of PGC reprograming I to promote proliferation, survival and expression of the gonadal germline program as marked by MVH. We show that PRMT5 regulates gene expression by promoting methylation of the Sm spliceosomal proteins and significantly altering the spliced repertoire of RNAs in mammalian embryonic cells and primordial cells. © 2014 The Authors.

  5. Multi-site-specific 16S rRNA methyltransferase RsmF from Thermus thermophilus

    DEFF Research Database (Denmark)

    Demirci, Hasan; Larsen, Line H G; Hansen, Trine

    2010-01-01

    Cells devote a significant effort toward the production of multiple modified nucleotides in rRNAs, which fine tune the ribosome function. Here, we report that two methyltransferases, RsmB and RsmF, are responsible for all four 5-methylcytidine (m(5)C) modifications in 16S rRNA of Thermus...... thermophilus. Like Escherichia coli RsmB, T. thermophilus RsmB produces m(5)C967. In contrast to E. coli RsmF, which introduces a single m(5)C1407 modification, T. thermophilus RsmF modifies three positions, generating m(5)C1400 and m(5)C1404 in addition to m(5)C1407. These three residues are clustered near...

  6. Design, synthesis, and protein methyltransferase activity of a unique set of constrained amine containing compounds.

    Science.gov (United States)

    Zhou, Hao; Che, Xin; Bao, Guochen; Wang, Na; Peng, Li; Barnash, Kimberly D; Frye, Stephen V; James, Lindsey I; Bai, Xu

    2016-09-15

    Epigenetic alterations relate to various human diseases, and developing inhibitors of Kme regulatory proteins is considered to be a new frontier for drug discovery. We were inspired by the known multicyclic ligands, UNC669 and UNC926, which are the first reported small molecule ligands for a methyl-lysine binding domain. We hypothesized that reducing the conformational flexibility of the key amine moiety of UNC669 would result in a unique set of ligands. Twenty-five novel compounds containing a fused bi- or tricyclic amine or a spirocyclic amine were designed and synthesized. To gauge the potential of these amine-containing compounds to interact with Kme regulatory proteins, the compounds were screened against a panel of 24 protein methyltransferases. Compound 13 was discovered as a novel scaffold that interacts with SETD8 and could serve as a starting point for the future development of PKMT inhibitors. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. Expression, purification, and characterization of the protein repair l-isoaspartyl methyltransferase from Arabidopsis thaliana.

    Science.gov (United States)

    Thapar, N; Clarke, S

    2000-11-01

    Protein l-isoaspartate (d-aspartate) O-methyltransferase (EC 2.1.1. 77) is a repair enzyme that methylates abnormal l-isoaspartate residues in proteins which arise spontaneously as a result of aging. This enzyme initiates their conversion back into the normal l-aspartate form by a methyl esterification reaction. Previously, partial cDNAs of this enzyme were isolated from the higher plant Arabidopsis thaliana. In this study, we report the cloning and expression of a full-length cDNA of l-isoaspartyl methyltransferase from A. thaliana into Escherichia coli under the P(BAD) promoter, which offers a high level of expression under a tight regulatory control. The enzyme is found largely in the soluble fraction. We purified this recombinant enzyme to homogeneity using a series of steps involving DEAE-cellulose, gel filtration, and hydrophobic interaction chromatographies. The homogeneous enzyme was found to have maximum activity at 45 degrees C and a pH optimum from 7 to 8. The enzyme was found to have a wide range of affinities for l-isoaspartate-containing peptides and displayed relatively poor reactivity toward protein substrates. The best methyl-accepting substrates were KASA-l-isoAsp-LAKY (K(m) = 80 microM) and VYP-l-isoAsp-HA (K(m) = 310 microM). We also expressed the full-length form and a truncated version of this enzyme (lacking the N-terminal 26 amino acid residues) in E. coli under the T7 promoter. Both the full-length and the truncated forms were active, though overexpression of the truncated enzyme led to a complete loss of activity. Copyright 2000 Academic Press.

  8. Metabolomic profiles of arsenic (+3 oxidation state) methyltransferase knockout mice: effect of sex and arsenic exposure.

    Science.gov (United States)

    Huang, Madelyn C; Douillet, Christelle; Su, Mingming; Zhou, Kejun; Wu, Tao; Chen, Wenlian; Galanko, Joseph A; Drobná, Zuzana; Saunders, R Jesse; Martin, Elizabeth; Fry, Rebecca C; Jia, Wei; Stýblo, Miroslav

    2017-01-01

    Arsenic (+3 oxidation state) methyltransferase (As3mt) is the key enzyme in the pathway for methylation of inorganic arsenic (iAs). Altered As3mt expression and AS3MT polymorphism have been linked to changes in iAs metabolism and in susceptibility to iAs toxicity in laboratory models and in humans. As3mt-knockout mice have been used to study the association between iAs metabolism and adverse effects of iAs exposure. However, little is known about systemic changes in metabolism of these mice and how these changes lead to their increased susceptibility to iAs toxicity. Here, we compared plasma and urinary metabolomes of male and female wild-type (WT) and As3mt-KO (KO) C57BL/6 mice and examined metabolomic shifts associated with iAs exposure in drinking water. Surprisingly, exposure to 1 ppm As elicited only small changes in the metabolite profiles of either WT or KO mice. In contrast, comparisons of KO mice with WT mice revealed significant differences in plasma and urinary metabolites associated with lipid (phosphatidylcholines, cytidine, acyl-carnitine), amino acid (hippuric acid, acetylglycine, urea), and carbohydrate (L-sorbose, galactonic acid, gluconic acid) metabolism. Notably, most of these differences were sex specific. Sex-specific differences were also found between WT and KO mice in plasma triglyceride and lipoprotein cholesterol levels. Some of the differentially changed metabolites (phosphatidylcholines, carnosine, and sarcosine) are substrates or products of reactions catalyzed by other methyltransferases. These results suggest that As3mt KO alters major metabolic pathways in a sex-specific manner, independent of iAs treatment, and that As3mt may be involved in other cellular processes beyond iAs methylation.

  9. Epigenetic silencing of O6-methylguanine DNA methyltransferase gene in NiS-transformed cells.

    Science.gov (United States)

    Ji, Weidong; Yang, Linqing; Yu, Lei; Yuan, Jianhui; Hu, Dalin; Zhang, Wenjuan; Yang, Jianping; Pang, Yaqin; Li, Wenxue; Lu, Jiachun; Fu, Juan; Chen, Jiakun; Lin, Zhongning; Chen, Wen; Zhuang, Zhixiong

    2008-06-01

    Nickel (Ni) compounds are potent carcinogens and can induce malignant transformation of rodent and human cells. To uncover the molecular mechanisms of nickel sulfide (NiS)-induced cell transformation, we investigated epigenetic alterations in a set of DNA repair genes. The silencing of the O(6)-methylguanine DNA methyltransferase (MGMT) gene locus and upregulation of DNA methyltransferase 1 (DNMT1) expression was specifically detected in NiS-transformed human bronchial epithelial (16HBE) cells. In addition, we noted epigenetic alterations including DNA hypermethylation, reduced histone H4 acetylation and a decrease in the ratio of Lys-9 acetylated/methylated histone H3 at the MGMT CpG island in NiS-transformed 16HBE cells. Meanwhile, we identified concurrent binding of methyl-CpG-binding protein 2, methylated DNA-binding domain protein 2 and DNMT1 to the CpG island of the MGMT promoter, demonstrating that these components collaborate to maintain MGMT methylation in NiS-transformed cells. Moreover, depletion of DNMT1 by introduction of a small hairpin RNA construct into NiS-transformed cells resulted in a 30% inhibition of cell proliferation and led to increased MGMT gene expression by reversion of the epigenetic modifications at the MGMT promoter region. MGMT suppression and hypermethylation at the CpG island of the MGMT promoter occurred 6 days after NiS treatment, indicating that epigenetic modifications of MGMT might be an early event in tumorigenesis. Taken together, these observations demonstrate that epigenetic silencing of MGMT is associated with DNA hypermethylation, histone modifications and DNMT1 upregulation, which contribute to NiS-induced malignant transformation.

  10. Protein arginine methyltransferase 5 (PRMT5) is a novel coactivator of constitutive androstane receptor (CAR)

    Energy Technology Data Exchange (ETDEWEB)

    Kanno, Yuichiro, E-mail: ykanno@phar.toho-u.ac.jp; Inajima, Jun; Kato, Sayaka; Matsumoto, Maika; Tokumoto, Chikako; Kure, Yuki; Inouye, Yoshio

    2015-03-27

    The constitutive androstane receptor (CAR) plays a key role in the expression of xenobiotic/steroid and drug metabolizing enzymes and their transporters. In this study, we demonstrated that protein arginine methyltransferase 5 (PRMT5) is a novel CAR-interacting protein. Furthermore, the PRMT-dependent induction of a CAR reporter gene, which was independent of methyltransferase activity, was enhanced in the presence of steroid receptor coactivator 1 (SRC1), peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC-1α) or DEAD box DNA/RNA helicase DP97. Using tetracycline inducible-hCAR system in HepG2 cells, we showed that knockdown of PRMT5 with small interfering RNA suppressed tetracycline -induced mRNA expression of CYP2B6 but not of CYP2C9 or CYP3A4. PRMT5 enhanced phenobarbital-mediated transactivation of a phenobarbital-responsive enhancer module (PBREM)-driven reporter gene in co-operation with PGC-1α in rat primary hepatocytes. Based on these findings, we suggest PRMT5 to be a gene (or promoter)-selective coactivator of CAR by mediating the formation of complexes between hCAR and appropriate coactivators. - Highlights: • Nuclear receptor CAR interact with PRMT5. • PRMT5 enhances transcriptional activity of CAR. • PRMT5 synergistically enhances transactivity of CAR by the co-expression of SRC-1, DP97 or PGC1α. • PRMT5 is a gene-selective co-activator for hCAR.

  11. Sensitive detection of DNA methyltransferase using the dendritic rolling circle amplification-induced fluorescence.

    Science.gov (United States)

    Song, Weiling; Luan, Yawen; Guo, Xiaoyan; He, Peng; Zhang, Xiaoru

    2017-03-01

    The analysis of DNA methylation and MTase activities is very important in the early clinical diagnosis of cancer, on purposes of providing insights into the mechanism of gene repression and developing novel drugs of treating methylation-related diseases. Combining the dendritic rolling circle amplification and Mg(2+)-dependent DNAzyme with a function of catalyzing the generation of a fluorophore-labeled nucleic acid acting as readout signal for the analyses, a new fluorescent method for DNA methyltransferase detection was reported. In the presence of DNA methyltransferases (MTase), the methylation-responsive sequence of double-stranded DNA probe was methylated and then cleaved by the methylation-sensitive restriction endonuclease DpnI. The cleaved hybrid DNA probe then functioned as a signal primer to initiate the dendritic rolling circle amplification reaction, containing a circular DNA and a structurally tailored hairpin structure. Subsequently, the circular nucleic acid template produced a complementary sequence to the Mg(2+)-dependent DNAzyme and a sequence identical to the loop region of the co-added hairpin structure. At last, a fluorescence readout signal was afforded by the DNAzyme-catalyzed cleavage of a fluorophore/quencher-modified substrate. This method enabled the analysis of the target MTase with a detection limit up to 0.36 U mL(-1), and a dynamic range was obtained from 1.0 to 10 U mL(-1). Moreover, the proposed strategy was successfully applied in real sample assay. With this assay, the inhibitors of MTase were evaluated and screened which might be helpful for the discovery of anticancer drugs. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Expression patterns and miRNA regulation of DNA methyltransferases in chicken primordial germ cells.

    Directory of Open Access Journals (Sweden)

    Deivendran Rengaraj

    Full Text Available DNA methylation is widespread in most species, from bacteria to mammals, and is crucial for genomic imprinting, gene expression, and embryogenesis. DNA methylation occurs via two major classes of enzymatic reactions: maintenance-type methylation catalyzed by DNA (cytosine-5--methyltransferase (DNMT 1, and de novo methylation catalyzed by DNMT 3 alpha (DNMT3A and -beta (DNMT3B. The expression pattern and regulation of DNMT genes in primordial germ cells (PGCs and germ line cells has not been sufficiently established in birds. Therefore, we employed bioinformatics, RT-PCR, real-time PCR, and in situ hybridization analyses to examine the structural conservation and conserved expression patterns of chicken DNMT family genes. We further examined the regulation of a candidate de novo DNA methyltransferase gene, cDNMT3B by cotransfection of cDNMT3B 3'UTR- and cDNMT3B 3'UTR-specific miRNAs through a dual fluorescence reporter assay. All cDNMT family members were differentially detected during early embryonic development. Of interest, cDNMT3B expression was highly detected in early embryos and in PGCs. During germ line development and sexual maturation, cDNMT3B expression was reestablished in a female germ cell-specific manner. In the dual fluorescence reporter assay, cDNMT3B expression was significantly downregulated by four miRNAs: gga-miR-15c (25.82%, gga-miR-29b (30.01%, gga-miR-383 (30.0%, and gga-miR-222 (31.28%. Our data highlight the structural conservation and conserved expression patterns of chicken DNMTs. The miRNAs investigated in this study may induce downregulation of gene expression in chicken PGCs and germ cells.

  13. Molecular and biochemical characterization of the jasmonic acid methyltransferase gene from black cottonwood (Populus trichocarpa)

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Nan [ORNL; Yao, Jianzhuang [University of Tennessee, Knoxville (UTK); Chaiprasongsuk, Minta [University of Tennessee, Knoxville (UTK); Li, Guanglin [University of Tennessee, Knoxville (UTK); Guan, Ju [University of Tennessee, Knoxville (UTK); Tschaplinski, Timothy J [ORNL; Guo, Hong [University of Tennessee, Knoxville (UTK); Chen, Feng [University of Tennessee, Knoxville (UTK)

    2013-01-01

    Methyl jasmonate is a metabolite known to be produced by many plants and has roles in diverse biological processes. It is biosynthesized by the action of S-adenosyl-L-methionine:jasmonic acid carboxyl methyltransferase (JMT), which belongs to the SABATH family of methyltransferases. Herein is reported the isolation and biochemical characterization of a JMT gene from black cottonwood (Populus trichocarpa). The genome of P. trichocarpa contains 28 SABATH genes (PtSABATH1 to PtSABATH28). Recombinant PtSABATH3 expressed in Escherichia coli showed the highest level of activity with jasmonic acid (JA) among carboxylic acids tested. It was therefore renamed PtJMT1. PtJMT1 also displayed activity with benzoic acid (BA), with which the activity was about 22% of that with JA. PtSABATH2 and PtSABATH4 were most similar to PtJMT1 among all PtSABATHs. However, neither of them had activity with JA. The apparent Km values of PtJMT1 using JA and BA as substrate were 175 lM and 341 lM, respectively. Mutation of Ser-153 and Asn-361, two residues in the active site of PtJMT1, to Tyr and Ser respectively, led to higher specific activity with BA than with JA. Homology-based structural modeling indicated that substrate alignment, in which Asn-361 is involved, plays a role in determining the substrate specificity of PtJMT1. In the leaves of young seedlings of black cottonwood, the expression of PtJMT1 was induced by plant defense signal molecules methyl jasmonate and salicylic acid and a fungal elicitor alamethicin, suggesting that PtJMT1 may have a role in plant defense against biotic stresses. Phylogenetic analysis suggests that PtJMT1 shares a common ancestor with the Arabidopsis JMT, and functional divergence of these two apparent JMT orthologs has occurred since the split of poplar and Arabidopsis lineages.

  14. Pain vulnerability and DNA methyltransferase 3a involved in the affective dimension of chronic pain.

    Science.gov (United States)

    Wang, Wei; Li, Caiyue; Cai, Youqing; Pan, Zhizhong Z

    2017-01-01

    Chronic pain with comorbid emotional disorders is a prevalent neurological disease in patients under various pathological conditions, yet patients show considerable difference in their vulnerability to developing chronic pain. Understanding the neurobiological basis underlying this pain vulnerability is essential to develop targeted therapies of higher efficiency in pain treatment of precision medicine. However, this pain vulnerability has not been addressed in preclinical pain research in animals to date. In this study, we investigated individual variance in both sensory and affective/emotional dimensions of pain behaviors in response to chronic neuropathic pain condition in a mouse model of chronic pain. We found that mice displayed considerably diverse sensitivities in the chronic pain-induced anxiety- and depression-like behaviors of affective pain. Importantly, the mouse group that was more vulnerable to developing anxiety was also more vulnerable to developing depressive behavior under the chronic pain condition. In contrast, there was relatively much less variance in individual responses in the sensory dimension of pain sensitization. Molecular analysis revealed that those mice vulnerable to developing the emotional disorders showed a significant reduction in the protein level of DNA methyltransferase 3a in the emotion-processing central nucleus of the amygdala. In addition, social stress also revealed significant individual variance in anxiety behavior in mice. These findings suggest that individual pain vulnerability may be inherent mostly in the emotional/affective component of chronic pain and remain consistent in different aspects of negative emotion, in which adaptive changes in the function of DNA methyltransferase 3a for DNA methylation in central amygdala may play an important role. This may open a new avenue of basic research into the neurobiological mechanisms underlying pain vulnerability.

  15. Human calmodulin methyltransferase: expression, activity on calmodulin, and Hsp90 dependence.

    Directory of Open Access Journals (Sweden)

    Sophia Magen

    Full Text Available Deletion of the first exon of calmodulin-lysine N-methyltransferase (CaM KMT, previously C2orf34 has been reported in two multigene deletion syndromes, but additional studies on the gene have not been reported. Here we show that in the cells from 2p21 deletion patients the loss of CaM KMT expression results in accumulation of hypomethylated calmodulin compared to normal controls, suggesting that CaM KMT is essential for calmodulin methylation and there are no compensatory mechanisms for CaM methylation in humans. We have further studied the expression of this gene at the transcript and protein levels. We have identified 2 additional transcripts in cells of the 2p21 deletion syndrome patients that start from alternative exons positioned outside the deletion region. One of them starts in the 2(nd known exon, the other in a novel exon. The transcript starting from the novel exon was also identified in a variety of tissues from normal individuals. These new transcripts are not expected to produce proteins. Immunofluorescent localization of tagged CaM KMT in HeLa cells indicates that it is present in both the cytoplasm and nucleus of cells whereas the short isoform is localized to the Golgi apparatus. Using Western blot analysis we show that the CaM KMT protein is broadly expressed in mouse tissues. Finally we demonstrate that the CaM KMT interacts with the middle portion of the Hsp90 molecular chaperon and is probably a client protein since it is degraded upon treatment of cells with the Hsp90 inhibitor geldanamycin. These findings suggest that the CaM KMT is the major, possibly the single, methyltransferase of calmodulin in human cells with a wide tissue distribution and is a novel Hsp90 client protein. Thus our data provides basic information for a gene potentially contributing to the patient phenotype of two contiguous gene deletion syndromes.

  16. Clinical and Microbiological Aspects of Linezolid Resistance Mediated by the cfr Gene Encoding a 23S rRNA Methyltransferase▿

    Science.gov (United States)

    Arias, Cesar A.; Vallejo, Martha; Reyes, Jinnethe; Panesso, Diana; Moreno, Jaime; Castañeda, Elizabeth; Villegas, Maria V.; Murray, Barbara E.; Quinn, John P.

    2008-01-01

    The cfr (chloramphenicol-florfenicol resistance) gene encodes a 23S rRNA methyltransferase that confers resistance to linezolid. Detection of linezolid resistance was evaluated in the first cfr-carrying human hospital isolate of linezolid and methicillin-resistant Staphylococcus aureus (designated MRSA CM-05) by dilution and diffusion methods (including Etest). The presence of cfr was investigated in isolates of staphylococci colonizing the patient's household contacts and clinical isolates recovered from patients in the same unit where MRSA CM-05 was isolated. Additionally, 68 chloramphenicol-resistant Colombian MRSA isolates recovered from hospitals between 2001 and 2004 were screened for the presence of the cfr gene. In addition to erm(B), the erm(A) gene was also detected in CM-05. The isolate belonged to sequence type 5 and carried staphylococcal chromosomal cassette mec type I. We were unable to detect the cfr gene in any of the human staphylococci screened (either clinical or colonizing isolates). Agar and broth dilution methods detected linezolid resistance in CM-05. However, the Etest and disk diffusion methods failed to detect resistance after 24 h of incubation. Oxazolidinone resistance mediated by the cfr gene is rare, and acquisition by a human isolate appears to be a recent event in Colombia. The detection of cfr-mediated linezolid resistance might be compromised by the use of the disk diffusion or Etest method. PMID:18174304

  17. Increasing the Fill Factor of Inverted P3HT:PCBM Solar Cells Through Surface Modification of Al-Doped ZnO via Phosphonic Acid-Anchored C60 SAMs

    DEFF Research Database (Denmark)

    Stubhan, Tobias; Salinas, Michael; Ebel, Alexander

    2012-01-01

    The influence of aluminum-doped zinc oxide (AZO) electron extraction layers modified with self-assembled monolayers (SAMs) on inverted polymer solar cells is investigated. It is found that AZO modification with phosphonic acid-anchored Fullerene–SAMs leads to a reduction of the series resistance,...

  18. Detecting Organic Compounds Released from Iron Oxidizing Bacteria using Sample Analysis at Mars (SAM) Like Instrument Protocols

    Science.gov (United States)

    Glavin, D. P.; Popa, R.; Martin, M. G.; Freissinet, C.; Fisk, M. R.; Dworkin, J. P.; Mahaffy, P. R.

    2012-01-01

    Mars is a planet of great interest for Astrobiology since its past environmental conditions are thought to have been favourable for the emergence life. At present, the Red Planet is extremely cold and dry and the surface is exposed to intense UV and ionizing radiation, conditions generally considered to be incompatible with life as we know it on Earth. It was proposed that the shallow subsurface of Mars, where temperatures can be above freezing and liquid water can exist on rock surfaces, could harbor chemolithoautotrophic bacteria such as the iron oxidizing microorganism Pseudomonas sp. HerB. The Mars Science Laboratory (MSL) mission will provide the next opportunity to carry out in situ measurements for organic compounds of possible biological origin on Mars. One instrument onboard MSL, called the Sample Analysis at Mars (SAM) instrument suite, will carry out a broad and sensitive search for organic compounds in surface samples using either high temperature pyrolysis or chemical extraction followed by gas chromatography mass spectrometry. We present gas chromatograph mass spectrometer (GC/MS) data on crushed olivine rock powders that have been inoculated with Pseudomonas sp. HerB at different concentrations ranging from approx 10(exp 2) to 10(exp 7) cells per gram. The inoculated olivine samples were heated under helium carrier gas flow at 500 C and the pyrolysis products concentrated using a SAM-like hydrocarbon trap set at -20 C followed by trap heating and analysis by GC/Ms. In addition, the samples were also extracted using a low temperature "one-pot" chemical extraction technique using N-methyl, N-(tert-butyldimethylsilyl) trifluoroacetamide (MTBSTFA) as the silylating agent prior to GC/MS analysis. We identified several aldehydes, thiols, and alkene nitriles after pyrolysis GC/MS analysis of the bacteria that were not found in the olivine control samples that had not been inoculated with bacteria. The distribution of pyrolysis products extracted from the

  19. Expression of DNA methyltransferases in breast cancer patients and to analyze the effect of natural compounds on DNA methyltransferases and associated proteins.

    Science.gov (United States)

    Mirza, Sameer; Sharma, Gayatri; Parshad, Rajinder; Gupta, Sidhartha Datta; Pandya, Pranav; Ralhan, Ranju

    2013-03-01

    The DNA methylation mediated by specific DNA methyltransferases (DNMTs), results in the epigenetic silencing of multiple genes which are implicated in human breast cancer. We hypothesized that the natural compounds modulate the expression of DNMTs and their associated proteins in the breast cancer cell lines and affect the methylation mediated gene silencing. The DNMTs transcript expression was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) in the tumors and the adjacent normal breast tissues of the patients with invasive ductal breast carcinoma. We tested the hypothesis that the natural compounds, viz., epigallocatechin gallate (EGCG), genistein, withaferin A, curcumin, resveratrol, and guggulsterone, have demethylation potential. To investigate this hypothesis, we analyzed the DNMTs expression at the transcript levels, followed by the analysis of DNMT1 and its associated proteins (HDAC1, MeCP2, and MBD2). The increased DNMTs transcripts expression, viz., DNMT1, DNMT3a, and DNMT3b, in the breast cancer tissues suggest involvement of the DNMTs in the breast carcinogenesis. Quantitative RT-PCR analysis revealed that the treatment with natural compounds, viz., EGCG, genistein, withaferin A, curcumin, resveratrol, and guggulsterone, resulted in a significant decrease in the transcript levels of all the DNMTs investigated. Importantly, these natural compounds decreased the protein levels of DNMT1, HDAC1, and MeCP2. Our results demonstrate that the natural compounds, EGCG, genistein, withaferin A, curcumin, resveratrol, and guggulsterone, have the potential to reverse the epigenetic changes. Moreover, their lack of toxicity makes these natural compounds promising candidates for the chemoprevention of the breast cancer. In-depth future mechanistic studies aimed to elucidate how these compounds affect the gene transcription are warranted.

  20. Novel Gas Sensor Arrays Based on High-Q SAM-Modified Piezotransduced Single-Crystal Silicon Bulk Acoustic Resonators

    Directory of Open Access Journals (Sweden)

    Yuan Zhao

    2017-06-01

    Full Text Available This paper demonstrates a novel micro-size (120 μm × 200 μm piezoelectric gas sensor based on a piezotransduced single-crystal silicon bulk acoustic resonator (PSBAR. The PSBARs operate at 102 MHz and possess high Q values (about 2000, ensuring the stability of the measurement. A corresponding gas sensor array is fabricated by integrating three different self-assembled monolayers (SAMs modified PSBARs. The limit of detection (LOD for ethanol vapor is demonstrated to be as low as 25 ppm with a sensitivity of about 1.5 Hz/ppm. Two sets of identification code bars based on the sensitivities and the adsorption energy constants are utilized to successfully discriminate isopropanol (IPA, ethanol, hexane and heptane vapors at low and high gas partial pressures, respectively. The proposed sensor array shows the potential to form a portable electronic nose system for volatile organic compound (VOC differentiation.