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Sample records for arginine protein translocase

  1. Subunit Organization in the TatA Complex of the Twin Arginine Protein Translocase

    OpenAIRE

    White, Gaye F.; Schermann, Sonya M.; Bradley, Justin; Roberts, Andrew; Greene, Nicholas P.; Berks, Ben C.; Thomson, Andrew J.

    2009-01-01

    The Tat system is used to transport folded proteins across the cytoplasmic membrane in bacteria and archaea and across the thylakoid membrane of plant chloroplasts. Multimers of the integral membrane TatA protein are thought to form the protein-conducting element of the Tat pathway. Nitroxide radicals were introduced at selected positions within the transmembrane helix of Escherichia coli TatA and used to probe the structure of detergent-solubilized TatA complexes by EPR spectroscopy. A compa...

  2. A mutation leading to super-assembly of twin-arginine translocase (Tat) protein complexes

    NARCIS (Netherlands)

    Patel, Roshani; Vasilev, Cvetelin; Beck, Daniel; Monteferrante, Carmine G; van Dijl, Jan Maarten; Hunter, C Neil; Smith, Corinne; Robinson, Colin

    2014-01-01

    The Tat system transports folded proteins across the bacterial plasma membrane. The mechanism is believed to involve coalescence of a TatC-containing unit with a separate TatA complex, but the full translocation complex has never been visualised and the assembly process is poorly defined. We report

  3. Identification of protein-protein interactions between the TatB and TatC subunits of the twin-arginine translocase system and respiratory enzyme specific chaperones.

    Science.gov (United States)

    Kuzniatsova, Lalita; Winstone, Tara M L; Turner, Raymond J

    2016-04-01

    The Twin-arginine translocation (Tat) pathway serves for translocation of fully folded proteins across the cytoplasmic membrane in bacterial and chloroplast thylakoid membranes. The Escherichia coli Tat system consists of three core components: TatA, TatB, and TatC. The TatB and TatC subunits form the receptor complex for Tat dependent proteins. The TatB protein is composed of a single transmembrane helix and cytoplasmic domain. The structure of TatC revealed six transmembrane helices. Redox Enzyme Maturation Proteins (REMPs) are system specific chaperones, which play roles in the maturation of Tat dependent respiratory enzymes. Here we applied the in vivo bacterial two-hybrid technique to investigate interaction of REMPs with the TatBC proteins, finding that all but the formate dehydrogenase REMP dock to TatB or TatC. We focused on the NarJ subfamily, where DmsD--the REMP for dimethyl sulfoxide reductase in E. coli--was previously shown to interact with TatB and TatC. We found that these REMPs interact with TatC cytoplasmic loops 1, 2 and 4, with the exception of NarJ, that only interacts with 1 and 4. An in vitro isothermal titration calorimetry study was applied to confirm the evidence of interactions between TatC fragments and DmsD chaperone. Using a peptide overlapping array, it was shown that the different NarJ subfamily REMPs interact with different regions of the TatB cytoplasmic domains. The results demonstrate a role of REMP chaperones in targeting respiratory enzymes to the Tat system. The data suggests that the different REMPs may have different mechanisms for this task. PMID:26826271

  4. Rhomboid protease AarA mediates quorum-sensing in Providencia stuartii by activating TatA of the twin-arginine translocase

    OpenAIRE

    Stevenson, Lindsay G.; Strisovsky, Kvido; Clemmer, Katy M.; Bhatt, Shantanu; Freeman, Matthew; Rather, Philip N.

    2007-01-01

    The Providencia stuartii AarA protein is a member of the rhomboid family of intramembrane serine proteases and is required for the production of an unknown quorum-sensing molecule. In a screen to identify rhomboid-encoding genes from Proteus mirabilis, tatA was identified as a multicopy suppressor and restored extracellular signal production as well as complementing all other phenotypes of a Prov. stuartii aarA mutant. TatA is a component of the twin-arginine translocase (Tat) protein secreti...

  5. Unique Photobleaching Phenomena of the Twin-Arginine Translocase Respiratory Enzyme Chaperone DmsD

    OpenAIRE

    Rivardo, Fabrizio; Leach, Thorin G.H.; Chan, Catherine S.; Winstone, Tara M. L.; Ladner, Carol L.; Sarfo, Kwabena J.; Turner, Raymond J.

    2014-01-01

    DmsD is a chaperone of the redox enzyme maturation protein family specifically required for biogenesis of DMSO reductase in Escherichia coli. It exists in multiple folding forms, all of which are capable of binding its known substrate, the twin-arginine leader sequence of the DmsA catalytic subunit. It is important for maturation of the reductase and targeting to the cytoplasmic membrane for translocation. Here, we demonstrate that DmsD exhibits an irreversible photobleaching phenomenon upon ...

  6. Mitochondrial OXA Translocase Plays a Major Role in Biogenesis of Inner-Membrane Proteins.

    Science.gov (United States)

    Stiller, Sebastian B; Höpker, Jan; Oeljeklaus, Silke; Schütze, Conny; Schrempp, Sandra G; Vent-Schmidt, Jens; Horvath, Susanne E; Frazier, Ann E; Gebert, Natalia; van der Laan, Martin; Bohnert, Maria; Warscheid, Bettina; Pfanner, Nikolaus; Wiedemann, Nils

    2016-05-10

    The mitochondrial inner membrane harbors three protein translocases. Presequence translocase and carrier translocase are essential for importing nuclear-encoded proteins. The oxidase assembly (OXA) translocase is required for exporting mitochondrial-encoded proteins; however, different views exist about its relevance for nuclear-encoded proteins. We report that OXA plays a dual role in the biogenesis of nuclear-encoded mitochondrial proteins. First, a systematic analysis of OXA-deficient mitochondria led to an unexpected expansion of the spectrum of OXA substrates imported via the presequence pathway. Second, biogenesis of numerous metabolite carriers depends on OXA, although they are not imported by the presequence pathway. We show that OXA is crucial for the biogenesis of the Tim18-Sdh3 module of the carrier translocase. The export translocase OXA is thus required for the import of metabolite carriers by promoting assembly of the carrier translocase. We conclude that OXA is of central importance for the biogenesis of the mitochondrial inner membrane. PMID:27166948

  7. The presence of disulfide bonds reveals an evolutionarily conserved mechanism involved in mitochondrial protein translocase assembly

    Science.gov (United States)

    Wrobel, Lidia; Sokol, Anna M.; Chojnacka, Magdalena; Chacinska, Agnieszka

    2016-01-01

    Disulfide bond formation is crucial for the biogenesis and structure of many proteins that are localized in the intermembrane space of mitochondria. The importance of disulfide bond formation within mitochondrial proteins was extended beyond soluble intermembrane space proteins. Tim22, a membrane protein and core component of the mitochondrial translocase TIM22, forms an intramolecular disulfide bond in yeast. Tim22 belongs to the Tim17/Tim22/Tim23 family of protein translocases. Here, we present evidence of the high evolutionary conservation of disulfide bond formation in Tim17 and Tim22 among fungi and metazoa. Topological models are proposed that include the location of disulfide bonds relative to the predicted transmembrane regions. Yeast and human Tim22 variants that are not oxidized do not properly integrate into the membrane complex. Moreover, the lack of Tim17 oxidation disrupts the TIM23 translocase complex. This underlines the importance of disulfide bond formation for mature translocase assembly through membrane stabilization of weak transmembrane domains. PMID:27265872

  8. It takes two to tango: two TatA paralogues and two redox enzyme-specific chaperones are involved in the localization of twin-arginine translocase substrates in Campylobacter jejuni.

    OpenAIRE

    Liu, Y.W.; Hitchcock, A.; Salmon, R.C.; Kelly, D J

    2014-01-01

    The food-borne zoonotic pathogen Campylobacter jejuni has complex electron transport chains required for growth in the host, many of which contain cofactored periplasmic enzymes localized by the twin-arginine translocase (TAT). We report here the identification of two paralogues of the TatA translocase component in C. jejuni strain NCTC 11168, encoded by cj1176c (tatA1) and cj0786 (tatA2). Deletion mutants constructed in either or both of the tatA1 and tatA2 genes displayed distinct growth an...

  9. Arginine Depletion by Arginine Deiminase Does Not Affect Whole Protein Metabolism or Muscle Fractional Protein Synthesis Rate in Mice

    Science.gov (United States)

    Marini, Juan C.; Didelija, Inka Cajo

    2015-01-01

    Due to the absolute need for arginine that certain cancer cells have, arginine depletion is a therapy in clinical trials to treat several types of cancers. Arginine is an amino acids utilized not only as a precursor for other important molecules, but also for protein synthesis. Because arginine depletion can potentially exacerbate the progressive loss of body weight, and especially lean body mass, in cancer patients we determined the effect of arginine depletion by pegylated arginine deiminase (ADI-PEG 20) on whole body protein synthesis and fractional protein synthesis rate in multiple tissues of mice. ADI-PEG 20 successfully depleted circulating arginine (<1 μmol/L), and increased citrulline concentration more than tenfold. Body weight and body composition, however, were not affected by ADI-PEG 20. Despite the depletion of arginine, whole body protein synthesis and breakdown were maintained in the ADI-PEG 20 treated mice. The fractional protein synthesis rate of muscle was also not affected by arginine depletion. Most tissues (liver, kidney, spleen, heart, lungs, stomach, small and large intestine, pancreas) were able to maintain their fractional protein synthesis rate; however, the fractional protein synthesis rate of brain, thymus and testicles was reduced due to the ADI-PEG 20 treatment. Furthermore, these results were confirmed by the incorporation of ureido [14C]citrulline, which indicate the local conversion into arginine, into protein. In conclusion, the intracellular recycling pathway of citrulline is able to provide enough arginine to maintain protein synthesis rate and prevent the loss of lean body mass and body weight. PMID:25775142

  10. Structural model for the protein-translocating element of the twin-arginine transport system

    Science.gov (United States)

    Rodriguez, Fernanda; Rouse, Sarah L.; Tait, Claudia E.; Harmer, Jeffrey; De Riso, Antonio; Timmel, Christiane R.; Sansom, Mark S. P.; Berks, Ben C.; Schnell, Jason R.

    2013-01-01

    The twin-arginine translocase (Tat) carries out the remarkable process of translocating fully folded proteins across the cytoplasmic membrane of prokaryotes and the thylakoid membrane of plant chloroplasts. Tat is required for bacterial pathogenesis and for photosynthesis in plants. TatA, the protein-translocating element of the Tat system, is a small transmembrane protein that assembles into ring-like oligomers of variable size. We have determined a structural model of the Escherichia coli TatA complex in detergent solution by NMR. TatA assembly is mediated entirely by the transmembrane helix. The amphipathic helix extends outwards from the ring of transmembrane helices, permitting assembly of complexes with variable subunit numbers. Transmembrane residue Gln8 points inward, resulting in a short hydrophobic pore in the center of the complex. Simulations of the TatA complex in lipid bilayers indicate that the short transmembrane domain distorts the membrane. This finding suggests that TatA facilitates protein transport by sensitizing the membrane to transient rupture. PMID:23471988

  11. Arginine-aromatic interactions and their effects on arginine-induced solubilization of aromatic solutes and suppression of protein aggregation

    KAUST Repository

    Shah, Dhawal

    2011-09-21

    We examine the interaction of aromatic residues of proteins with arginine, an additive commonly used to suppress protein aggregation, using experiments and molecular dynamics simulations. An aromatic-rich peptide, FFYTP (a segment of insulin), and lysozyme and insulin are used as model systems. Mass spectrometry shows that arginine increases the solubility of FFYTP by binding to the peptide, with the simulations revealing the predominant association of arginine to be with the aromatic residues. The calculations further show a positive preferential interaction coefficient, Γ XP, contrary to conventional thinking that positive Γ XP\\'s indicate aggregation rather than suppression of aggregation. Simulations with lysozyme and insulin also show arginine\\'s preference for aromatic residues, in addition to acidic residues. We use these observations and earlier results reported by us and others to discuss the possible implications of arginine\\'s interactions with aromatic residues on the solubilization of aromatic moieties and proteins. Our results also highlight the fact that explanations based purely on Γ XP, which measures average affinity of an additive to a protein, could obscure or misinterpret the underlying molecular mechanisms behind additive-induced suppression of protein aggregation. © 2011 American Institute of Chemical Engineers (AIChE).

  12. Evaluation of Porin Interaction with Adenine Nucleotide Translocase and Cyclophilin-D Proteins after Brain Ischemia and Reperfusion

    Directory of Open Access Journals (Sweden)

    Mohammad Ali Atlasi

    2011-01-01

    Full Text Available Objective (s Porin is a mitochondrial outer membrane channel, which usually functions as the pathway for the movement of various substances in and out of the mitochondria and is considered to be a component of the permeability transition (PT pore complex that plays a role in the PT. We addressed the hypothesis that porin interacts with other mitochondrial proteins after ischemic injury.Materials and MethodsFor this purpose, we used in vivo 4-vessel occlusion model of rat brain and porin purification method by hydroxyapatite column. After SDS gel electrophoresis and silver nitrate staining, Western blotting was done for porin, adenine nucleotide translocase and cyclophilin-D proteins.Results Porin was purified from mitochondrial mixture in ischemic brain and control groups. Investigation of interaction of adenine nucleotide transposes (ANT and cyclophilin-D with porin by Western blotting showed no proteins co-purified with porin from injured tissues.Conclusion The present study implies that there may not be interaction between porin, and ANT or cyclophilin-D, and if there is any, it is not maintained during the purification procedure.

  13. The regulatory PII protein controls arginine biosynthesis in Arabidopsis.

    Science.gov (United States)

    Ferrario-Méry, Sylvie; Besin, Evelyne; Pichon, Olivier; Meyer, Christian; Hodges, Michael

    2006-04-01

    In higher plants, PII is a nuclear-encoded plastid protein which is homologous to bacterial PII signalling proteins known to be involved in the regulation of nitrogen metabolism. A reduced ornithine, citrulline and arginine accumulation was observed in two Arabidopsis PII knock-out mutants in response to NH4+ resupply after N starvation. This difference could be explained by the regulation of a key enzyme of the arginine biosynthesis pathway, N-acetyl glutamate kinase (NAGK) by PII. In vitro assays using purified recombinant proteins showed the catalytic activation of Arabidopsis NAGK by PII giving the first evidence of a physiological role of the PII protein in higher plants. Using Arabidopsis transcriptome microarray (CATMA) and RT-PCR analyses, it was found that none of the genes involved in the arginine biosynthetic or catabolic pathways were differentially expressed in a PII knock-out mutant background. In conclusion, the observed changes in metabolite levels can be explained by the reduced activation of NAGK by PII. PMID:16545809

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

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    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.

  15. Haloarchaeal Protein Translocation via the Twin Arginine Translocation Pathway

    Energy Technology Data Exchange (ETDEWEB)

    Pohlschroder Mechthild

    2009-02-03

    Protein transport across hydrophobic membranes that partition cellular compartments is essential in all cells. The twin arginine translocation (Tat) pathway transports proteins across the prokaryotic cytoplasmic membranes. Distinct from the universally conserved Sec pathway, which secretes unfolded proteins, the Tat machinery is unique in that it secretes proteins in a folded conformation, making it an attractive pathway for the transport and secretion of heterologously expressed proteins that are Sec-incompatible. During the past 7 years, the DOE-supported project has focused on the characterization of the diversity of bacterial and archaeal Tat substrates as well as on the characterization of the Tat pathway of a model archaeon, Haloferax volcanii, a member of the haloarchaea. We have demonstrated that H. volcanii uses this pathway to transport most of its secretome.

  16. Evaluation of chemical labeling methods for identifying functional arginine residues of proteins by mass spectrometry.

    Science.gov (United States)

    Wanigasekara, Maheshika S K; Chowdhury, Saiful M

    2016-09-01

    Arginine residues undergo several kinds of post-translational modifications (PTMs). These PTMs are associated with several inflammatory diseases, such as rheumatoid arthritis, atherosclerosis, and diabetes. Mass spectrometric studies of arginine modified proteins and peptides are very important, not only to identify the reactive arginine residues but also to understand the tandem mass spectrometry behavior of these peptides for assigning the sequences unambiguously. Herein, we utilize tandem mass spectrometry to report the performance of two widely used arginine labeling reagents, 1,2-cyclohexanedione (CHD) and phenylglyoxal (PG) with several arginine containing peptides and proteins. Time course labeling studies were performed to demonstrate the selectivity of the reagents in proteins or protein digests. Structural studies on the proteins were also explored to better understand the reaction sites and position of arginine residues. We found CHD showed better labeling efficiencies compared to phenylglyoxal. Reactive arginine profiling on a purified albumin protein clearly pointed out the cellular glycation modification site for this protein with high confidence. We believe these detailed mass-spectrometric studies will provide significant input to profile reactive arginine residues in large-scale studies; therefore, targeted proteomics can be performed to the short listed reactive sites for cellular arginine modifications. PMID:27543028

  17. Detection of the proteins with different arginine methylation status induced by low dose irradiation

    International Nuclear Information System (INIS)

    Complete text of publication follows. Objective: The objective of this study is to detect the noble proteins that were functionally regulated by change of arginine methylation through irradiation of the low dose. The increase of the arginine methylation which is induced by low dose gamma-ray will have meaningful Introduction: Exposure of cells to low doses of radiation has well documented biological effect, but the underlying regulatory mechanisms are still poorly understood. Arginine methylation is a post translational modification that results in the formation of asymmetrical and symmetrical dimethylated arginines. Post-translational methylation of arginine residues of proteins involved in a growing number of cellular processes, including transcriptional regulation, cell signaling, RNA processing and DNA repair, biological influence. Methods: Human normal cell line Chang-liver was irradiation by gamma-ray of 0.02Gy, 0.2Gy. After irradiation, cells were incubated for 4h, 8h, 24h, and then harvested to prepare protein extracts. ASYM24(anti-dimethyl-Arginine, asymmetric) antibody was used to Western blot and immunoprecipitation. Proteins that show different degrees of intensity between the two samples were analyzed by Mass spectrometry. Results: We detected increased asymmetric arginine methylation of two proteins at 24h after a dose of 0.2Gy irradiation. The mass spectrometry identified that it is 27kDa and 73kDa proteins. The 27kDa is hypothetical protein that function does not know. The 73kDa protein is Mortalin, a member of the Heat shock 70 protein family, which correlate with the radioresistance response, control of cell proliferation and act as a chaperone. Conclusion: Low dose radiation induces the change of asymmetric arginine methylation modification of arginine residues of hypothetical protein and mortalin. We expect that increase of arginine methylation in mortarin and hypothetical protein correlates with the radioresistance, the functional study for

  18. Protein arginine methyltransferase 1 regulates herpes simplex virus replication through ICP27 RGG-box methylation

    International Nuclear Information System (INIS)

    Protein arginine methylation is involved in viral infection and replication through the modulation of diverse cellular processes including RNA metabolism, cytokine signaling, and subcellular localization. It has been suggested previously that the protein arginine methylation of the RGG-box of ICP27 is required for herpes simplex virus type-1 (HSV-1) viral replication and gene expression in vivo. However, a cellular mediator for this process has not yet been identified. In our current study, we show that the protein arginine methyltransferase 1 (PRMT1) is a cellular mediator of the arginine methylation of ICP27 RGG-box. We generated arginine substitution mutants in this domain and examined which arginine residues are required for methylation by PRMT1. R138, R148 and R150 were found to be the major sites of this methylation but additional arginine residues serving as minor methylation sites are still required to sustain the fully methylated form of ICP27 RGG. We also demonstrate that the nuclear foci-like structure formation, SRPK interactions, and RNA-binding activity of ICP27 are modulated by the arginine methylation of the ICP27 RGG-box. Furthermore, HSV-1 replication is inhibited by hypomethylation of this domain resulting from the use of general PRMT inhibitors or arginine mutations. Our data thus suggest that the PRMT1 plays a key role as a cellular regulator of HSV-1 replication through ICP27 RGG-box methylation.

  19. Protein arginine methyltransferase 1 regulates herpes simplex virus replication through ICP27 RGG-box methylation

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Jungeun; Shin, Bongjin; Park, Eui-Soon; Yang, Sujeong; Choi, Seunga [Department of Microbiology, Chungnam National University, 220 Gung-dong, Yuseong-gu, Daejon 305-764 (Korea, Republic of); BK21 Bio Brain Center, Chungnam National University, 220 Gung-dong, Yuseong-gu, Daejon 305-764 (Korea, Republic of); Kang, Misun [Department of Microbiology, Chungnam National University, 220 Gung-dong, Yuseong-gu, Daejon 305-764 (Korea, Republic of); Rho, Jaerang, E-mail: jrrho@cnu.ac.kr [Department of Microbiology, Chungnam National University, 220 Gung-dong, Yuseong-gu, Daejon 305-764 (Korea, Republic of); BK21 Bio Brain Center, Chungnam National University, 220 Gung-dong, Yuseong-gu, Daejon 305-764 (Korea, Republic of); GRAST, Chungnam National University, 220 Gung-dong, Yuseong-gu, Daejon 305-764 (Korea, Republic of)

    2010-01-01

    Protein arginine methylation is involved in viral infection and replication through the modulation of diverse cellular processes including RNA metabolism, cytokine signaling, and subcellular localization. It has been suggested previously that the protein arginine methylation of the RGG-box of ICP27 is required for herpes simplex virus type-1 (HSV-1) viral replication and gene expression in vivo. However, a cellular mediator for this process has not yet been identified. In our current study, we show that the protein arginine methyltransferase 1 (PRMT1) is a cellular mediator of the arginine methylation of ICP27 RGG-box. We generated arginine substitution mutants in this domain and examined which arginine residues are required for methylation by PRMT1. R138, R148 and R150 were found to be the major sites of this methylation but additional arginine residues serving as minor methylation sites are still required to sustain the fully methylated form of ICP27 RGG. We also demonstrate that the nuclear foci-like structure formation, SRPK interactions, and RNA-binding activity of ICP27 are modulated by the arginine methylation of the ICP27 RGG-box. Furthermore, HSV-1 replication is inhibited by hypomethylation of this domain resulting from the use of general PRMT inhibitors or arginine mutations. Our data thus suggest that the PRMT1 plays a key role as a cellular regulator of HSV-1 replication through ICP27 RGG-box methylation.

  20. A TatABC-type Tat translocase is required for unimpaired aerobic growth of Corynebacterium glutamicum ATCC13032.

    Directory of Open Access Journals (Sweden)

    Dan Oertel

    Full Text Available The twin-arginine translocation (Tat system transports folded proteins across the cytoplasmic membrane of bacteria and the thylakoid membrane of plant chloroplasts. Escherichia coli and other Gram-negative bacteria possess a TatABC-type Tat translocase in which each of the three inner membrane proteins TatA, TatB, and TatC performs a mechanistically distinct function. In contrast, low-GC Gram-positive bacteria, such as Bacillus subtilis, use a TatAC-type minimal Tat translocase in which the TatB function is carried out by a bifunctional TatA. In high-GC Gram-positive Actinobacteria, such as Mycobacterium tuberculosis and Corynebacterium glutamicum, tatA, tatB, and tatC genes can be identified, suggesting that these organisms, just like E. coli, might use TatABC-type Tat translocases as well. However, since contrary to this view a previous study has suggested that C. glutamicum might in fact use a TatAC translocase with TatB only playing a minor role, we reexamined the requirement of TatB for Tat-dependent protein translocation in this microorganism. Under aerobic conditions, the misassembly of the Rieske iron-sulfur protein QcrA was identified as a major reason for the severe growth defect of Tat-defective C. glutamicum mutant strains. Furthermore, our results clearly show that TatB, besides TatA and TatC, is strictly required for unimpaired aerobic growth. In addition, TatB was also found to be essential for the secretion of a heterologous Tat-dependent model protein into the C. glutamicum culture supernatant. Together with our finding that expression of the C. glutamicum TatB in an E. coli ΔtatB mutant strain resulted in the formation of an active Tat translocase, our results clearly indicate that a TatABC translocase is used as the physiologically relevant functional unit for Tat-dependent protein translocation in C. glutamicum and, most likely, also in other TatB-containing Actinobacteria.

  1. Hypothalamic protein profiles associated with inhibited feed intake of ducks fed with insufficient dietary arginine.

    Science.gov (United States)

    Wang, C; Zheng, A J; Xie, M; Huang, W; Xie, J J; Hou, S S

    2014-07-01

    An experiment was conducted to investigate the effect of arginine on feed intake regulation. One hundred and twenty six 1-day-old male White Pekin ducks (Anas platyrhynchos domestica) were randomly were allotted to one of two dietary treatments. The birds were fed diets containing 0.71% (deficient) or 1.27% (sufficient) arginine for 3 weeks. At 21 days of age, feed intake was determined and hypothalamic protein profiles were analyzed using isobaric tags for relative and absolute quantification technique. The birds fed with arginine-deficient diet had a lower final live BW and cumulative feed intake (P1.5-fold expressional changes between arginine-deficient and -sufficient dietary treatments. Nine of these proteins were upregulated and seven of them were downregulated. The identified proteins could be regrouped into six categories: protein processing, carbohydrate metabolism and energy production, transporter, cytoskeleton, immunity and neuronal development. Dietary arginine deficiency decreased expression of proteins involved in energy production (glycine amidinotransferase, aldolase B fructose-bisphosphate, aconitase, transaldolase, 6-phosphofructokinase type C-like) and oxygen transportation (haemoglobin subunit α expression). The proteomic alterations described here provides valuable insights into the interactions of arginine with appetite. PMID:24804691

  2. Aflatoxin B1 induced upregulation of protein arginine methyltransferase 5 in human cell lines.

    Science.gov (United States)

    Ghufran, Md Sajid; Ghosh, Krishna; Kanade, Santosh R

    2016-09-01

    The exposure of naturally occurring mycotoxins affects human health and play a vital role in cancer initiation and progression. Aflatoxin B1 is a difuranocoumarin mycotoxin, classified as a group I carcinogen. The present study was conducted to assess the effect of aflatoxin B1 on epigenetic regulatory proteins. The protein arginine methyltransferase 5 expression was induced upon aflatoxin B1 treatment in a dose and time dependent manner. Further global arginine methylation was also increased in the same manner. This is the first report showing the induction of epigenetic regulatory protein, protein arginine methyltransferase 5 upon aflatoxin B1 treatment. Further study is required to establish the detailed pathway of PRMT5 induction. PMID:27242039

  3. Radiometric assay for determining the incorporation of L-canavanine or L-arginine into protein

    International Nuclear Information System (INIS)

    Procedures for a radiometric assay of L-[guanidinooxy-14C]canavanine were developed which provide a convenient and accurate measure of the incorporation of [14C]canavanine into de novo-synthesized proteins. These methods are also applicable to determining [14C]arginine incorporation into protein. These procedures have been employed to study the synthesis of L-[guanidinooxy-14C]canavanine- and L-[guanidino-14C]arginine-containing proteins from the hemolymph of Manduca sexta and Heliothis virescens, two highly destructive insect pests

  4. Shrimp arginine kinase being a binding protein of WSSV envelope protein VP31

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    Ma, Cuiyan; Gao, Qiang; Liang, Yan; Li, Chen; Liu, Chao; Huang, Jie

    2016-03-01

    Viral entry into the host is the earliest stage of infection in the viral life cycle in which attachment proteins play a key role. VP31 (WSV340/WSSV396), an envelope protein of white spot syndrome virus (WSSV), contains an Arg-Gly-Asp (RGD) peptide domain known as a cellular attachment site. At present, the process of VP31 interacting with shrimp host cells has not been explored. Therefore, the VP31 gene was cloned into pET30a (+), expressed in Escherichia coli strain BL21 and purified with immobilized metal ion affinity chromatography. Four gill cellular proteins of shrimp (Fenneropenaeus chinensis) were pulled down by an affinity column coupled with recombinant VP31 (rVP31), and the amino acid sequences were identified with MALDI-TOF/TOF mass spectrometry. Hemocyanin, beta-actin, arginine kinase (AK), and an unknown protein were suggested as the putative VP31 receptor proteins. SDS-PAGE showed that AK is the predominant binding protein of VP31. An i n vitro binding activity experiment indicated that recombinant AK's (rAK) binding activity with rVP31 is comparable to that with the same amount of WSSV. These results suggested that AK, as a member of the phosphagen kinase family, plays a role in WSSV infection. This is the first evidence showing that AK is a binding protein of VP31. Further studies on this topic will elucidate WSSV infection mechanism in the future.

  5. Functional Diversity of Human Mitochondrial J-proteins Is Independent of Their Association with the Inner Membrane Presequence Translocase.

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    Sinha, Devanjan; Srivastava, Shubhi; D'Silva, Patrick

    2016-08-12

    Mitochondrial J-proteins play a critical role in governing Hsp70 activity and, hence, are essential for organellar protein translocation and folding. In contrast to yeast, which has a single J-protein Pam18, humans involve two J-proteins, DnaJC15 and DnaJC19, associated with contrasting cellular phenotype, to transport proteins into the mitochondria. Mutation in DnaJC19 results in dilated cardiomyopathy and ataxia syndrome, whereas expression of DnaJC15 regulates the response of cancer cells to chemotherapy. In the present study we have comparatively assessed the biochemical properties of the J-protein paralogs in relation to their association with the import channel. Both DnaJC15 and DnaJC19 formed two distinct subcomplexes with Magmas at the import channel. Knockdown analysis suggested an essential role for Magmas and DnaJC19 in organellar protein translocation and mitochondria biogenesis, whereas DnaJC15 had dispensable supportive function. The J-proteins were found to have equal affinity for Magmas and could stimulate mitochondrial Hsp70 ATPase activity by equivalent levels. Interestingly, we observed that DnaJC15 exhibits bifunctional properties. At the translocation channel, it involves conserved interactions and mechanism to translocate the precursors into mitochondria. In addition to protein transport, DnaJC15 also showed a dual role in yeast where its expression elicited enhanced sensitivity of cells to cisplatin that required the presence of a functional J-domain. The amount of DnaJC15 expressed in the cell was directly proportional to the sensitivity of cells. Our analysis indicates that the differential cellular phenotype displayed by human mitochondrial J-proteins is independent of their activity and association with Magmas at the translocation channel. PMID:27330077

  6. Twin-arginine-dependent translocation of SufI in the absence of cytosolic helper proteins.

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    Holzapfel, Eva; Moser, Michael; Schiltz, Emile; Ueda, Takuya; Betton, Jean-Michel; Müller, Matthias

    2009-06-16

    The twin-arginine translocation (Tat) machinery present in bacterial and thylakoidal membranes is able to transport fully folded proteins. Folding of some Tat precursor proteins requires dedicated chaperones that also sequester the signal sequence during the maturation process. Whether or not signal sequence-binding chaperones are a general prerequisite for all Tat substrate proteins is not known. Here, we have studied the propensity of Tat signal sequences of Escherichia coli to interact with general chaperones and peptidyl-prolyl-cis,trans-isomerases. Site-specific photocross-linking revealed a clear specificity for FK506-binding proteins. Nevertheless transport of the Tat substrate SufI into inverted inner membrane vesicles of E. coli was found to occur in the bona fide absence of any cytosolic chaperone. Our results suggest that in E. coli, cytosolic chaperones are not essential for the twin-arginine-dependent export of cofactor-less substrates. PMID:19432418

  7. Study of the five Rickettsia prowazekii proteins annotated as ATP/ADP translocases (Tlc): Only Tlc1 transports ATP/ADP, while Tlc4 and Tlc5 transport other ribonucleotides.

    Science.gov (United States)

    Audia, Jonathon P; Winkler, Herbert H

    2006-09-01

    The obligate intracytoplasmic pathogen Rickettsia prowazekii relies on the transport of many essential compounds from the cytoplasm of the eukaryotic host cell in lieu of de novo synthesis, an evolutionary outcome undoubtedly linked to obligatory growth in this metabolite-replete niche. The paradigm for the study of rickettsial transport systems is the ATP/ADP translocase Tlc1, which exchanges bacterial ADP for host cell ATP as a source of energy, rather than as a source of adenylate. Interestingly, the R. prowazekii genome encodes four open reading frames that are highly homologous to the well-characterized ATP/ADP translocase Tlc1. Therefore, by annotation, the R. prowazekii genome encodes a total of five ATP/ADP translocases: Tlc1, Tlc2, Tlc3, Tlc4, and Tlc5. We have confirmed by quantitative reverse transcriptase PCR that mRNAs corresponding to all five tlc homologues are expressed in R. prowazekii growing in L-929 cells and have shown their heterologous protein expression in Escherichia coli, suggesting that none of the tlc genes are pseudogenes in the process of evolutionary meltdown. However, we demonstrate by heterologous expression in E. coli that only Tlc1 functions as an ATP/ADP transporter. A survey of nucleotides and nucleosides has determined that Tlc4 transports CTP, UTP, and GDP. Intriguingly, although GTP was not transported by Tlc4, it was an inhibitor of CTP and UTP uptake and demonstrated a K(i) similar to that of GDP. In addition, we demonstrate that Tlc5 transports GTP and GDP. We postulate that Tlc4 and Tlc5 serve the primary function of maintaining intracellular pools of nucleotides for rickettsial nucleic acid biosynthesis and do not provide the cell with nucleoside triphosphates as an energy source, as is the case for Tlc1. Although heterologous expression of Tlc2 and Tlc3 was observed in E. coli, we were unable to identify substrates for these proteins. PMID:16923893

  8. The bacterial Sec-translocase: structure and mechanism

    OpenAIRE

    Lycklama a Nijeholt, Jelger A.; Driessen, Arnold J. M.

    2012-01-01

    Most bacterial secretory proteins pass across the cytoplasmic membrane via the translocase, which consists of a protein-conducting channel SecYEG and an ATP-dependent motor protein SecA. The ancillary SecDF membrane protein complex promotes the final stages of translocation. Recent years have seen a major advance in our understanding of the structural and biochemical basis of protein translocation, and this has led to a detailed model of the translocation mechanism.

  9. Periplasmic Binding Proteins in Thermophiles: Characterization and Potential Application of an Arginine-Binding Protein from Thermotoga maritima: A Brief Thermo-Story

    Directory of Open Access Journals (Sweden)

    Sabato D'Auria

    2013-02-01

    Full Text Available Arginine-binding protein from the extremophile Thermotoga maritima is a 27.7 kDa protein possessing the typical two-domain structure of the periplasmic binding proteins family. The protein is characterized by a very high specificity and affinity to bind to arginine, also at high temperatures. Due to its features, this protein could be taken into account as a potential candidate for the design of a biosensor for arginine. It is important to investigate the stability of proteins when they are used for biotechnological applications. In this article, we review the structural and functional features of an arginine-binding protein from the extremophile Thermotoga maritima with a particular eye on its potential biotechnological applications.

  10. Structure of the TatC core of the twin-arginine protein transport system.

    Science.gov (United States)

    Rollauer, Sarah E; Tarry, Michael J; Graham, James E; Jääskeläinen, Mari; Jäger, Franziska; Johnson, Steven; Krehenbrink, Martin; Liu, Sai-Man; Lukey, Michael J; Marcoux, Julien; McDowell, Melanie A; Rodriguez, Fernanda; Roversi, Pietro; Stansfeld, Phillip J; Robinson, Carol V; Sansom, Mark S P; Palmer, Tracy; Högbom, Martin; Berks, Ben C; Lea, Susan M

    2012-12-13

    The twin-arginine translocation (Tat) pathway is one of two general protein transport systems found in the prokaryotic cytoplasmic membrane and is conserved in the thylakoid membrane of plant chloroplasts. The defining, and highly unusual, property of the Tat pathway is that it transports folded proteins, a task that must be achieved without allowing appreciable ion leakage across the membrane. The integral membrane TatC protein is the central component of the Tat pathway. TatC captures substrate proteins by binding their signal peptides. TatC then recruits TatA family proteins to form the active translocation complex. Here we report the crystal structure of TatC from the hyperthermophilic bacterium Aquifex aeolicus. This structure provides a molecular description of the core of the Tat translocation system and a framework for understanding the unique Tat transport mechanism. PMID:23201679

  11. Mutation of exposed hydrophobic amino acids to arginine to increase protein stability

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    Czaplicki Jerzy

    2004-07-01

    Full Text Available Abstract Background One strategy to increase the stability of proteins is to reduce the area of water-accessible hydrophobic surface. Results In order to test it, we replaced 14 solvent-exposed hydrophobic residues of acetylcholinesterase by arginine. The stabilities of the resulting proteins were tested using denaturation by high temperature, organic solvents, urea and by proteolytic digestion. Conclusion Altough the mutational effects were rather small, this strategy proved to be successful since half of the mutants showed an increased stability. This stability may originate from the suppression of unfavorable interactions of nonpolar residues with water or from addition of new hydrogen bonds with the solvent. Other mechanisms may also contribute to the increased stability observed with some mutants. For example, introduction of a charge at the surface of the protein may provide a new coulombic interaction on the protein surface.

  12. Formation of functional Tat translocases from heterologous components

    OpenAIRE

    Widdick David A; Buchanan Grant; Guymer David; Hicks Matthew G; Caldelari Isabelle; Berks Ben C; Palmer Tracy

    2006-01-01

    Abstract Background The Tat pathway transports folded proteins across the cytoplasmic membrane of bacteria and the thylakoid membrane of plants. In Eschericha coli, Tat transport requires the integral membrane proteins TatA, TatB and TatC. In this study we have tested the ability of tat genes from the eubacterial species Pseudomonas syringae, Streptomyces coelicolor and Aquifex aeolicus, to compensate for the absence of the cognate E. coli tat gene, and thus to form functional Tat translocase...

  13. Role of type II protein arginine methyltransferase 5 in the regulation of Circadian Per1 gene.

    Directory of Open Access Journals (Sweden)

    Jungtae Na

    Full Text Available Circadian clocks are the endogenous oscillators that regulate rhythmic physiological and behavioral changes to correspond to daily light-dark cycles. Molecular dissections have revealed that transcriptional feedback loops of the circadian clock genes drive the molecular oscillation, in which PER/CRY complexes inhibit the transcriptional activity of the CLOCK/BMAL1 heterodimer to constitute a negative feedback loop. In this study, we identified the type II protein arginine methyltransferase 5 (PRMT5 as an interacting molecule of CRY1. Although the Prmt5 gene was constitutively expressed, increased interaction of PRMT5 with CRY1 was observed when the Per1 gene was repressed both in synchronized mouse liver and NIH3T3 cells. Moreover, rhythmic recruitment of PRMT5 and CRY1 to the Per1 gene promoter was found to be associated with an increased level of histone H4R3 dimethylation and Per1 gene repression. Consistently, decreased histone H4R3 dimethylation and altered rhythmic Per1 gene expression were observed in Prmt5-depleted cells. Taken together, these findings provide an insight into the link between histone arginine methylation by PRMT5 and transcriptional regulation of the circadian Per1 gene.

  14. A rigorous method for multigenic families' functional annotation: the peptidyl arginine deiminase (PADs proteins family example

    Directory of Open Access Journals (Sweden)

    Blanc M

    2005-11-01

    Full Text Available Abstract Background large scale and reliable proteins' functional annotation is a major challenge in modern biology. Phylogenetic analyses have been shown to be important for such tasks. However, up to now, phylogenetic annotation did not take into account expression data (i.e. ESTs, Microarrays, SAGE, .... Therefore, integrating such data, like ESTs in phylogenetic annotation could be a major advance in post genomic analyses. We developed an approach enabling the combination of expression data and phylogenetic analysis. To illustrate our method, we used an example protein family, the peptidyl arginine deiminases (PADs, probably implied in Rheumatoid Arthritis. Results the analysis was performed as follows: we built a phylogeny of PAD proteins from the NCBI's NR protein database. We completed the phylogenetic reconstruction of PADs using an enlarged sequence database containing translations of ESTs contigs. We then extracted all corresponding expression data contained in EST database This analysis allowed us 1/To extend the spectrum of homologs-containing species and to improve the reconstruction of genes' evolutionary history. 2/To deduce an accurate gene expression pattern for each member of this protein family. 3/To show a correlation between paralogous sequences' evolution rate and pattern of tissular expression. Conclusion coupling phylogenetic reconstruction and expression data is a promising way of analysis that could be applied to all multigenic families to investigate the relationship between molecular and transcriptional evolution and to improve functional annotation.

  15. Obese women on a low energy rice and bean diet: effects of leucine, arginine or glycine supplementation on protein turnover

    Directory of Open Access Journals (Sweden)

    J.S. Marchini

    2001-10-01

    Full Text Available This study examined if leucine, arginine or glycine supplementation in adult obese patients (body mass index of 33 ± 4 kg/m² consuming a Brazilian low energy and protein diet (4.2 MJ/day and 0.6 g protein/kg affects protein and amino acid metabolism. After four weeks adaptation to this diet, each subject received supplements of these amino acids (equivalent to 0.2 g protein kg-1 day-1 in random order. On the seventh day of each amino acid supplementation, a single-dose 15N-glycine study was carried out. There were no significant differences in protein flux, synthesis or breakdown. The protein flux (grams of nitrogen, gN/9 h was 55 ± 24 during the nonsupplemented diet intake and 39 ± 10, 44 ± 22 and 58 ± 35 during the leucine-, glycine- and arginine-supplemented diet intake, respectively; protein synthesis (gN/9 h was 57 ± 24, 36 ± 10, 41 ± 22 and 56 ± 36, respectively; protein breakdown (gN/9 h was 51 ± 24, 34 ± 10, 32 ± 28 and 53 ± 35, respectively; kinetic balance (gN/9 h was 3.2 ± 1.8, 4.1 ± 1.7, 3.4 ± 2.9 and 3.9 ± 1.6. There was no difference in amino acid profiles due to leucine, arginine or glycine supplementation. The present results suggest that 0.6 g/kg of dietary protein is enough to maintain protein turnover in obese women consuming a reduced energy diet and that leucine, arginine or glycine supplementation does not change kinetic balance or protein synthesis.

  16. Expression and Characterization of ArgR, An Arginine Regulatory Protein in Corynebacterium crenatum

    Institute of Scientific and Technical Information of China (English)

    CHEN Xue Lan; ZHANG Bin; TANG Li; JIAO Hai Tao; XU Heng Yi; XU Feng; XU Hong; WEI Hua; XIONG Yong Hua

    2014-01-01

    Objective Corynebacterium crenatum MT, a mutant from C. crenatum AS 1.542 with a lethal argR gene, exhibits high arginine production. To confirm the effect of ArgR on arginine biosynthesis in C. crenatum, an intact argR gene from wild-type AS 1.542 was introduced into C. crenatum MT, resulting in C. crenatum MT. sp, and the changes of transcriptional levels of the arginine biosynthetic genes and arginine production were compared between the mutant strain and the recombinant strain. Methods Quantitative real-time polymerase chain reaction was employed to analyze the changes of the related genes at the transcriptional level, electrophoretic mobility shift assays were used to determine ArgR binding with the argCJBDF, argGH, and carAB promoter regions, and arginine production was determined with an automated amino acid analyzer. Results Arginine production assays showed a 69.9%reduction in arginine from 9.01±0.22 mg/mL in C. crenatum MT to 2.71±0.13 mg/mL (P Conclusion The arginine biosynthetic genes in C. crenatum are clearly controlled by the negative regulator ArgR, and intact ArgR in C. crenatum MT results in a significant descrease in arginine production.

  17. Structure of the twin-arginine signal-binding protein DmsD from Escherichia coli

    International Nuclear Information System (INIS)

    The crystal structure of DmsD from E. coli has been determined at 2.4 Å resolution. The translocation of folded proteins via the twin-arginine translocation (Tat) pathway is regulated to prevent the futile export of inactive substrate. DmsD is part of a class of cytoplasmic chaperones that play a role in preventing certain redox proteins from premature transport. DmsD from Escherichia coli has been crystallized in space group P41212, with unit-cell parameters a = b = 97.45, c = 210.04 Å, in the presence of a small peptide. The structure has been solved by molecular replacement to a resolution of 2.4 Å and refined to an R factor of 19.4%. There are four molecules in the asymmetric unit that may mimic a higher order structure in vivo. There appears to be density for the peptide in a predicted binding pocket, which lends support to its role as the signal-recognition surface for this class of proteins

  18. Modulation of Epstein–Barr Virus Nuclear Antigen 2-dependent transcription by protein arginine methyltransferase 5

    International Nuclear Information System (INIS)

    Highlights: ► Catalytic active PRMT5 substantially binds to the EBNA2 RG domain. ► PRMT5 augments the EBNA2-dependent transcription. ► PRMT5 triggers the symmetric dimethylation of the EBNA2 RG domain. ► PRMT5 enhances the promoter occupancy of EBNA2 on its target promoters. -- Abstract: Epstein–Barr Virus Nuclear Antigen (EBNA) 2 features an Arginine–Glycine repeat (RG) domain at amino acid positions 335–360, which is a known target for protein arginine methyltransferaser 5 (PRMT5). In this study, we performed protein affinity pull-down assays to demonstrate that endogenous PRMT5 derived from lymphoblastoid cells specifically associated with the protein bait GST-E2 RG. Transfection of a plasmid expressing PRMT5 induced a 2.5- to 3-fold increase in EBNA2-dependent transcription of both the LMP1 promoter in AKATA cells, which contain the EBV genome endogenously, and a Cp-Luc reporter plasmid in BJAB cells, which are EBV negative. Furthermore, we showed that there was a 2-fold enrichment of EBNA2 occupancy in target promoters in the presence of exogenous PRMT5. Taken together, we show that PRMT5 triggers the symmetric dimethylation of EBNA2 RG domain to coordinate with EBNA2-mediated transcription. This modulation suggests that PRMT5 may play a role in latent EBV infection

  19. Comparative analysis of twin-arginine (Tat)-dependent protein secretion of a heterologous model protein (GFP) in three different Gram-positive bacteria

    NARCIS (Netherlands)

    Meissner, Daniel; Vollstedt, Angela; van Dijl, Jan Maarten; Freudl, Roland

    2007-01-01

    In contrast to the general protein secretion (Sec) system, the twin-arginine translocation (Tat) export pathway allows the translocation of proteins across the bacterial plasma membrane in a fully folded conformation. Due to this feature, the Tat pathway provides an attractive alternative to the sec

  20. Protein arginine methyltransferase 5 (PRMT5) is a novel coactivator of constitutive androstane receptor (CAR).

    Science.gov (United States)

    Kanno, Yuichiro; 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. PMID:25721668

  1. Protein arginine methyltransferase 5 is associated with malignant phenotype and peritoneal metastasis in gastric cancer.

    Science.gov (United States)

    Kanda, Mitsuro; Shimizu, Dai; Fujii, Tsutomu; Tanaka, Haruyoshi; Shibata, Masahiro; Iwata, Naoki; Hayashi, Masamichi; Kobayashi, Daisuke; Tanaka, Chie; Yamada, Suguru; Nakayama, Goro; Sugimoto, Hiroyuki; Koike, Masahiko; Fujiwara, Michitaka; Kodera, Yasuhiro

    2016-09-01

    Identification of novel gastric cancer (GC)-related molecules is necessary to improve management of patients with GC in both diagnostic and therapeutic aspects. The aim of the present study was to determine whether protein arginine methyltransferase 5 (PRMT5) acts as an oncogene in the progression of GC and whether it serves as a novel diagnostic marker and therapeutic target. We conducted global expression profiling of GC cell lines and RNA interference experiments to evaluate the effect of PRMT5 expression on the phenotype of GC cells. We analysed tissues of 179 patients with GC to assess the association of PRMT5 mRNA levels with clinicopathological factors. Differential expression of PRMT5 mRNA by GC cell lines correlated positively with the levels of GEMIN2, STAT3 and TGFB3. PRMT5 knockdown reduced the proliferation, invasion and migration of a GC cell line. PRMT5 mRNA levels were significantly higher in GC tissues than the corresponding adjacent normal tissues and were independent of tumour depth, differentiation and lymph node metastasis. High PRMT5 expression was an independent risk factor of positive peritoneal lavage cytology (odds ratio 3.90, P=0.003) and decreased survival. PRMT5 enhances the malignant phenotype of GC cell lines and its expression in gastric tissues may serve as a biomarker for patient stratification and a potential target of therapy. PMID:27315569

  2. Novel Protein Arginine Methyltransferase 8 Isoform Is Essential for Cell Proliferation.

    Science.gov (United States)

    Hernandez, Sarah; Dominko, Tanja

    2016-09-01

    Identification of molecular mechanisms that regulate cellular replicative lifespan is needed to better understand the transition between a normal and a neoplastic cell phenotype. We have previously reported that low oxygen-mediated activity of FGF2 leads to an increase in cellular lifespan and acquisition of regeneration competence in human dermal fibroblasts (iRC cells). Though cells display a more plastic developmental phenotype, they remain non-tumorigenic when injected into SCID mice (Page et al. [2009] Cloning Stem Cells 11:417-426; Page et al. [2011] Eng Part A 17:2629-2640) allowing for investigation of mechanisms that regulate increased cellular lifespan in a non-tumorigenic system. Analysis of chromatin modification enzymes by qRT-PCR revealed a 13.3-fold upregulation of the arginine methyltransferase PRMT8 in iRC cells. Increased protein expression was confirmed in both iRC and human embryonic stem cells-the first demonstration of endogenous human PRMT8 expression outside the brain. Furthermore, iRC cells express a novel PRMT8 mRNA variant. Using siRNA-mediated knockdown we demonstrated that this novel variant was required for proliferation of human dermal fibroblasts (hDFs) and grade IV glioblastomas. PRMT8 upregulation in a non-tumorigenic system may offer a potential diagnostic biomarker and a therapeutic target for cells in pre-cancerous and cancerous states. J. Cell. Biochem. 117: 2056-2066, 2016. © 2016 Wiley Periodicals, Inc. PMID:26851891

  3. Protein arginine methyltransferase 5 (PRMT5) is a novel coactivator of constitutive androstane receptor (CAR)

    International Nuclear Information System (INIS)

    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

  4. 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.

  5. The Ergogenic Potential of Arginine

    Directory of Open Access Journals (Sweden)

    La Bounty Paul M

    2004-12-01

    Full Text Available Abstract Arginine is a conditionally essential amino acid that is involved in protein synthesis, the detoxification of ammonia, and its conversion to glucose as well as being catabolized to produce energy. In addition to these physiological functions, arginine has been purported to have ergogenic potential. Athletes have taken arginine for three main reasons: 1 its role in the secretion of endogenous growth hormone; 2 its involvement in the synthesis of creatine; 3 its role in augmenting nitric oxide. These aspects of arginine supplementation will be discussed as well as a review of clinical investigations involving exercise performance and arginine ingestion.

  6. Characterization of an ATP translocase identified in the plant pathogen, Candidatus Liberibacter asiaticus

    Science.gov (United States)

    ATP/ADP translocases allow for the transport of ATP across a lipid bilayer, which is normally impermeable to this molecule due to its size and charge. These transport proteins appear to be unique to mitochondria, plant plastids, and obligate-intracellular bacteria. Of the bacterial ATP/ADP translo...

  7. Formation of functional Tat translocases from heterologous components

    Directory of Open Access Journals (Sweden)

    Widdick David A

    2006-07-01

    Full Text Available Abstract Background The Tat pathway transports folded proteins across the cytoplasmic membrane of bacteria and the thylakoid membrane of plants. In Eschericha coli, Tat transport requires the integral membrane proteins TatA, TatB and TatC. In this study we have tested the ability of tat genes from the eubacterial species Pseudomonas syringae, Streptomyces coelicolor and Aquifex aeolicus, to compensate for the absence of the cognate E. coli tat gene, and thus to form functional Tat translocases with E. coli Tat components. Results All three subunits of the Tat system from the Gram positive organism Streptomyces coelicolor were able to form heterologous translocases with substantive Tat transport activity. However, only the TatA and TatB proteins of Pseudomonas syringae were able to functionally interact with the E. coli Tat system even though the two organisms are closely related. Of the Tat components from the phylogenetically distant hyperthermophillic bacterium Aquifex aeolicus only the TatA proteins showed any detectable level of heterologous functionality. The heterologously expressed TatA proteins of S. coelicolor and A. aeolicus were found exclusively in the membrane fraction. Conclusion Our results show that of the three Tat proteins, TatA is most likely to show cross-species complementation. By contrast, TatB and TatC do not always show cross-complementation, probably because they must recognise heterologous signal peptides. Since heterologously-expressed S. coelicolor TatA protein was functional and found only in the membrane fraction, it suggests that soluble forms of Streptomyces TatA reported by others do not play a role in protein export.

  8. l-Arginine grafted alginate hydrogel beads: A novel pH-sensitive system for specific protein delivery

    Directory of Open Access Journals (Sweden)

    Mohamed S. Mohy Eldin

    2015-05-01

    Full Text Available Novel pH-sensitive hydrogels based on l-arginine grafted alginate (Arg-g-Alg hydrogel beads were synthesized and utilized as a new carrier for protein delivery (BSA in specific pH media. l-arginine was grafted onto the polysaccharide backbone of virgin alginate via amine functions. Evidences of grafting of alginate were extracted from FT-IR and thermal analysis, while the morphological structure of Arg-g-Alg hydrogel beads was investigated by SEM photographs. Factors affecting on the grafting process e.g. l-arginine concentration, reaction time, reaction temperature, reaction pH, and crosslinking conditions, have been studied. Whereas, grafting efficiency of each factor was evaluated. Grafting of alginate has improved both thermal and morphological properties of Arg-g-Alg hydrogel beads. The swelling behavior of Arg-g-Alg beads was determined as a function of pH and compared with virgin calcium alginate beads. The cumulative in vitro release profiles of BSA loaded beads were studied at different pHs for simulating the physiological environments of the gastrointestinal tract. The amount of BSA released from neat alginate beads at pH 2 was almost 15% after 5 h, while the Arg-g-Alg beads at the same conditions were clearly higher than 45%, then it increased to 90% at pH 7.2. Accordingly, grafting of alginate has improved its release profile behavior particularly in acidic media. The preliminary results clearly suggested that the Arg-g-Alg hydrogel may be a potential candidate for polymeric carrier for oral delivery of protein or drugs.

  9. Study of the Five Rickettsia prowazekii Proteins Annotated as ATP/ADP Translocases (Tlc): Only Tlc1 Transports ATP/ADP, While Tlc4 and Tlc5 Transport Other Ribonucleotides

    OpenAIRE

    Audia, Jonathon P.; Winkler, Herbert H.

    2006-01-01

    The obligate intracytoplasmic pathogen Rickettsia prowazekii relies on the transport of many essential compounds from the cytoplasm of the eukaryotic host cell in lieu of de novo synthesis, an evolutionary outcome undoubtedly linked to obligatory growth in this metabolite-replete niche. The paradigm for the study of rickettsial transport systems is the ATP/ADP translocase Tlc1, which exchanges bacterial ADP for host cell ATP as a source of energy, rather than as a source of adenylate. Interes...

  10. Human Papillomavirus Type 1 E1^E4 Protein Is a Potent Inhibitor of the Serine-Arginine (SR) Protein Kinase SRPK1 and Inhibits Phosphorylation of Host SR Proteins and of the Viral Transcription and Replication Regulator E2

    OpenAIRE

    Prescott, Emma L.; Brimacombe, Claire L.; Hartley, Margaret; Bell, Ian; Graham, Sheila; Roberts, Sally

    2014-01-01

    The serine-arginine-specific protein kinase SRPK1 is a common binding partner of the E1^E4 protein of diverse human papillomavirus types. We show here for the first time that the interaction between HPV1 E1^E4 and SRPK1 leads to potent inhibition of SRPK1 phosphorylation of host serine-arginine (SR) proteins that have critical roles in mRNA metabolism, including pre-mRNA processing, mRNA export, and translation. Furthermore, we show that SRPK1 phosphorylates serine residues of SR/RS dipeptide...

  11. Guipi decoction effects on arginine vasopressin protein and gene expression in the hippocampus, ventromedial hypothalamic nucleus, and prefrontal lobe in rats with spleen deficiency

    Institute of Scientific and Technical Information of China (English)

    Huinan Qian; Xueqin Hu; Libo Shen

    2008-01-01

    BACKGROUND: Arginine vasopressin has been shown to enhance learning in experimental animal models.OBJECTIVE: To determine whether Guipi decoction enhances memory and learning by increasing arginine vasopressin levels, and to verify the influence of Guipi decoction on arginine vasopressin protein and gene expression in the hippocampal CAI region, prefrontal lobe cortex, and ventral nucleus of hypothalamus in rats with spleen deficiency.DESIGN, TIME AND SETTING: The randomized, neuropharmacological, control study was performed in the College of Basic Medical Sciences, Beijing University of Chinese Medicine between March 2002 and March 2005.MATERIALS: Sixty, healthy, male, Wistar rats were used to establish spleen deficiency models according to the traditional Chinese medicine principle of bitter drugs for purgation, improper diet, and overstrain. Arginine vasopressin-I polyclonal anti-rabbit antibody immunohistocbemistry kit and arginine vasopressin in situ hybridization kit were provided by Department of Neuroanatomy in Shanghai Second Military Medical University of Chinese PLA.METHODS: Sixty rats were divided into five groups at random: normal control (n = 11), model (n = 13), Guipi decoction (n = 12), recipe control A (n = 12), and recipe control B groups (n = 12). Rats in the latter four groups received 7.5 g/kg of the drugs by intragastric administration each morning, which comprised Dahuang, Houpu, and Zhishi, prepared at a ratio of 2:1 : 1. The rats were lasted every other day, but were allowed free access to water at all times. The rats were forced to swim in 25℃ water until fatigued. Rats in the Guipi decoction and two recipe control groups were intragastrically administered 7.5 g/kg Guipi decoction, Chaihu Shugan powder, and Tianwang Buxin pellets, respectively, each afternoon. Rats in the normal group were intragastrically administered the same amount of normal saline. All rats were treated for 6 weeks.MAIN OUTCOME MEASURES: At 6 weeks after drug

  12. Protein loop compaction and the origin of the effect of arginine and glutamic acid mixtures on solubility, stability and transient oligomerization of proteins.

    Science.gov (United States)

    Blobel, Jascha; Brath, Ulrika; Bernadó, Pau; Diehl, Carl; Ballester, Lidia; Sornosa, Alejandra; Akke, Mikael; Pons, Miquel

    2011-12-01

    Addition of a 50 mM mixture of L: -arginine and L: -glutamic acid (RE) is extensively used to improve protein solubility and stability, although the origin of the effect is not well understood. We present Small Angle X-ray Scattering (SAXS) and Nuclear Magnetic Resonance (NMR) results showing that RE induces protein compaction by collapsing flexible loops on the protein core. This is suggested to be a general mechanism preventing aggregation and improving resistance to proteases and to originate from the polyelectrolyte nature of RE. Molecular polyelectrolyte mixtures are expected to display long range correlation effects according to dressed interaction site theory. We hypothesize that perturbation of the RE solution by dissolved proteins is proportional to the volume occupied by the protein. As a consequence, loop collapse, minimizing the effective protein volume, is favored in the presence of RE. PMID:21390527

  13. Identification of a nuclear localization motif in the serine/arginine protein kinase PSRPK of physarum polycephalum

    Directory of Open Access Journals (Sweden)

    Tian Shengli

    2009-08-01

    Full Text Available Abstract Background Serine/arginine (SR protein-specific kinases (SRPKs are conserved in a wide range of organisms, from humans to yeast. Studies showed that SRPKs can regulate the nuclear import of SR proteins in cytoplasm, and regulate the sub-localization of SR proteins in the nucleus. But no nuclear localization signal (NLS of SRPKs was found. We isolated an SRPK-like protein PSRPK (GenBank accession No. DQ140379 from Physarum polycephalum previously, and identified a NLS of PSRPK in this study. Results We carried out a thorough molecular dissection of the different domains of the PSRPK protein involved in its nuclear localization. By truncation of PSRPK protein, deletion of and single amino acid substitution in a putative NLS and transfection of mammalian cells, we observed the distribution of PSRPK fluorescent fusion protein in mammalian cells using confocal microscopy and found that the protein was mainly accumulated in the nucleus; this indicated that the motif contained a nuclear localization signal (NLS. Further investigation with truncated PSPRK peptides showed that the NLS (318PKKGDKYDKTD328 was localized in the alkaline Ω-loop of a helix-loop-helix motif (HLHM of the C-terminal conserved domain. If the 318PKKGDK322 sequence was deleted from the loop or K320 was mutated to T320, the PSRPK fluorescent fusion protein could not enter and accumulate in the nucleus. Conclusion This study demonstrated that the 318PKKGDKYDKTD328 peptides localized in the C-terminal conserved domain of PSRPK with the Ω-loop structure could play a crucial role in the NLS function of PSRPK.

  14. Two proteins with ornithine acetyltransferase activity show different functions in Streptomyces clavuligerus: Oat2 modulates clavulanic acid biosynthesis in response to arginine.

    Science.gov (United States)

    de la Fuente, A; Martín, J F; Rodríguez-García, A; Liras, P

    2004-10-01

    The oat2 gene, located in the clavulanic acid gene cluster in Streptomyces clavuligerus, is similar to argJ, which encodes N-acetylornithine:glutamic acid acetyltransferase activity. Purified proteins obtained by expression in Escherichia coli of the argJ and oat2 genes of S. clavuligerus posses N-acetyltransferase activity. The kinetics and substrate specificities of both proteins are very similar. Deletion of the oat2 gene did not affect the total N-acetylornithine transferase activity and slightly reduced the formation of clavulanic acid under standard culture conditions. However, the oat2 mutant produced more clavulanic acid than the parental strain in cultures supplemented with high levels (above 1 mM) of arginine. The purified S. clavuligerus ArgR protein bound the arginine box in the oat2 promoter, and the expression of oat2 was higher in mutants with a disruption in argR (arginine-deregulated), confirming that the Arg boxes of oat2 are functional in vivo. Our results suggest that the Oat2 protein or one of its reaction products has a regulatory role that modulates clavulanic acid biosynthesis in response to high arginine concentrations. PMID:15375131

  15. Identification of the methylation preference region in heterogeneous nuclear ribonucleoprotein K by protein arginine methyltransferase 1 and its implication in regulating nuclear/cytoplasmic distribution

    International Nuclear Information System (INIS)

    Research highlights: → Verifying by direct methylation assay the substrate sites of PRMT1 in the hnRNP K protein. → Identifying the preferred PMRT1 methylation regions in hnRNP K by kinetic analysis. → Linking methylation in regulating nuclear localization of hnRNP K. -- Abstract: Protein arginine methylation plays crucial roles in numerous cellular processes. Heterogeneous nuclear ribonucleoprotein K (hnRNP K) is a multi-functional protein participating in a variety of cellular functions including transcription and RNA processing. HnRNP K is methylated at multiple sites in the glycine- and arginine-rich (RGG) motif. Using various RGG domain deletion mutants of hnRNP K as substrates, here we show by direct methylation assay that protein arginine methyltransferase 1 (PRMT1) methylated preferentially in a.a. 280-307 of the RGG motif. Kinetic analysis revealed that deletion of a.a. 280-307, but not a.a. 308-327, significantly inhibited rate of methylation. Importantly, nuclear localization of hnRNP K was significantly impaired in mutant hnRNP K lacking the PRMT1 methylation region or upon pharmacological inhibition of methylation. Together our results identify preferred PRMT1 methylation sequences of hnRNP K by direct methylation assay and implicate a role of arginine methylation in regulating intracellular distribution of hnRNP K.

  16. Arginine transport in catabolic disease states.

    Science.gov (United States)

    Pan, Ming; Choudry, Haroon A; Epler, Mark J; Meng, Qinghe; Karinch, Anne; Lin, Chengmao; Souba, Wiley

    2004-10-01

    Arginine appears to be a semiessential amino acid in humans during critical illness. Catabolic disease states such as sepsis, injury, and cancer cause an increase in arginine utilization, which exceeds body production, leading to arginine depletion. This is aggravated by the reduced nutrient intake that is associated with critical illness. Arginine depletion may have negative consequences on tissue function under these circumstances. Nutritional regimens containing arginine have been shown to improve nitrogen balance and lymphocyte function, and stimulate arginine transport in the liver. We have studied the effects of stress mediators on arginine transport in vascular endothelium, liver, and gut epithelium. In vascular endothelium, endotoxin stimulates arginine uptake, an effect that is mediated by the cytokine tumor necrosis factor-alpha (TNF-alpha) and by the cyclo-oxygenase pathway. This TNF-alpha stimulation involves the activation of intracellular protein kinase C (PKC). A significant increase in hepatic arginine transport activity also occurs following burn injury and in rats with progressive malignant disease. Surgical removal of the growing tumor results in a normalization of the accelerated hepatic arginine transport within days. Chronic metabolic acidosis and sepsis individually augment intestinal arginine transport in rats and Caco-2 cell culture. PKC and mitogen-activated protein kinases are involved in mediating the sepsis/acidosis stimulation of arginine transport. Understanding the regulation of plasma membrane arginine transport will enhance our knowledge of nutrition and metabolism in seriously ill patients and may lead to the design of improved nutritional support formulas. PMID:15465794

  17. Arginine and nitrogen storage.

    Science.gov (United States)

    Llácer, José L; Fita, Ignacio; Rubio, Vicente

    2008-12-01

    When nitrogen is abundant, prokaryotic and eukaryotic oxygen-producing photosynthetic organisms store nitrogen as arginine, by relieving feedback inhibition of the arginine biosynthesis controlling enzyme, N-acetylglutamate kinase (NAGK). The signalling protein PII, an ancient and widely distributed nitrogen/carbon/ADP/ATP sensor, mediates feedback inhibition relief of NAGK by binding to this enzyme. PII phosphorylation or PII binding of ADP or 2-oxoglutarate prevents PII-NAGK complex formation. Crystal structures of NAGK, cyanobacterial and plant PII and corresponding PII-NAGK complexes have been recently determined. In these complexes, two polar PII trimers sandwich one ring-like NAGK hexamer. Each PII subunit contacts one NAGK subunit, triggering a symmetry-restricted narrowing of the NAGK ring, with concomitant adoption by the arginine sites of a low-affinity conformation. PMID:19013524

  18. Effect of counter ions of arginine as an additive for the solubilization of protein and aromatic compounds.

    Science.gov (United States)

    Yoshizawa, Shunsuke; Arakawa, Tsutomu; Shiraki, Kentaro

    2016-10-01

    Arginine is widely used in biotechnological application, but mostly with chloride counter ion. Here, we examined the effects of various anions on solubilization of aromatic compounds and reduced lysozyme and on refolding of the lysozyme. All arginine salts tested increased the solubility of propyl gallate with acetate much more effectively than chloride. The effects of arginine salts were compared with those of sodium or guanidine salts, indicating that the ability of anions to modulate the propyl gallate solubility is independent of the cation. Comparison of transfer free energy of propyl gallate between sodium and arginine salts indicates that the interaction of propyl gallate is more favorable with arginine than sodium. On the contrary, the solubility of aromatic amino acids is only slightly modulated by anions, implying that there is specific interaction between acetic acid and propyl gallate. Unlike their effects on the solubility of small aromatic compounds, the solubility of reduced lysozyme was much higher in arginine chloride than in arginine acetate or sulfate. Consistent with high solubility, refolding of reduced lysozyme was most effective in arginine chloride. These results suggest potential broader applications of arginine modulated by different anions. PMID:27234496

  19. Redox specificity of 2-hydroxyacid-coupled NAD(+/NADH dehydrogenases: a study exploiting "reactive" arginine as a reporter of protein electrostatics.

    Directory of Open Access Journals (Sweden)

    Pooja Gupta

    Full Text Available With "reactive" arginine as a kinetic reporter, 2-hydroxyacid dehydrogenases are assessed in basis of their specialization as NAD(+-reducing or NADH-oxidizing enzymes. Specifically, M4 and H4 lactate dehydrogenases (LDHs and cytoplasmic and mitochondrial malate dehydrogenases (MDHs are compared to assess if their coenzyme specificity may involve electrostatics of cationic or neutral nicotinamide structure as the basis. The enzymes from diverse eukaryote and prokaryote sources thus are assessed in "reactivity" of functionally-critical arginine as a function of salt concentration and pH. Electrostatic calculations were performed on "reactive" arginines and found good correspondence with experiment. The reductive and oxidative LDHs and MDHs are assessed in their count over ionizable residues and in placement details of the residues in their structures as proteins. The variants found to be high or low in ΔpKa of "reactive" arginine are found to be also strong or weak cations that preferentially oxidize NADH (neutral nicotinamide structure or reduce NAD(+ (cationic nicotinamide structure. The ionized groups of protein structure may thus be important to redox specificity of the enzyme on basis of electrostatic preference for the oxidized (cationic nicotinamide or reduced (neutral nicotinamide coenzyme. Detailed comparisons of isozymes establish that the residues contributing in their redox specificity are scrambled in structure of the reductive enzyme.

  20. Evolutionary conservation of dual Sec translocases in the cyanelles of Cyanophora paradoxa

    Directory of Open Access Journals (Sweden)

    Löffelhardt Wolfgang

    2008-11-01

    Full Text Available Abstract Background Cyanelles, the peptidoglycan-armored plastids of glaucocystophytes, occupy a unique bridge position in between free-living cyanobacteria and chloroplasts. In some respects they side with cyanobacteria whereas other features are clearly shared with chloroplasts. The Sec translocase, an example for "conservative sorting" in the course of evolution, is found in the plasma membrane of all prokaryotes, in the thylakoid membrane of chloroplasts and in both these membrane types of cyanobacteria. Results In this paper we present evidence for a dual location of the Sec translocon in the thylakoid as well as inner envelope membranes of the cyanelles from Cyanophora paradoxa, i. e. conservative sorting sensu stricto. The prerequisite was the generation of specific antisera directed against cyanelle SecY that allowed immunodetection of the protein on SDS gels from both membrane types separated by sucrose density gradient floatation centrifugation. Immunoblotting of blue-native gels yielded positive but differential results for both the thylakoid and envelope Sec complexes, respectively. In addition, heterologous antisera directed against components of the Toc/Tic translocons and binding of a labeled precursor protein were used to discriminate between inner and outer envelope membranes. Conclusion The envelope translocase can be envisaged as a prokaryotic feature missing in higher plant chloroplasts but retained in cyanelles, likely for protein transport to the periplasm. Candidate passengers are cytochrome c6 and enzymes of peptidoglycan metabolism. The minimal set of subunits of the Toc/Tic translocase of a primitive plastid is proposed.

  1. Identification in Lupin Seed of a Serine-Endopeptidase Activity Cleaving between Twin Arginine Pairs and Causing Limited Proteolysis of Seed Storage Proteins

    Institute of Scientific and Technical Information of China (English)

    Chiara Magni; Fabio Sessa; Gabriella Tedeschi; Armando Negri; Alessio Scarafoni; Alessandro Consonni; Marcello Duranti

    2012-01-01

    The occurrence of twin-arginine motifs (-R-R-) in the amino acid sequences of animal pro-proteins frequently defines the cleavage site(s) for their structural/functional maturation.No information is available on the presence and possible biological meaning of these motifs in the seed storage proteins.In this work,a novel endopeptidase activity with cleavage specificity to twin-arginine pairs has been detected in mature dry Lupinus albus seeds.The endopeptidase was tested with a number of endogenous and exogenous protein substrates,which were selected according to the presence of one or more twin-arginine residue motifs in their amino acid sequences.The observed hydrolysis patterns were limited and highly specific.Partial proteolysis led to stable polypeptide fragments that were characterized by 1-and 2-D electrophoresis.Selected polypeptides were submitted to N-terminal amino acid sequencing and mass spectrometry analyses.These approaches,supported by bioinformatic analysis of the available sequences,allowed the conclusion that the polypeptide cleavage events had occurred at the peptide bonds comprised between twin-arginine residue pairs with all tested protein substrates.The endopeptidase activity was inhibited by 4-(2-AminoEthyl)Benzene-Sulphonyl Fluoride hydrochloride (AEBSF),leupeptin,and serine proteinase protein inhibitors,while it was not affected by pepstatin,transEpoxysuccinyI-L-leucylamido(4-guanidino)butane (E64),and ethylenediaminetetraacetic acid (EDTA),thus qualifying the Arg-Arg cleaving enzyme as a serine endopeptidase.The structural features of storage proteins from lupin and other legume seeds strongly support the hypothesis that the occurrence of an endopeptidase activity cleaving-R-R-bonds may be functional to facilitate their degradation at germination and possibly generate polypeptide fragments with specific biological activity.

  2. Uncovering the protein lysine and arginine methylation network in Arabidopsis chloroplasts.

    Science.gov (United States)

    Alban, Claude; Tardif, Marianne; Mininno, Morgane; Brugière, Sabine; Gilgen, Annabelle; Ma, Sheng; Mazzoleni, Meryl; Gigarel, Océane; Martin-Laffon, Jacqueline; Ferro, Myriam; Ravanel, Stéphane

    2014-01-01

    Post-translational modification of proteins by the addition of methyl groups to the side chains of Lys and Arg residues is proposed to play important roles in many cellular processes. In plants, identification of non-histone methylproteins at a cellular or subcellular scale is still missing. To gain insights into the extent of this modification in chloroplasts we used a bioinformatics approach to identify protein methyltransferases targeted to plastids and set up a workflow to specifically identify Lys and Arg methylated proteins from proteomic data used to produce the Arabidopsis chloroplast proteome. With this approach we could identify 31 high-confidence Lys and Arg methylation sites from 23 chloroplastic proteins, of which only two were previously known to be methylated. These methylproteins are split between the stroma, thylakoids and envelope sub-compartments. They belong to essential metabolic processes, including photosynthesis, and to the chloroplast biogenesis and maintenance machinery (translation, protein import, division). Also, the in silico identification of nine protein methyltransferases that are known or predicted to be targeted to plastids provided a foundation to build the enzymes/substrates relationships that govern methylation in chloroplasts. Thereby, using in vitro methylation assays with chloroplast stroma as a source of methyltransferases we confirmed the methylation sites of two targets, plastid ribosomal protein L11 and the β-subunit of ATP synthase. Furthermore, a biochemical screening of recombinant chloroplastic protein Lys methyltransferases allowed us to identify the enzymes involved in the modification of these substrates. The present study provides a useful resource to build the methyltransferases/methylproteins network and to elucidate the role of protein methylation in chloroplast biology. PMID:24748391

  3. Uncovering the Protein Lysine and Arginine Methylation Network in Arabidopsis Chloroplasts

    OpenAIRE

    Alban, Claude; Tardif, Marianne; Mininno, Morgane; Brugiere, Sabine; Gilgen, Annabelle; Ma, Sheng; Mazzoleni, Meryl; Gigarel, Oceane; Martin-Laffon, Jacqueline; Ferro, Myriam

    2014-01-01

    Post-translational modification of proteins by the addition of methyl groups to the side chains of Lys and Arg residues is proposed to play important roles in many cellular processes. In plants, identification of non-histone methylproteins at a cellular or subcellular scale is still missing. To gain insights into the extent of this modification in chloroplasts we used a bioinformatics approach to identify protein methyltransferases targeted to plastids and set up a workflow to specifically id...

  4. Identification of a novel antimicrobial peptide from human hepatitis B virus core protein arginine-rich domain (ARD.

    Directory of Open Access Journals (Sweden)

    Heng-Li Chen

    Full Text Available The rise of multidrug-resistant (MDR pathogens causes an increasing challenge to public health. Antimicrobial peptides are considered a possible solution to this problem. HBV core protein (HBc contains an arginine-rich domain (ARD at its C-terminus, which consists of 16 arginine residues separated into four clusters (ARD I to IV. In this study, we demonstrated that the peptide containing the full-length ARD I-IV (HBc147-183 has a broad-spectrum antimicrobial activity at micro-molar concentrations, including some MDR and colistin (polymyxin E-resistant Acinetobacter baumannii. Furthermore, confocal fluorescence microscopy and SYTOX Green uptake assay indicated that this peptide killed Gram-negative and Gram-positive bacteria by membrane permeabilization or DNA binding. In addition, peptide ARD II-IV (HBc153-176 and ARD I-III (HBc147-167 were found to be necessary and sufficient for the activity against P. aeruginosa and K. peumoniae. The antimicrobial activity of HBc ARD peptides can be attenuated by the addition of LPS. HBc ARD peptide was shown to be capable of direct binding to the Lipid A of lipopolysaccharide (LPS in several in vitro binding assays. Peptide ARD I-IV (HBc147-183 had no detectable cytotoxicity in various tissue culture systems and a mouse animal model. In the mouse model by intraperitoneal (i.p. inoculation with Staphylococcus aureus, timely treatment by i.p. injection with ARD peptide resulted in 100-fold reduction of bacteria load in blood, liver and spleen, as well as 100% protection of inoculated animals from death. If peptide was injected when bacterial load in the blood reached its peak, the protection rate dropped to 40%. Similar results were observed in K. peumoniae using an IVIS imaging system. The finding of anti-microbial HBc ARD is discussed in the context of commensal gut microbiota, development of intrahepatic anti-viral immunity and establishment of chronic infection with HBV. Our current results suggested that

  5. The Critical Role of Protein Arginine Methyltransferase prmt8 in Zebrafish Embryonic and Neural Development Is Non-Redundant with Its Paralogue prmt1

    OpenAIRE

    Lin, Yu-Ling; Tsai, Yun-Jung; Liu, Yu-Fang; Cheng, Yi-Chuan; Hung, Chuan-Mao; Lee, Yi-Jen; Pan, Huichin; Li, Chuan

    2013-01-01

    Protein arginine methyltransferase (PRMT) 1 is the most conserved and widely distributed PRMT in eukaryotes. PRMT8 is a vertebrate-restricted paralogue of PRMT1 with an extra N-terminal sequence and brain-specific expression. We use zebrafish (Danio rerio) as a vertebrate model to study PRMT8 function and putative redundancy with PRMT1. The transcripts of zebrafish prmt8 were specifically expressed in adult zebrafish brain and ubiquitously expressed from zygotic to early segmentation stage be...

  6. A Proline/Arginine-Rich End Leucine-Rich Repeat Protein (PRELP) Variant Is Uniquely Expressed in Chronic Lymphocytic Leukemia Cells

    OpenAIRE

    Eva Mikaelsson; Anders Österborg; Mahmood Jeddi-Tehrani; Parviz Kokhaei; Mahyar Ostadkarampour; Reza Hadavi; Mehran Gholamin; Mehdi Akhondi; Fazel Shokri; Hodjattallah Rabbani; Håkan Mellstedt

    2013-01-01

    Proline/arginine-rich end leucine-rich repeat protein (PRELP) belongs to the small leucine-rich proteoglycan (SLRP) family, normally expressed in extracellular matrix of collagen-rich tissues. We have previously reported on another SLRP, fibromodulin (FMOD) in patients with chronic lymphocytic leukemia (CLL). PRELP is structurally similar to FMOD with adjacent localization on chromosome 1 (1q32.1). As cluster-upregulation of genes may occur in malignancies, the aim of our study was to analyze...

  7. Mass Spectrometric Identification of the Arginine and Lysine deficient Proline Rich Glutamine Rich Wheat Storage Proteins

    Science.gov (United States)

    Tandem mass spectrometry (MS/MS) of enzymatic digest has made possible identification of a wide variety of proteins and complex samples prepared by such techniques as RP-HPLC or 2-D gel electrophoresis. Success requires peptide fragmentation to be indicative of the peptide amino acid sequence. The f...

  8. Variable stoichiometry of the TatA component of the twin-arginine protein transport system observed by in vivo single-molecule imaging

    OpenAIRE

    Leake, Mark C.; Greene, Nicholas P.; Godun, Rachel M.; Granjon, Thierry; Buchanan, Grant; Chen, Shuyun; Berry, Richard M; Palmer, Tracy; Berks, Ben C.

    2008-01-01

    The twin-arginine translocation (Tat) system transports folded proteins across the bacterial cytoplasmic membrane and the thylakoid membrane of plant chloroplasts. The essential components of the Tat pathway are the membrane proteins TatA, TatB, and TatC. TatA is thought to form the protein translocating element of the Tat system. Current models for Tat transport make predictions about the oligomeric state of TatA and whether, and how, this state changes during the transport cycle. We determi...

  9. Arginine residues at internal positions in a protein are always charged

    OpenAIRE

    Harms, Michael J.; Schlessman, Jamie L.; Sue, Gloria R; García-Moreno E., Bertrand

    2011-01-01

    Many functionally essential ionizable groups are buried in the hydrophobic interior of proteins. A systematic study of Lys, Asp, and Glu residues at 25 internal positions in staphylococcal nuclease showed that their pKa values can be highly anomalous, some shifted by as many as 5.7 pH units relative to normal pKa values in water. Here we show that, in contrast, Arg residues at the same internal positions exhibit no detectable shifts in pKa; they are all charged at pH ≤ 10. Twenty-three of the...

  10. Supplemental L-arginine Modulates Developmental Pulmonary Hypertension in Broiler Chickens Fed Reduced-Protein Diets and Reared at High Altitude

    Directory of Open Access Journals (Sweden)

    Sharifi MR

    2015-06-01

    Full Text Available This experiment was conducted to evaluate the effects of supplemental L-arginine (ARG in reduced-protein diets on cardiopulmonary performance and intestinal morphology in the broilers reared at high altitude. A total of 156 day-old male broilers were randomly assigned to 3 treatments and 4 replicates of 13 chicks and reared up to 42 days of age. Treatment groups were designed as a normal-protein diet (NPD, a reduced-protein diet (RPD with 30 g/Kg less crude protein compared to the NPD, and a reduced-protein diet plus 4 g/Kg L-arginine (RPD + ARG. There were no significant differences among dietary treatments for intestinal morphology and weight gain. Feed conversion ratio was improved in the chickens fed RPD + ARG compared to those fed RPD alone. The right to total ventricular weight ratio (RV:TV was significantly increased in the chickens fed RPD when compared to those fed NPD or RPD + ARG. Serum nitric oxide and amplitude of the S waves of electrocardiogram significantly declined by reducing dietary protein content. Relative expression of endothelin-1 (ET-1 gene was higher in the heart and lungs of chicks fed RPD than those fed NPD and it was off set when ARG supplemented to RPD (P. In conclusion, supplementing reduced-protein diets with ARG would be an effective strategy to prevent the development of pulmonary hypertension by increase in nitric oxide, and decrease in RV:TV and ET-1 gene expression.

  11. Genetic evidence for a tight cooperation of TatB and TatC during productive recognition of twin-arginine (Tat signal peptides in Escherichia coli.

    Directory of Open Access Journals (Sweden)

    Frank Lausberg

    Full Text Available The twin arginine translocation (Tat pathway transports folded proteins across the cytoplasmic membrane of bacteria. Tat signal peptides contain a consensus motif (S/T-R-R-X-F-L-K that is thought to play a crucial role in substrate recognition by the Tat translocase. Replacement of the phenylalanine at the +2 consensus position in the signal peptide of a Tat-specific reporter protein (TorA-MalE by aspartate blocked export of the corresponding TorA(D(+2-MalE precursor, indicating that this mutation prevents a productive binding of the TorA(D(+2 signal peptide to the Tat translocase. Mutations were identified in the extreme amino-terminal regions of TatB and TatC that synergistically suppressed the export defect of TorA(D(+2-MalE when present in pairwise or triple combinations. The observed synergistic suppression activities were even more pronounced in the restoration of membrane translocation of another export-defective precursor, TorA(KQ-MalE, in which the conserved twin arginine residues had been replaced by lysine-glutamine. Collectively, these findings indicate that the extreme amino-terminal regions of TatB and TatC cooperate tightly during recognition and productive binding of Tat-dependent precursor proteins and, furthermore, that TatB and TatC are both involved in the formation of a specific signal peptide binding site that reaches out as far as the end of the TatB transmembrane segment.

  12. 5′-Single-stranded/duplex DNA junctions are loading sites for E. coli UvrD translocase

    OpenAIRE

    Tomko, Eric J.; Jia, Haifeng; Park, Jeehae; Maluf, Nasib K.; Ha, Taekjip; Lohman, Timothy M.

    2010-01-01

    Single-molecule analyses of the dual DNA helicase/translocase enzyme UvrD reveal the particular substrate preferences of its monomeric, translocase form, with implications for understanding UvrD's translocase and anti-recombinase roles.

  13. Serine-arginine protein kinase 1 (SRPK1) inhibition as a potential novel targeted therapeutic strategy in prostate cancer.

    Science.gov (United States)

    Mavrou, A; Brakspear, K; Hamdollah-Zadeh, M; Damodaran, G; Babaei-Jadidi, R; Oxley, J; Gillatt, D A; Ladomery, M R; Harper, S J; Bates, D O; Oltean, S

    2015-08-13

    Angiogenesis is required for tumour growth and is induced principally by vascular endothelial growth factor A (VEGF-A). VEGF-A pre-mRNA is alternatively spliced at the terminal exon to produce two families of isoforms, pro- and anti-angiogenic, only the former of which is upregulated in prostate cancer (PCa). In renal epithelial cells and colon cancer cells, the choice of VEGF splice isoforms is controlled by the splicing factor SRSF1, phosphorylated by serine-arginine protein kinase 1 (SRPK1). Immunohistochemistry staining of human samples revealed a significant increase in SRPK1 expression both in prostate intra-epithelial neoplasia lesions as well as malignant adenocarcinoma compared with benign prostate tissue. We therefore tested the hypothesis that the selective upregulation of pro-angiogenic VEGF in PCa may be under the control of SRPK1 activity. A switch in the expression of VEGF165 towards the anti-angiogenic splice isoform, VEGF165b, was seen in PC-3 cells with SRPK1 knockdown (KD). PC-3 SRPK1-KD cells resulted in tumours that grew more slowly in xenografts, with decreased microvessel density. No effect was seen as a result of SRPK1-KD on growth, proliferation, migration and invasion capabilities of PC-3 cells in vitro. Small-molecule inhibitors of SRPK1 switched splicing towards the anti-angiogenic isoform VEGF165b in PC-3 cells and decreased tumour growth when administered intraperitoneally in an orthotopic mouse model of PCa. Our study suggests that modulation of SRPK1 and subsequent inhibition of tumour angiogenesis by regulation of VEGF splicing can alter prostate tumour growth and supports further studies for the use of SRPK1 inhibition as a potential anti-angiogenic therapy in PCa. PMID:25381816

  14. FlpS, the FNR-Like Protein of Streptococcus suis Is an Essential, Oxygen-Sensing Activator of the Arginine Deiminase System

    Directory of Open Access Journals (Sweden)

    Jörg Willenborg

    2016-07-01

    Full Text Available Streptococcus (S. suis is a zoonotic pathogen causing septicemia and meningitis in pigs and humans. During infection S. suis must metabolically adapt to extremely diverse environments of the host. CcpA and the FNR family of bacterial transcriptional regulators are important for metabolic gene regulation in various bacteria. The role of CcpA in S. suis is well defined, but the function of the FNR-like protein of S. suis, FlpS, is yet unknown. Transcriptome analyses of wild-type S. suis and a flpS mutant strain suggested that FlpS is involved in the regulation of the central carbon, arginine degradation and nucleotide metabolism. However, isotopologue profiling revealed no substantial changes in the core carbon and amino acid de novo biosynthesis. FlpS was essential for the induction of the arcABC operon of the arginine degrading pathway under aerobic and anaerobic conditions. The arcABC-inducing activity of FlpS could be associated with the level of free oxygen in the culture medium. FlpS was necessary for arcABC-dependent intracellular bacterial survival but redundant in a mice infection model. Based on these results, we propose that the core function of S. suis FlpS is the oxygen-dependent activation of the arginine deiminase system.

  15. The RNA-binding protein HOS5 and serine/arginine-rich proteins RS40 and RS41 participate in miRNA biogenesis in Arabidopsis

    KAUST Repository

    Chen, Tao

    2015-07-30

    MicroRNAs are a class of small regulatory RNAs that are generated from primary miRNA (pri-miRNA) transcripts with a stem-loop structure. Accuracy of the processing of pri-miRNA into mature miRNA in plants can be enhanced by SERRATE (SE) and HYPONASTIC LEAVES 1 (HYL1). HYL1 activity is regulated by the FIERY2 (FRY2)/RNA polymerase II C-terminal domain phosphatase-like 1 (CPL1). Here, we discover that HIGH OSMOTIC STRESS GENE EXPRESSION 5 (HOS5) and two serine/arginine-rich splicing factors RS40 and RS41, previously shown to be involved in pre-mRNA splicing, affect the biogenesis of a subset of miRNA. These proteins are required for correct miRNA strand selection and the maintenance of miRNA levels. FRY2 dephosphorylates HOS5 whose phosphorylation status affects its subnuclear localization. HOS5 and the RS proteins bind both intronless and intron-containing pri-miRNAs. Importantly, all of these splicing-related factors directly interact with both HYL1 and SE in nuclear splicing speckles. Our results indicate that these splicing factors are directly involved in the biogenesis of a group of miRNA.

  16. The Hydrophobic Region of the DmsA Twin-Arginine Leader Peptide Determines Specificity with Chaperone DmsD

    OpenAIRE

    Winstone, Tara M. L.; Tran, Vy A.; Turner, Raymond J.

    2013-01-01

    The system specific chaperone DmsD plays a role in the maturation of the catalytic subunit of dimethyl sulfoxide (DMSO) reductase, DmsA. Pre-DmsA contains a 45-amino acid twin-arginine leader peptide that is important for targeting and translocation of folded and cofactor-loaded DmsA by the twin-arginine translocase. DmsD has previously been shown to interact with the complete twin-arginine leader peptide of DmsA. In this study, isothermal titration calorimetry was used to investigate the the...

  17. Response of arginine vasopressin-enhanced green fluorescent protein fusion gene in the hypothalamus of adjuvant-induced arthritic rats

    Czech Academy of Sciences Publication Activity Database

    Suzuki, H.; Onaka, T.; Kasai, M.; Kawasaki, M.; Ohnishi, H.; Otsubo, H.; Saito, T.; Hashimoto, H.; Yokoyama, T.; Fujihara, H.; Dayanithi, Govindan; Murphy, D.; Nakamura, T.; Ueta, Y.

    2009-01-01

    Roč. 21, č. 3 (2009), s. 183-190. ISSN 0953-8194 Institutional research plan: CEZ:AV0Z50390512 Keywords : arginine vasopressin * Corticotrophin-releasing hormone * GFP Subject RIV: FH - Neurology Impact factor: 3.700, year: 2009

  18. Basis of Arginine Sensitivity of Microbial N-Acetyl-l-Glutamate Kinases: Mutagenesis and Protein Engineering Study with the Pseudomonas aeruginosa and Escherichia coli Enzymes▿

    Science.gov (United States)

    Fernández-Murga, M. Leonor; Rubio, Vicente

    2008-01-01

    N-Acetylglutamate kinase (NAGK) catalyzes the second step of arginine biosynthesis. In Pseudomonas aeruginosa, but not in Escherichia coli, this step is rate limiting and feedback and sigmoidally inhibited by arginine. Crystal structures revealed that arginine-insensitive E. coli NAGK (EcNAGK) is homodimeric, whereas arginine-inhibitable NAGKs, including P. aeruginosa NAGK (PaNAGK), are hexamers in which an extra N-terminal kinked helix (N-helix) interlinks three dimers. By introducing single amino acid replacements in PaNAGK, we prove the functionality of the structurally identified arginine site, as arginine site mutations selectively decreased the apparent affinity for arginine. N-helix mutations affecting R24 and E17 increased and decreased, respectively, the apparent affinity of PaNAGK for arginine, as predicted from enzyme structures that revealed the respective formation by these residues of bonds favoring inaccessible and accessible arginine site conformations. N-helix N-terminal deletions spanning ≥16 residues dissociated PaNAGK to active dimers, those of ≤20 residues decreased the apparent affinity for arginine, and complete N-helix deletion (26 residues) abolished arginine inhibition. Upon attachment of the PaNAGK N-terminal extension to the EcNAGK N terminus, EcNAGK remained dimeric and arginine insensitive. We concluded that the N-helix and its C-terminal portion after the kink are essential but not sufficient for hexamer formation and arginine inhibition, respectively; that the N-helix modulates NAGK affinity for arginine and mediates signal transmission between arginine sites, thus establishing sigmoidal arginine inhibition kinetics; that the mobile αH-β16 loop of the arginine site is the modulatory signal receiver; and that the hexameric architecture is not essential for arginine inhibition but is functionally essential for physiologically relevant arginine control of NAGK. PMID:18263723

  19. Basis of arginine sensitivity of microbial N-acetyl-L-glutamate kinases: mutagenesis and protein engineering study with the Pseudomonas aeruginosa and Escherichia coli enzymes.

    Science.gov (United States)

    Fernández-Murga, M Leonor; Rubio, Vicente

    2008-04-01

    N-acetylglutamate kinase (NAGK) catalyzes the second step of arginine biosynthesis. In Pseudomonas aeruginosa, but not in Escherichia coli, this step is rate limiting and feedback and sigmoidally inhibited by arginine. Crystal structures revealed that arginine-insensitive E. coli NAGK (EcNAGK) is homodimeric, whereas arginine-inhibitable NAGKs, including P. aeruginosa NAGK (PaNAGK), are hexamers in which an extra N-terminal kinked helix (N-helix) interlinks three dimers. By introducing single amino acid replacements in PaNAGK, we prove the functionality of the structurally identified arginine site, as arginine site mutations selectively decreased the apparent affinity for arginine. N-helix mutations affecting R24 and E17 increased and decreased, respectively, the apparent affinity of PaNAGK for arginine, as predicted from enzyme structures that revealed the respective formation by these residues of bonds favoring inaccessible and accessible arginine site conformations. N-helix N-terminal deletions spanning > or = 16 residues dissociated PaNAGK to active dimers, those of arginine, and complete N-helix deletion (26 residues) abolished arginine inhibition. Upon attachment of the PaNAGK N-terminal extension to the EcNAGK N terminus, EcNAGK remained dimeric and arginine insensitive. We concluded that the N-helix and its C-terminal portion after the kink are essential but not sufficient for hexamer formation and arginine inhibition, respectively; that the N-helix modulates NAGK affinity for arginine and mediates signal transmission between arginine sites, thus establishing sigmoidal arginine inhibition kinetics; that the mobile alphaH-beta16 loop of the arginine site is the modulatory signal receiver; and that the hexameric architecture is not essential for arginine inhibition but is functionally essential for physiologically relevant arginine control of NAGK. PMID:18263723

  20. Amino-acid selective experiments on uniformly 13C and 15N labeled proteins by MAS NMR: Filtering of lysines and arginines

    Science.gov (United States)

    Jehle, Stefan; Rehbein, Kristina; Diehl, Anne; van Rossum, Barth-Jan

    2006-12-01

    Amino-acid selective magic-angle spinning (MAS) NMR experiments can aid the assignment of ambiguous cross-peaks in crowded spectra of solid proteins. In particular for larger proteins, data analysis can be hindered by severe resonance overlap. In such cases, filtering techniques may provide a good alternative to site-specific spin-labeling to obtain unambiguous assignments that can serve as starting points in the assignment procedure. In this paper we present a simple pulse sequence that allows selective excitation of arginine and lysine residues. To achieve this, we make use of a combination of specific cross-polarization for selective excitation [M. Baldus, A.T. Petkova, J. Herzfeld, R.G. Griffin, Cross polarization in the tilted frame: assignment and spectral simplification in heteronuclear spin systems, Mol. Phys. 95 (1998) 1197-1207.] and spin diffusion for transfer along the amino-acid side-chain. The selectivity of the filter is demonstrated with the excitation of lysine and arginine side-chain resonances in a uniformly 13C and 15N labeled protein preparation of the α-spectrin SH3 domain. It is shown that the filter can be applied as a building block in a 13C- 13C lysine-only correlation experiment.

  1. Physiological implications of arginine metabolism in plants.

    Science.gov (United States)

    Winter, Gudrun; Todd, Christopher D; Trovato, Maurizio; Forlani, Giuseppe; Funck, Dietmar

    2015-01-01

    Nitrogen is a limiting resource for plant growth in most terrestrial habitats since large amounts of nitrogen are needed to synthesize nucleic acids and proteins. Among the 21 proteinogenic amino acids, arginine has the highest nitrogen to carbon ratio, which makes it especially suitable as a storage form of organic nitrogen. Synthesis in chloroplasts via ornithine is apparently the only operational pathway to provide arginine in plants, and the rate of arginine synthesis is tightly regulated by various feedback mechanisms in accordance with the overall nutritional status. While several steps of arginine biosynthesis still remain poorly characterized in plants, much wider attention has been paid to inter- and intracellular arginine transport as well as arginine-derived metabolites. A role of arginine as alternative source besides glutamate for proline biosynthesis is still discussed controversially and may be prevented by differential subcellular localization of enzymes. Apparently, arginine is a precursor for nitric oxide (NO), although the molecular mechanism of NO production from arginine remains unclear in higher plants. In contrast, conversion of arginine to polyamines is well documented, and in several plant species also ornithine can serve as a precursor for polyamines. Both NO and polyamines play crucial roles in regulating developmental processes as well as responses to biotic and abiotic stress. It is thus conceivable that arginine catabolism serves on the one hand to mobilize nitrogen storages, while on the other hand it may be used to fine-tune development and defense mechanisms against stress. This review summarizes the recent advances in our knowledge about arginine metabolism, with a special focus on the model plant Arabidopsis thaliana, and pinpoints still unresolved critical questions. PMID:26284079

  2. Basis of Arginine Sensitivity of Microbial N-Acetyl-l-Glutamate Kinases: Mutagenesis and Protein Engineering Study with the Pseudomonas aeruginosa and Escherichia coli Enzymes▿

    OpenAIRE

    Fernández-Murga, M. Leonor; Rubio, Vicente

    2008-01-01

    N-Acetylglutamate kinase (NAGK) catalyzes the second step of arginine biosynthesis. In Pseudomonas aeruginosa, but not in Escherichia coli, this step is rate limiting and feedback and sigmoidally inhibited by arginine. Crystal structures revealed that arginine-insensitive E. coli NAGK (EcNAGK) is homodimeric, whereas arginine-inhibitable NAGKs, including P. aeruginosa NAGK (PaNAGK), are hexamers in which an extra N-terminal kinked helix (N-helix) interlinks three dimers. By introducing single...

  3. The glove-like structure of the conserved membrane protein TatC provides insight into signal sequence recognition in twin-arginine translocation.

    Science.gov (United States)

    Ramasamy, Sureshkumar; Abrol, Ravinder; Suloway, Christian J M; Clemons, William M

    2013-05-01

    In bacteria, two signal-sequence-dependent secretion pathways translocate proteins across the cytoplasmic membrane. Although the mechanism of the ubiquitous general secretory pathway is becoming well understood, that of the twin-arginine translocation pathway, responsible for translocation of folded proteins across the bilayer, is more mysterious. TatC, the largest and most conserved of three integral membrane components, provides the initial binding site of the signal sequence prior to pore assembly. Here, we present two crystal structures of TatC from the thermophilic bacteria Aquifex aeolicus at 4.0 Å and 6.8 Å resolution. The membrane architecture of TatC includes a glove-shaped structure with a lipid-exposed pocket predicted by molecular dynamics to distort the membrane. Correlating the biochemical literature to these results suggests that the signal sequence binds in this pocket, leading to structural changes that facilitate higher order assemblies. PMID:23583035

  4. Time Resolved EPR Study on the Photoinduced Long-Range Charge-Separated State in Protein: Electron Tunneling Mediated by Arginine Residue in Human Serum Albumin.

    Science.gov (United States)

    Fuki, Masaaki; Murai, Hisao; Tachikawa, Takashi; Kobori, Yasuhiro

    2016-05-19

    To elucidate how local molecular conformations play a role on electronic couplings for the long-range photoinduced charge-separated (CS) states in protein systems, we have analyzed time-resolved electron paramagnetic resonance (TREPR) spectra by polarized laser irradiations of 9,10-anthraquinone-1-sulfonate (AQ1S(-)) bound to human serum albumin (HSA). Analyses of the magnetophotoselection effects on the EPR spectra and a docking simulation clarified the molecular geometry and the electronic coupling of the long-range CS states of AQ1S(•2-)-tryptophan214 radical cation (W214(•+)) separated by 1.2 nm. The ligand of AQ1S(-) has been demonstrated to be bound to the drug site I in HSA. Molecular conformations of the binding region were estimated by the docking simulations, indicating that an arginine218 (R218(+)) residue bound to AQ1S(•2-) mediates the long-range electron-transfer. The energetics of triad states of AQ1S(•2-)-R218(+)-W214(•+) and AQ1S(-)-R218(•)-W214(•+) have been computed on the basis of the density functional molecular orbital calculations, providing the clear evidence for the long-range electronic couplings of the CS states in terms of the superexchange tunneling model through the arginine residue. PMID:27116363

  5. Intra-domain Cross-talk Regulates Serine-arginine Protein Kinase 1-dependent Phosphorylation and Splicing Function of Transformer 2β1.

    Science.gov (United States)

    Jamros, Michael A; Aubol, Brandon E; Keshwani, Malik M; Zhang, Zhaiyi; Stamm, Stefan; Adams, Joseph A

    2015-07-10

    Transformer 2β1 (Tra2β1) is a splicing effector protein composed of a core RNA recognition motif flanked by two arginine-serine-rich (RS) domains, RS1 and RS2. Although Tra2β1-dependent splicing is regulated by phosphorylation, very little is known about how protein kinases phosphorylate these two RS domains. We now show that the serine-arginine protein kinase-1 (SRPK1) is a regulator of Tra2β1 and promotes exon inclusion in the survival motor neuron gene 2 (SMN2). To understand how SRPK1 phosphorylates this splicing factor, we performed mass spectrometric and kinetic experiments. We found that SRPK1 specifically phosphorylates 21 serines in RS1, a process facilitated by a docking groove in the kinase domain. Although SRPK1 readily phosphorylates RS2 in a splice variant lacking the N-terminal RS domain (Tra2β3), RS1 blocks phosphorylation of these serines in the full-length Tra2β1. Thus, RS2 serves two new functions. First, RS2 positively regulates binding of the central RNA recognition motif to an exonic splicing enhancer sequence, a phenomenon reversed by SRPK1 phosphorylation on RS1. Second, RS2 enhances ligand exchange in the SRPK1 active site allowing highly efficient Tra2β1 phosphorylation. These studies demonstrate that SRPK1 is a regulator of Tra2β1 splicing function and that the individual RS domains engage in considerable cross-talk, assuming novel functions with regard to RNA binding, splicing, and SRPK1 catalysis. PMID:26013829

  6. A proline/arginine-rich end leucine-rich repeat protein (PRELP) variant is uniquely expressed in chronic lymphocytic leukemia cells.

    Science.gov (United States)

    Mikaelsson, Eva; Österborg, Anders; Jeddi-Tehrani, Mahmood; Kokhaei, Parviz; Ostadkarampour, Mahyar; Hadavi, Reza; Gholamin, Mehran; Akhondi, Mehdi; Shokri, Fazel; Rabbani, Hodjattallah; Mellstedt, Håkan

    2013-01-01

    Proline/arginine-rich end leucine-rich repeat protein (PRELP) belongs to the small leucine-rich proteoglycan (SLRP) family, normally expressed in extracellular matrix of collagen-rich tissues. We have previously reported on another SLRP, fibromodulin (FMOD) in patients with chronic lymphocytic leukemia (CLL). PRELP is structurally similar to FMOD with adjacent localization on chromosome 1 (1q32.1). As cluster-upregulation of genes may occur in malignancies, the aim of our study was to analyze PRELP expression in CLL. PRELP was expressed (RT-PCR) in all CLL patients (30/30), as well as in some patients with mantle cell lymphoma (3/5), but not in healthy donor leukocytes (0/20) or tumor samples from other hematological malignancies (0/35). PRELP was also detected in CLL cell-lines (4/4) but not in cell-lines from other hematological tumors (0/9). PRELP protein was detected in all CLL samples but not in normal leukocytes. Deglycosylation experiments revealed a CLL-unique 38 kDa core protein, with an intact signal peptide. This 38 kDa protein was, in contrast to the normal 55 kDa size, not detected in serum which, in combination with the uncleaved signal peptide, suggests cellular retention. The unique expression of a 38 kDa PRELP in CLL cells may suggest involvement in the pathobiology of CLL and merits further studies. PMID:23826326

  7. A proline/arginine-rich end leucine-rich repeat protein (PRELP variant is uniquely expressed in chronic lymphocytic leukemia cells.

    Directory of Open Access Journals (Sweden)

    Eva Mikaelsson

    Full Text Available Proline/arginine-rich end leucine-rich repeat protein (PRELP belongs to the small leucine-rich proteoglycan (SLRP family, normally expressed in extracellular matrix of collagen-rich tissues. We have previously reported on another SLRP, fibromodulin (FMOD in patients with chronic lymphocytic leukemia (CLL. PRELP is structurally similar to FMOD with adjacent localization on chromosome 1 (1q32.1. As cluster-upregulation of genes may occur in malignancies, the aim of our study was to analyze PRELP expression in CLL. PRELP was expressed (RT-PCR in all CLL patients (30/30, as well as in some patients with mantle cell lymphoma (3/5, but not in healthy donor leukocytes (0/20 or tumor samples from other hematological malignancies (0/35. PRELP was also detected in CLL cell-lines (4/4 but not in cell-lines from other hematological tumors (0/9. PRELP protein was detected in all CLL samples but not in normal leukocytes. Deglycosylation experiments revealed a CLL-unique 38 kDa core protein, with an intact signal peptide. This 38 kDa protein was, in contrast to the normal 55 kDa size, not detected in serum which, in combination with the uncleaved signal peptide, suggests cellular retention. The unique expression of a 38 kDa PRELP in CLL cells may suggest involvement in the pathobiology of CLL and merits further studies.

  8. Homology modeling of major intrinsic proteins in rice, maize and Arabidopsis: comparative analysis of transmembrane helix association and aromatic/arginine selectivity filters

    Directory of Open Access Journals (Sweden)

    Sankararamakrishnan Ramasubbu

    2007-04-01

    Full Text Available Abstract Background The major intrinsic proteins (MIPs facilitate the transport of water and neutral solutes across the lipid bilayers. Plant MIPs are believed to be important in cell division and expansion and in water transport properties in response to environmental conditions. More than 30 MIP sequences have been identified in Arabidopsis thaliana, maize and rice. Plasma membrane intrinsic proteins (PIPs, tonoplast intrinsic proteins (TIPs, Nod26-like intrinsic protein (NIPs and small and basic intrinsic proteins (SIPs are subfamilies of plant MIPs. Despite sequence diversity, all the experimentally determined structures belonging to the MIP superfamily have the same "hour-glass" fold. Results We have structurally characterized 39 rice and 31 maize MIPs and compared them with that of Arabidopsis. Homology models of 105 MIPs from all three plant species were built. Structure-based sequence alignments were generated and the residues in the helix-helix interfaces were analyzed. Small residues (Gly/Ala/Ser/Thr are found to be highly conserved as a group in the helix-helix interface of MIP structures. Individual families sometimes prefer one or another of the residues from this group. The narrow aromatic/arginine (ar/R selectivity filter in MIPs has been shown to provide an important constriction for solute permeability. Ar/R regions were analyzed and compared between the three plant species. Seventeen TIP, NIP and SIP members from rice and maize have ar/R signatures that are not found in Arabidopsis. A subgroup of rice and maize NIPs has small residues in three of the four positions in the ar/R tetrad, resulting in a wider constriction. These MIP members could transport larger solute molecules. Conclusion Small residues are group-conserved in the helix-helix interface of MIP structures and they seem to be important for close helix-helix interactions. Such conservation might help to preserve the hour-glass fold in MIP structures. Analysis and

  9. Carnitine-acylcarnitine translocase deficiency with severe hypoglycemia and auriculo ventricular block. Translocase assay in permeabilized fibroblasts.

    OpenAIRE

    Pande, S V; Brivet, M; Slama, A.; Demaugre, F; Aufrant, C; Saudubray, J.M.

    1993-01-01

    Deficiency of the enzymes of mitochondrial fatty acid oxidation and related carnitine dependent steps have been shown to be one of the causes of the fasting-induced hypoketotic hypoglycemia. We describe here carnitine-acylcarnitine translocase deficiency in a neonate who died eight days after birth. The proband showed severe fasting-induced hypoketotic hypoglycemia, high plasma creatine kinase, heartbeat disorder, hypothermia, and hyperammonemia. The plasma-free carnitine on day three was onl...

  10. Genetics Home Reference: ornithine translocase deficiency

    Science.gov (United States)

    ... coma. Episodes of illness may coincide with the introduction of high-protein formulas or solid foods into ... Library of Medicine Lister Hill National Center for Biomedical Communications 8600 Rockville Pike, Bethesda, MD 20894, USA ...

  11. The effects of arginine glutamate, a promising excipient for protein formulation, on cell viability: Comparisons with NaCl.

    Science.gov (United States)

    Kheddo, Priscilla; Golovanov, Alexander P; Mellody, Kieran T; Uddin, Shahid; van der Walle, Christopher F; Dearman, Rebecca J

    2016-06-01

    The effects of an equimolar mixture of l-arginine and l-glutamate (Arg·Glu) on cell viability and cellular stress using in vitro cell culture systems are examined with reference to NaCl, in the context of monoclonal antibody formulation. Cells relevant to subcutaneous administration were selected: the human monocyte cell line THP-1, grown as a single cell suspension, and adherent human primary fibroblasts. For THP-1 cells, the mechanism of cell death caused by relatively high salt concentrations was investigated and effects on cell activation/stress assessed as a function of changes in membrane marker and cytokine (interleukin-8) expression. These studies demonstrated that Arg·Glu does not have any further detrimental effects on THP-1 viability in comparison to NaCl at equivalent osmolalities, and that both salts at higher concentrations cause cell death by apoptosis; there was no significant effect on measures of THP-1 cellular stress/activation. For adherent fibroblasts, both salts caused significant toxicity at ~400mOsm/kg, although Arg·Glu caused a more precipitous subsequent decline in viability than did NaCl. These data indicate that Arg·Glu is of equivalent toxicity to NaCl and that the mechanism of toxicity is such that cell death is unlikely to trigger inflammation upon subcutaneous injection in vivo. PMID:26873863

  12. Severe Hypomyelination and Developmental Defects Are Caused in Mice Lacking Protein Arginine Methyltransferase 1 (PRMT1) in the Central Nervous System.

    Science.gov (United States)

    Hashimoto, Misuzu; Murata, Kazuya; Ishida, Junji; Kanou, Akihiko; Kasuya, Yoshitoshi; Fukamizu, Akiyoshi

    2016-01-29

    Protein arginine methyltransferase 1 (PRMT1) is involved in cell proliferation, DNA damage response, and transcriptional regulation. Although PRMT1 is extensively expressed in the CNS at embryonic and perinatal stages, the physiological role of PRMT1 has been poorly understood. Here, to investigate the primary function of PRMT1 in the CNS, we generated CNS-specific PRMT1 knock-out mice by the Cre-loxP system. These mice exhibited postnatal growth retardation with tremors, and most of them died within 2 weeks after birth. Brain histological analyses revealed prominent cell reduction in the white matter tracts of the mutant mice. Furthermore, ultrastructural analysis demonstrated that myelin sheath was almost completely ablated in the CNS of these animals. In agreement with hypomyelination, we also observed that most major myelin proteins including myelin basic protein (MBP), 2',3'-cyclic-nucleotide 3'-phosphodiesterase (CNPase), and myelin-associated glycoprotein (MAG) were dramatically decreased, although neuronal and astrocytic markers were preserved in the brain of CNS-specific PRMT1 knock-out mice. These animals had a reduced number of OLIG2(+) oligodendrocyte lineage cells in the white matter. We found that expressions of transcription factors essential for oligodendrocyte specification and further maturation were significantly suppressed in the brain of the mutant mice. Our findings provide evidence that PRMT1 is required for CNS development, especially for oligodendrocyte maturation processes. PMID:26637354

  13. Chemical modification of arginine residues in the lactose repressor

    International Nuclear Information System (INIS)

    The lactose repressor protein was chemically modified with 2,3-butanedione and phenylglyoxal. Arginine reaction was quantitated by either amino aced analysis or incorporation of 14C-labeled phenylglyoxal. Inducer binding activity was unaffected by the modification of arginine residues, while both operator and nonspecific DNA binding activities were diminished, although to differing degrees. The correlation of the decrease in DNA binding activities with the modification of ∼ 1-2 equiv of arginine per monomer suggests increased reactivity of a functionally essential residue(s). For both reagents, operator DNA binding activity was protected by the presence of calf thymus DNA, and the extent of reaction with phenylglyoxal was simultaneously diminished. This protection presumably results from steric restriction of reagent access to an arginine(s) that is (are) essential for DNA binding interactions. These experiments suggest that there is (are) an essential reactive arginine(s) critical for repressor binding to DNA

  14. Protein arginine methyltransferase 1 may be involved in pregnane x receptor-activated overexpression of multidrug resistance 1 gene during acquired multidrug resistant

    Science.gov (United States)

    Li, Tingting; Kong, Ah-Ng Tony; Ma, Zhiqiang; Liu, Haiyan; Liu, Pinghua; Xiao, Yu; Jiang, Xuehua; Wang, Ling

    2016-01-01

    Purpose Pregnane x receptor (PXR) - activated overexpression of the multidrug resistance 1 (MDR1) gene is an important way for tumor cells to acquire drug resistance. However, the detailed mechanism still remains unclear. In the present study, we aimed to investigate whether protein arginine methyl transferase 1(PRMT1) is involved in PXR - activated overexpression of MDR1 during acquired multidrug resistant. Experimental Design Arginine methyltransferase inhibitor 1 (AMI-1) was used to pharmacologically block PRMT1 in resistant breast cancer cells (MCF7/adr). The mRNA and protein levels of MDR1 were detected by real-time PCR and western blotting analysis. Immunofluorescence microscopy and co-immunoprecipitation were used to investigate the physical interaction between PXR and PRMT1. Then, 136 candidate compounds were screened for PRMT1 inhibitors. Lastly, luciferase reporter gene and nude mice bearing resistant breast cancer xenografts were adopted to investigate the anti-tumor effect of PRMT1 inhibitors when combined with adriamycin. Results AMI-1 significantly suppressed the expression of MDR1 in MCF7/adr cells and increased cells sensitivity of MCF7/adr to adriamycin. Physical interaction between PRMT1 and PXR exists in MCF7/adr cells, which could be disrupted by AMI-1. Those results suggest that PRMT1 may be involved in PXR-activated overexpression of MDR1 in resistant breast cancer cells, and AMI-1 may suppress MDR1 by disrupting the interaction between PRMT1 and PXR. Then, five compounds including rutin, isoquercitrin, salvianolic acid A, naproxen, and felodipline were identified to be PRMT1 inhibitors. Finally, those PRMT1 inhibitors were observed to significantly decrease MDR1 promoter activity in vitro and enhance the antitumor effect of adriamycin in nude mice that bearing resistant breast cancer xenografts. Conclusions PRMT1 may be an important co-activator of PXR in activating MDR1 gene during acquired resistance, and PRMT1 inhibitor combined with

  15. A Tat ménage à trois--The role of Bacillus subtilis TatAc in twin-arginine protein translocation.

    Science.gov (United States)

    Goosens, Vivianne J; De-San-Eustaquio-Campillo, Alba; Carballido-López, Rut; van Dijl, Jan Maarten

    2015-10-01

    The twin-arginine translocation system (Tat) is a protein transport system that moves fully folded and cofactor-containing proteins across membranes of bacteria, archaea and thylakoids. The minimal Tat pathway is composed of two subunits, TatA and TatC. In some organisms TatA has been duplicated and evolved to form a third specialized subunit, TatB. The Bacillus subtilis genome encodes two TatC subunits (TatCd and TatCy) and three TatA subunits (TatAd, TatAy and TatAc). These subunits combine to form two parallel minimal pathways, TatAy-TatCy and TatAd-TatCd. The purpose and role of the third TatA component, TatAc, has remained ambiguous. In this study we examined the translocation of two natively expressed TatAy-TatCy-dependent substrates, EfeB and QcrA, in various Tat-deficient genetic backgrounds. More specifically, we examined the ability of different mutated TatAy subunits to complement for the absence of wild-type TatAy. We further detailed a graded growth phenotype associated with the functional translocation of EfeB. We found that in various instances where specific amino acid substitutions were made in TatAy, a definite TatAc-associated growth phenotype occurred in genetic backgrounds lacking TatAc. Altogether, our findings show that TatAy and TatAc interact and that this TatAy-TatAc interaction, although not essential, supports the translocation of the Tat substrate EfeB when TatAy function is compromised. This implies that the third TatA-like protein in B. subtilis could represent an intermediate evolutionary step in TatA-TatB specialization. PMID:26239117

  16. Structural analysis of substrate binding by the TatBC component of the twin-arginine protein transport system.

    Science.gov (United States)

    Tarry, Michael J; Schäfer, Eva; Chen, Shuyun; Buchanan, Grant; Greene, Nicholas P; Lea, Susan M; Palmer, Tracy; Saibil, Helen R; Berks, Ben C

    2009-08-11

    The Tat system transports folded proteins across the bacterial cytoplasmic membrane and the thylakoid membrane of plant chloroplasts. In Escherichia coli substrate proteins initially bind to the integral membrane TatBC complex which then recruits the protein TatA to effect translocation. Overproduction of TatBC and the substrate protein SufI in the absence of TatA led to the accumulation of TatBC-SufI complexes that could be purified using an affinity tag on the substrate. Three-dimensional structures of the TatBC-SufI complexes and unliganded TatBC were obtained by single-particle electron microscopy and random conical tilt reconstruction. Comparison of the structures shows that substrate molecules bind on the periphery of the TatBC complex and that substrate binding causes a significant reduction in diameter of the TatBC part of the complex. Although the TatBC complex contains multiple copies of the signal peptide-binding TatC protomer, purified TatBC-SufI complexes contain only 1 or 2 SufI molecules. Where 2 substrates are present in the TatBC-SufI complex, they are bound at adjacent sites. These observations imply that only certain TatC protomers within the complex interact with substrate or that there is a negative cooperativity of substrate binding. Similar TatBC-substrate complexes can be generated by an alternative in vitro reconstitution method and using a different substrate protein. PMID:19666509

  17. Arginine Biosynthesis in Thermotoga maritima: Characterization of the Arginine-Sensitive N-Acetyl-l-Glutamate Kinase

    OpenAIRE

    Fernández-Murga, M. Leonor; Gil-Ortiz, Fernando; Llácer, José L.; Rubio, Vicente

    2004-01-01

    To help clarify the control of arginine synthesis in Thermotoga maritima, the putative gene (argB) for N-acetyl-l-glutamate kinase (NAGK) from this microorganism was cloned and overexpressed, and the resulting protein was purified and shown to be a highly thermostable and specific NAGK that is potently and selectively inhibited by arginine. Therefore, NAGK is in T. maritima the feedback control point of arginine synthesis, a process that in this organism involves acetyl group recycling and ap...

  18. Induction of the arginine vasopressin-enhanced green fluorescent protein fusion transgene in the rat locus coeruleus

    Czech Academy of Sciences Publication Activity Database

    Todoroki, M.; Ueta, Y.; Fujihara, H.; Otsubo, H.; Shibata, M.; Hashimoto, H.; Kabayashi, M.; Sakamoto, H.; Kawata, M.; Dayanithi, Govindan; Murphy, D.; Hiro, H.; Takahashi, E.; Nagata, S.

    2010-01-01

    Roč. 13, č. 4 (2010), s. 281-292. ISSN 1025-3890 Institutional research plan: CEZ:AV0Z50390703 Keywords : colchicine * green fluorescent protein * hypothalamus Subject RIV: FH - Neurology Impact factor: 2.553, year: 2010

  19. L-arginine

    Science.gov (United States)

    ... your health provider.Medications for high blood pressure (ACE inhibitors)L-arginine seems to decrease blood pressure. Taking ... with certain medications for high blood pressure, called ACE inhibitors might cause your blood pressure to go too ...

  20. Expression of Major Capsid Protein of Cainine Parvovirus by Yeast (Pichia pastoris and Efficient Purification using Arginine in Affinity Chromatography

    Directory of Open Access Journals (Sweden)

    Ying He

    2016-06-01

    Full Text Available An immunochromatographic (IC assay was developed for rapid detection of canine parvovirus using the monoclonal antibodies (McAbs against canine parvovirus (CPV-2. To prepare the McAbs, gene encoding the VP2 protein of CPV-2a was expressed in a Pichia pastoris expression vector pPICZ-A. The recombinant VP2 was similar antigenically function to the native capsid protein as demonstrated by Western blotting using CPV- 2 polyclonal antiserum. McAbs against CPV-2 were produced by fusing myeloma cell line SP2/0 with spleen cells from Balb/C mice immunized with purified recombinant VP2 protein. By ELISA it was shown that the McAbs specifically recognized VP2 epitopes of CPV-2 but not those of other canine viruses such as Canine distemper virus (CDV or canine adenovirus (CAV. An IC assay developed with the McAbs was suitable for rapid detection of canine parvovirus. Fecal samples (120 from dogs suspected of CPV-2 infection were analyzed by both haemaglutination (HA assay and the IC assay, and 52 and 53 samples were found positive for CPV-2, respectively. Comparison between the two different assays revealed that IC assay is as sensitive as HA; the sensitivity and specificity for the IC assay is 98.6% and 98.1%, respectively.

  1. The TatA component of the twin-arginine protein transport system forms channel complexes of variable diameter

    OpenAIRE

    Gohlke, Ulrich; Pullan, Lee; McDevitt, Christopher A.; Porcelli, Ida; de Leeuw, Erik; Palmer, Tracy; Saibil, Helen R.; Berks, Ben C.

    2005-01-01

    The Tat system mediates Sec-independent transport of folded precursor proteins across the bacterial plasma membrane or the chloroplast thylakoid membrane. Tat transport involves distinct high-molecular-weight TatA and TatBC complexes. Here we report the 3D architecture of the TatA complex from Escherichia coli obtained by single-particle electron microscopy and random conical tilt reconstruction. TatA forms ring-shaped structures of variable diameter in which the internal channels are large e...

  2. Arginine: Its pKa value revisited

    Science.gov (United States)

    Fitch, Carolyn A; Platzer, Gerald; Okon, Mark; Garcia-Moreno E, Bertrand; McIntosh, Lawrence P

    2015-01-01

    Using complementary approaches of potentiometry and NMR spectroscopy, we have determined that the equilibrium acid dissociation constant (pKa value) of the arginine guanidinium group is 13.8 ± 0.1. This is substantially higher than that of ∼12 often used in structure-based electrostatics calculations and cited in biochemistry textbooks. The revised intrinsic pKa value helps explains why arginine side chains in proteins are always predominantly charged, even at pH values as great as 10. The high pKa value also reinforces the observation that arginine side chains are invariably protonated under physiological conditions of near neutral pH. This occurs even when the guanidinium moiety is buried in a hydrophobic micro-environment, such as that inside a protein or a lipid membrane, thought to be incompatible with the presence of a charged group. PMID:25808204

  3. Arginine, citrulline and nitric oxide metabolism in sepsis

    Science.gov (United States)

    Arginine has vasodilatory effects, via its conversion by nitric oxide (NO) synthase into NO, and immunomodulatory actions that play important roles in sepsis. Protein breakdown affects arginine availability, and the release of asymmetric dimethylarginine, an inhibitor of NO synthase, may therefore a...

  4. On the mechanism of arginine requirement for adenovirus synthesis

    International Nuclear Information System (INIS)

    The effects of arginine deprivation on the synthesis and processing of viral proteins and the assembly of incomplete and complete virions were studied during infection with human adenovirus type 2. Arginine deprivation greatly reduced the synthesis of all viral proteins, particularly the precursor to core protein VII. The inhibition was completely reversible by the addition of arginine to the medium. Arginine deprivation between 7 and 20 hours post-infection inhibited the processing of PVII to VII, suggesting that PVII is not cleaved autocatalytically. The assembly of incomplete virions was sensitive to arginine deprivation only prior to 20 hours, while the assembly of complete virions was dependent on the continuous presence of arginine. This observation supports the hypothesis that incomplete virions are precursors of complete virions. The experiments on the PVII-specific endoprotease activity showed that arginine deprivation caused only slight reduction in the in vitro activity, although no activity was observed in vivo. The present results lead to the hypothesis that arginine deficiency inhibits the synthesis of a functional protein essential for virion maturation, other than the synthesis of processing of PVII. (author)

  5. Arginine biosynthesis in Thermotoga maritima: characterization of the arginine-sensitive N-acetyl-L-glutamate kinase.

    Science.gov (United States)

    Fernández-Murga, M Leonor; Gil-Ortiz, Fernando; Llácer, José L; Rubio, Vicente

    2004-09-01

    To help clarify the control of arginine synthesis in Thermotoga maritima, the putative gene (argB) for N-acetyl-L-glutamate kinase (NAGK) from this microorganism was cloned and overexpressed, and the resulting protein was purified and shown to be a highly thermostable and specific NAGK that is potently and selectively inhibited by arginine. Therefore, NAGK is in T. maritima the feedback control point of arginine synthesis, a process that in this organism involves acetyl group recycling and appears not to involve classical acetylglutamate synthase. The inhibition of NAGK by arginine was found to be pH independent and to depend sigmoidally on the concentration of arginine, with a Hill coefficient (N) of approximately 4, and the 50% inhibitory arginine concentration (I0.5) was shown to increase with temperature, approaching above 65 degrees C the I0.50 observed at 37 degrees C with the mesophilic NAGK of Pseudomonas aeruginosa (the best-studied arginine-inhibitable NAGK). At 75 degrees C, the inhibition by arginine of T. maritima NAGK was due to a large increase in the Km for acetylglutamate triggered by the inhibitor, but at 37 degrees C arginine also substantially decreased the Vmax of the enzyme. The NAGKs of T. maritima and P. aeruginosa behaved in gel filtration as hexamers, justifying the sigmoidicity and high Hill coefficient of arginine inhibition, and arginine or the substrates failed to disaggregate these enzymes. In contrast, Escherichia coli NAGK is not inhibited by arginine and is dimeric, and thus the hexameric architecture may be an important determinant of arginine sensitivity. Potential thermostability determinants of T. maritima NAGK are also discussed. PMID:15342584

  6. The effects of in ovo feeding arginine, beta-hydroxy-beta-methyl-butyrate, and protein on jejunal digestive and absorptive activity in embryonic and neonatal turkey poults.

    Science.gov (United States)

    Foye, O T; Ferket, P R; Uni, Z

    2007-11-01

    In ovo feeding, injecting nutrients into the amnion of the avian embryo, may enhance jejunal nutrient uptake, activity of the intestinal enzymes, and posthatch growth. This hypothesis was tested in the following in ovo feeding (IOF) experiments. In experiment 1, 400 eggs were evenly distributed among 4 nutritional treatments at 23 d of embryonic development (23E) and administered 1 of 4 treatments as a 2 x 2 factorial arrangement of arginine (ARG 0, 0.7%) and beta-hydroxy-beta-methyl-butyrate (HMB 0, 0.1%). Tissues were assayed for maltase, sucrase, and leucine aminopeptidase (LAP) at 25E, hatch, and 3, 7, and 14 d. In experiment 2, all IOF procedures were repeated and treatments were administered at 21E: injected or noninjected control, 21% egg white protein (EWP), 21% EWP + 0.1% HMB. In experiment 3, two hundred eggs were evenly distributed among the following treatments at 23E: noninjected control or 0.7% ARG + 0.1% HMB + 21% EWP. Jejunal samples were assayed for glucose or alanine uptake at 23E, 25E, and hatch (experiment 2), and hatch and 7 d (experiment 3), respectively. All poults were fed a turkey starter diet ad libitum immediately upon hatching. There was a highly significant HMB x ARG interaction on jejunal sucrase, maltase, and LAP activities at 25E and 14 d. Poults in ovo (IO) fed HMB + ARG had approximately a 2- to 3-fold increase in jejunal sucrase, maltase, and LAP activities at 25E, and a 3-fold increase at 14 d, over other treatments. Poults IO fed EWP + HMB (experiment 2) had enhanced glucose uptake at 25E, whereas poults IO fed ARG + HMB + EWP (experiment 3) had enhanced alanine uptake at hatch and 7 d. These studies demonstrate that IOF ARG, HMB, and EWP may enhance jejunal nutrient uptake and digestion in turkeys. PMID:17954584

  7. The critical role of protein arginine methyltransferase prmt8 in zebrafish embryonic and neural development is non-redundant with its paralogue prmt1.

    Directory of Open Access Journals (Sweden)

    Yu-ling Lin

    Full Text Available Protein arginine methyltransferase (PRMT 1 is the most conserved and widely distributed PRMT in eukaryotes. PRMT8 is a vertebrate-restricted paralogue of PRMT1 with an extra N-terminal sequence and brain-specific expression. We use zebrafish (Danio rerio as a vertebrate model to study PRMT8 function and putative redundancy with PRMT1. The transcripts of zebrafish prmt8 were specifically expressed in adult zebrafish brain and ubiquitously expressed from zygotic to early segmentation stage before the neuronal development. Whole-mount in situ hybridization revealed ubiquitous prmt8 expression pattern during early embryonic stages, similar to that of prmt1. Knockdown of prmt8 with antisense morpholino oligonucleotide phenocopied prmt1-knockdown, with convergence/extension defects at gastrulation. Other abnormalities observed later include short body axis, curled tails, small and malformed brain and eyes. Catalytically inactive prmt8 failed to complement the morphants, indicating the importance of methyltransferase activity. Full-length prmt8 but not prmt1 cRNA can rescue the phenotypic changes. Nevertheless, cRNA encoding Prmt1 fused with the N-terminus of Prmt8 can rescue the prmt8 morphants. In contrast, N-terminus- deleted but not full-length prmt8 cRNA can rescue the prmt1 morphants as efficiently as prmt1 cRNA. Abnormal brain morphologies illustrated with brain markers and loss of fluorescent neurons in a transgenic fish upon prmt8 knockdown confirm the critical roles of prmt8 in neural development. In summery, our study is the first report showing the expression and function of prmt8 in early zebrafish embryogenesis. Our results indicate that prmt8 may play important roles non-overlapping with prmt1 in embryonic and neural development depending on its specific N-terminus.

  8. An evolutionarily conserved arginine is essential for Tre1 G protein-coupled receptor function during germ cell migration in Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Angela R Kamps

    Full Text Available BACKGROUND: G protein-coupled receptors (GPCRs play central roles in mediating cellular responses to environmental signals leading to changes in cell physiology and behaviors, including cell migration. Numerous clinical pathologies including metastasis, an invasive form of cell migration, have been linked to abnormal GPCR signaling. While the structures of some GPCRs have been defined, the in vivo roles of conserved amino acid residues and their relationships to receptor function are not fully understood. Trapped in endoderm 1 (Tre1 is an orphan receptor of the rhodopsin class that is necessary for primordial germ cell migration in Drosophila melanogaster embryos. In this study, we employ molecular genetic approaches to identify residues in Tre1 that are critical to its functions in germ cell migration. METHODOLOGY/PRINCIPAL FINDINGS: First, we show that the previously reported scattershot mutation is an allele of tre1. The scattershot allele results in an in-frame deletion of 8 amino acids at the junction of the third transmembrane domain and the second intracellular loop of Tre1 that dramatically impairs the function of this GPCR in germ cell migration. To further refine the molecular basis for this phenotype, we assayed the effects of single amino acid substitutions in transgenic animals and determined that the arginine within the evolutionarily conserved E/N/DRY motif is critical for receptor function in mediating germ cell migration within an intact developing embryo. CONCLUSIONS/SIGNIFICANCE: These structure-function studies of GPCR signaling in native contexts will inform future studies into the basic biology of this large and clinically important family of receptors.

  9. Inhibition of peptidyl-arginine deiminases reverses protein-hypercitrullination and disease in mouse models of multiple sclerosis

    Directory of Open Access Journals (Sweden)

    Mario A. Moscarello

    2013-03-01

    Multiple sclerosis (MS is the most common CNS-demyelinating disease of humans, showing clinical and pathological heterogeneity and a general resistance to therapy. We first discovered that abnormal myelin hypercitrullination, even in normal-appearing white matter, by peptidylarginine deiminases (PADs correlates strongly with disease severity and might have an important role in MS progression. Hypercitrullination is known to promote focal demyelination through reduced myelin compaction. Here we report that 2-chloroacetamidine (2CA, a small-molecule, PAD active-site inhibitor, dramatically attenuates disease at any stage in independent neurodegenerative as well as autoimmune MS mouse models. 2CA reduced PAD activity and protein citrullination to pre-disease status. In the autoimmune models, disease induction uniformly induced spontaneous hypercitrullination with citrulline+ epitopes targeted frequently. 2CA rapidly suppressed T cell autoreactivity, clearing brain and spinal cord infiltrates, through selective removal of newly activated T cells. 2CA essentially prevented disease when administered before disease onset or before autoimmune induction, making hypercitrullination, and specifically PAD enzymes, a therapeutic target in MS models and thus possibly in MS.

  10. The Selective Estrogen Receptor Modulator Raloxifene Regulates Arginine-Vasopressin Gene Expression in Human Female Neuroblastoma Cells Through G Protein-Coupled Estrogen Receptor and ERK Signaling.

    Science.gov (United States)

    Grassi, Daniela; Ghorbanpoor, Samar; Acaz-Fonseca, Estefania; Ruiz-Palmero, Isabel; Garcia-Segura, Luis M

    2015-10-01

    The selective estrogen receptor modulator raloxifene reduces blood pressure in hypertensive postmenopausal women. In the present study we have explored whether raloxifene regulates gene expression of arginine vasopressin (AVP), which is involved in the pathogenesis of hypertension. The effect of raloxifene was assessed in human female SH-SY5Y neuroblastoma cells, which have been recently identified as a suitable cellular model to study the estrogenic regulation of AVP. Raloxifene, within a concentration ranging from 10(-10) M to 10(-6) M, decreased the mRNA levels of AVP in SH-SY5Y cells with maximal effect at 10(-7) M. This effect of raloxifene was imitated by an agonist (±)-1-[(3aR*,4S*,9bS*)-4-(6-bromo-1,3-benzodioxol-5-yl)-3a,4,5,9b-tetrahydro-3H-cyclopenta[c]quinolin-8-yl]-ethanone of G protein-coupled estrogen receptor-1 (GPER) and blocked by an antagonist (3aS*,4R*,9bR*)-4-(6-bromo-1,3-benzodioxol-5-yl)-3a,4,5,9b-3H-cyclopenta[c]quinoline of GPER and by GPER silencing. Raloxifene induced a time-dependent increase in the level of phosphorylated ERK1 and ERK2, by a mechanism blocked by the GPER antagonist. The treatment of SH-SY5Y cells with either a MAPK/ERK kinase 1/2-specific inhibitor (1,4-diamino-2, 3-dicyano-1,4-bis(2-aminophenylthio)butadine) or a protein kinase C inhibitor (sotrastaurin) blocked the effects of raloxifene on the phosphorylation of ERK1/2 and the regulation of AVP mRNA levels. These results reveal a mechanism mediating the regulation of AVP expression by raloxifene, involving the activation of GPER, which in turn activates protein kinase C, MAPK/ERK kinase, and ERK. The regulation of AVP by raloxifene and GPER may have implications for the treatment of blood hypertension(.). PMID:26200092

  11. Lysine and arginine requirements of Salminus brasiliensis

    Directory of Open Access Journals (Sweden)

    Jony Koji Dairiki

    2013-08-01

    Full Text Available The objective of this work was to determine the dietary lysine (DL and dietary arginine (DA requirements of dourado (Salminus brasiliensis, through dose-response trials using the amino acid profiles of whole carcasses as a reference. Two experiments were carried out in a completely randomized design (n=4. In the first experiment, groups of 12 feed-conditioned dourado juveniles (11.4±0.2 g were stocked in 60 L cages placed in 300 L plastic indoor tanks in a closed circulation system. Fish were fed for 60 days on diets containing 1.0, 1.5, 2.0, 2.5, 3.0, or 3.5 % dietary lysine. In the second experiment, dourado juveniles (27.0±0.8 g were fed for 60 days on semipurified diets containing arginine at 1.0, 1.5, 2.0, 2.5 or 3.0%, in similar conditions to those of the first experiment. Optimal DL requirements, as determined by broken-line analysis method for final weight, weight gain and specific growth rate, were 2.15% DL or 5% lysine in dietary protein, and 1.48% DA or 3.43% arginine in dietary protein. The best feed conversion ratio is attained with 2.5% DL or 5.8% lysine in dietary protein and 1.4% DA or 3.25% arginine in dietary protein.

  12. Arginine: Its pKa value revisited

    OpenAIRE

    Fitch, Carolyn A.; Platzer, Gerald; Okon, Mark; Garcia-Moreno E, Bertrand; McIntosh, Lawrence P.

    2015-01-01

    Using complementary approaches of potentiometry and NMR spectroscopy, we have determined that the equilibrium acid dissociation constant (pKa value) of the arginine guanidinium group is 13.8 ± 0.1. This is substantially higher than that of ∼12 often used in structure-based electrostatics calculations and cited in biochemistry textbooks. The revised intrinsic pKa value helps explains why arginine side chains in proteins are always predominantly charged, even at pH values as great as 10. The hi...

  13. Octa-arginine mediated delivery of wild-type Lnk protein inhibits TPO-induced M-MOK megakaryoblastic leukemic cell growth by promoting apoptosis.

    Directory of Open Access Journals (Sweden)

    Chung Yeng Looi

    Full Text Available BACKGROUND: Lnk plays a non-redundant role by negatively regulating cytokine signaling of TPO, SCF or EPO. Retroviral expression of Lnk has been shown to suppress hematopoietic leukemic cell proliferation indicating its therapeutic value in cancer therapy. However, retroviral gene delivery carries risks of insertional mutagenesis. To circumvent this undesired consequence, we fused a cell permeable peptide octa-arginine to Lnk and evaluated the efficacy of inhibition of leukemic cell proliferation in vitro. METHODOLOGY/PRINCIPAL FINDINGS: In this study, proliferation assays, flow cytometry, Western Blot analyses were performed on wild-type (WT, mutant Lnk R8 or BSA treated M-MOK cells. We found that delivered WT, but not mutant Lnk R8 blocked TPO-induced M-MOK megakaryoblastic leukemic cell proliferation. In contrast, WT Lnk R8 showed no growth inhibitive effect on non-hematopoietic HELA or COS-7 cell. Moreover, we demonstrated that TPO-induced M-MOK cell growth inhibition by WT Lnk R8 was dose-dependent. Penetrated WT Lnk R8 induced cell cycle arrest and apoptosis. Immunoprecipitation and Western blots data indicated WT Lnk R8 interacted with endogeneous Jak2 and downregulated Jak-Stat and MAPK phosphorylation level in M-MOK cells after TPO stimulation. Treatment with specific inhibitors (TG101348 and PD98059 indicated Jak-Stat and MAPK pathways were crucial for TPO-induced proliferation of M-MOK cells. Further analyses using TF-1 and HEL leukemic cell-lines showed that WT Lnk R8 inhibited Jak2-dependent cell proliferation. Using cord blood-derived CD34+ stem cells, we found that delivered WT Lnk R8 blocked TPO-induced megakaryopoiesis in vitro. CONCLUSIONS/SIGNIFICANCE: Intracellular delivery of WT Lnk R8 fusion protein efficiently inhibited TPO-induced M-MOK leukemic cell growth by promoting apoptosis. WT Lnk R8 protein delivery may provide a safer and more practical approach to inhibit leukemic cell growth worthy of further development.

  14. Protein: FBB5 [TP Atlas

    Lifescience Database Archive (English)

    Full Text Available FBB5 RNA modification secDF Protein translocase subunit SecDF 300852 Thermus thermophilus (strai ... n HB8 / ATCC 27634 / DSM ... 579) 3168575 Q5SKE6 3AQP, 2RRN, 3AQO 21562494 #201 ...

  15. Protein: FBB5 [TP Atlas

    Lifescience Database Archive (English)

    Full Text Available FBB5 RNA modification secY Protein translocase subunit SecY 300852 Thermus thermophilus (strain ... HB8 / ATCC 27634 / DSM ... 579) 3169829 Q5SHQ8 2ZJS, 2ZQP 18923527 #20081016 ...

  16. Arginine metabolism in wounds

    International Nuclear Information System (INIS)

    Arginine metabolism in wounds was investigated in the rat in 1) lambda-carrageenan-wounded skeletal muscle, 2) Schilling chambers, and 3) subcutaneous polyvinyl alcohol sponges. All showed decreased arginine and elevated ornithine contents and high arginase activity. Arginase could be brought to the wound by macrophages, which were found to contain arginase activity. However, arginase was expressed by macrophages only after cell lysis and no arginase was released by viable macrophages in vitro. Thus the extracellular arginase of wounds may derive from dead macrophages within the injured tissue. Wound and peritoneal macrophages exhibited arginase deiminase activity as demonstrated by the conversion of [guanido-14C]arginine to radiolabeled citrulline during culture, the inhibition of this reaction by formamidinium acetate, and the lack of prokaryotic contamination of the cultures. These findings and the known metabolic fates of the products of arginase and arginine deiminase in the cellular populations of the wound suggest the possibility of cooperativity among cells for the production of substrates for collagen synthesis

  17. L-arginine

    Science.gov (United States)

    ... pain (angina pectoris). Leg pain associated with poor blood flow (peripheral arterial disease). Research suggests that taking L-arginine by mouth ... by IV) for up to 8 weeks increases blood flow in people with peripheral arterial disease. However, long-term use (up to 6 months) ...

  18. Solubilization of aromatic and hydrophobic moieties by arginine in aqueous solutions

    Science.gov (United States)

    Li, Jianguo; Garg, Manju; Shah, Dhawal; Rajagopalan, Raj

    2010-08-01

    Experiments hold intriguing, circumstantial clues to the mechanisms behind arginine-mediated solubilization of small organic drugs and suppression of protein aggregation driven by hydrophobic or aromatic associations, but how exactly arginine's molecular structure and interactions contribute to its function remains unclear since attention has focused so far on the thermodynamics of the preferential exclusion or binding of arginine. Here, we examine, through molecular dynamics simulations, how arginine solubilizes nanoscale particles with hydrophobic surfaces or aromatic-ring-type surface interactions. We show that preferential, hydrophobic, and dispersion interactions of arginine's guanidinium group with the particles lead to a surfactant-like behavior of arginine around the particles and to a solvation layer with a protective polar mask creating a hydrophilic shell. Additionally, arginine-arginine association around the solvation layer further prevents aggregative contacts. The results shed some light on the mechanistic basis of arginine's function as a suppressant of protein aggregation, although the complex energy landscapes and kinetic pathways of aggregation are protein-dependent and pose formidable challenges to developing comprehensive mechanistic pictures. Our results suggest arginine's mode of interaction with hydrophobic patches and aromatic residues could reduce aggregation-prone intermediate states of proteins and shield protein-protein aggregative contacts. The approach used here offers a systematic way of exploring implications of other amino acid/excipient interactions by studying interactions of the excipient with particles grafted with amino acids.

  19. Proteome-wide analysis of arginine monomethylation reveals widespread occurrence in human cells.

    Science.gov (United States)

    Larsen, Sara C; Sylvestersen, Kathrine B; Mund, Andreas; Lyon, David; Mullari, Meeli; Madsen, Maria V; Daniel, Jeremy A; Jensen, Lars J; Nielsen, Michael L

    2016-01-01

    The posttranslational modification of proteins by arginine methylation is functionally important, yet the breadth of this modification is not well characterized. Using high-resolution mass spectrometry, we identified 8030 arginine methylation sites within 3300 human proteins in human embryonic kidney 293 cells, indicating that the occurrence of this modification is comparable to phosphorylation and ubiquitylation. A site-level conservation analysis revealed that arginine methylation sites are less evolutionarily conserved compared to arginines that were not identified as modified by methylation. Through quantitative proteomics and RNA interference to examine arginine methylation stoichiometry, we unexpectedly found that the protein arginine methyltransferase (PRMT) family of arginine methyltransferases catalyzed methylation independently of arginine sequence context. In contrast to the frequency of somatic mutations at arginine methylation sites throughout the proteome, we observed that somatic mutations were common at arginine methylation sites in proteins involved in mRNA splicing. Furthermore, in HeLa and U2OS cells, we found that distinct arginine methyltransferases differentially regulated the functions of the pre-mRNA splicing factor SRSF2 (serine/arginine-rich splicing factor 2) and the RNA transport ribonucleoprotein HNRNPUL1 (heterogeneous nuclear ribonucleoprotein U-like 1). Knocking down PRMT5 impaired the RNA binding function of SRSF2, whereas knocking down PRMT4 [also known as coactivator-associated arginine methyltransferase 1 (CARM1)] or PRMT1 increased the RNA binding function of HNRNPUL1. High-content single-cell imaging additionally revealed that knocking down CARM1 promoted the nuclear accumulation of SRSF2, independent of cell cycle phase. Collectively, the presented human arginine methylome provides a missing piece in the global and integrative view of cellular physiology and protein regulation. PMID:27577262

  20. Cryo-electron microscopy structure of a yeast mitochondrial preprotein translocase.

    Science.gov (United States)

    Model, Kirstin; Meisinger, Chris; Kühlbrandt, Werner

    2008-11-28

    The translocase of the outer mitochondrial membrane (TOM) complex is the main entry gate for proteins imported into mitochondria. We determined the structure of the native, unstained approximately 550-kDa core-Tom20 complex from Saccharomycescerevisiae by cryo-electron microscopy at 18-A resolution. The complex is triangular, measuring 145 A on edge, and has near-3-fold symmetry. Its bulk is made up of three globular approximately 50-A domains. Three elliptical pores on the c-face merge into one central approximately 70-A cavity with a cage-like assembly on the opposite t-face. Nitrilotriacetic acid-gold labeling indicates that three Tom22 subunits in the TOM complex are located at the perimeter of the complex near the interface of the globular domains. We assign Tom22, which controls complex assembly, to three peripheral protrusions on the c-face, while the Tom20 subunit is tentatively assigned to the central protrusion on this surface. Based on our three-dimensional map, we propose a model of transient interactions and functional dynamics of the TOM assembly. PMID:18706915

  1. Active displacement of RecA filaments by UvrD translocase activity.

    Science.gov (United States)

    Petrova, Vessela; Chen, Stefanie H; Molzberger, Eileen T; Tomko, Eric; Chitteni-Pattu, Sindhu; Jia, Haifeng; Ordabayev, Yerdos; Lohman, Timothy M; Cox, Michael M

    2015-04-30

    The UvrD helicase has been implicated in the disassembly of RecA nucleoprotein filaments in vivo and in vitro. We demonstrate that UvrD utilizes an active mechanism to remove RecA from the DNA. Efficient RecA removal depends on the availability of DNA binding sites for UvrD and/or the accessibility of the RecA filament ends. The removal of RecA from DNA also requires ATP hydrolysis by the UvrD helicase but not by RecA protein. The RecA-removal activity of UvrD is slowed by RecA variants with enhanced DNA-binding properties. The ATPase rate of UvrD during RecA removal is much slower than the ATPase activity of UvrD when it is functioning either as a translocase or a helicase on DNA in the absence of RecA. Thus, in this context UvrD may operate in a specialized disassembly mode. PMID:25824953

  2. Inhibition of lytic infection of pseudorabies virus by arginine depletion

    International Nuclear Information System (INIS)

    Pseudorabies virus (PRV) is a member of Alphahepesviruses; it is an enveloped virus with a double-stranded DNA genome. Polyamines (such as spermine and spermidine) are ubiquitous in animal cells and participate in cellular proliferation and differentiation. Previous results of our laboratory showed that the PRV can accomplish lytic infection either in the presence of exogenous spermine (or spermidine) or depletion of cellular polyamines. The amino acid arginine is a precursor of polyamine biosynthesis. In this work, we investigated the role of arginine in PRV infection. It was found that the plaque formation of PRV was inhibited by arginase (enzyme catalyzing the conversion of arginine into ornithine and urea) treatment whereas this inhibition can be reversed by exogenous arginine, suggesting that arginine is essential for PRV proliferation. Western blotting was conducted to study the effect of arginine depletion on the levels of structural proteins of PRV in virus-infected cells. Four PRV structural proteins (gB, gE, UL47, and UL48) were chosen for examination, and results revealed that the levels of viral proteins were obviously reduced in long time arginase treatment. However, the overall protein synthesis machinery was apparently not influenced by arginase treatment either in mock or PRV-infected cells. Analyzing with native gel, we found that arginase treatment affected the mobility of PRV structural proteins, suggesting the conformational change of viral proteins by arginine depletion. Heat shock proteins, acting as molecular chaperons, participate in protein folding and translocation. Our results demonstrated that long time arginase treatment could reduce the expression of cellular heat shock proteins 70 (hsc70 and hsp70), and transcriptional suppression of heat shock protein 70 gene promoter was one of the mechanisms involved in this reduced expression

  3. Two adenine nucleotide translocase paralogues involved in cell proliferation and spermatogenesis in the silkworm Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Ryohei Sugahara

    Full Text Available Mitochondrial adenine nucleotide translocase (ANT specifically acts in ADP/ATP exchange through the mitochondrial inner membrane. This transporter protein thereby plays a significant role in energy metabolism in eukaryotic cells. Most mammals have four paralogous ANT genes (ANT1-4 and utilize these paralogues in different types of cells. The fourth paralogue of ANT (ANT4 is present only in mammals and reptiles and is exclusively expressed in testicular germ cells where it is required for meiotic progression in the spermatocytes. Here, we report that silkworms harbor two ANT paralogues, the homeostatic paralogue (BmANTI1 and the testis-specific paralogue (BmANTI2. The BmANTI2 protein has an N-terminal extension in which the positions of lysine residues in the amino acid sequence are distributed as in human ANT4. An expression analysis showed that BmANTI2 transcripts were restricted to the testis, suggesting the protein has a role in the progression of spermatogenesis. By contrast, BmANTI1 was expressed in all tissues tested, suggesting it has an important role in homeostasis. We also observed that cultured silkworm cells required BmANTI1 for proliferation. The ANTI1 protein of the lepidopteran Plutella xylostella (PxANTI1, but not those of other insect species (or PxANTI2, restored cell proliferation in BmANTI1-knockdown cells suggesting that ANTI1 has similar energy metabolism functions across the Lepidoptera. Our results suggest that BmANTI2 is evolutionarily divergent from BmANTI1 and has developed a specific role in spermatogenesis similar to that of mammalian ANT4.

  4. Arginine methylation regulates the p53 response

    DEFF Research Database (Denmark)

    Jansson, Martin; Durant, Stephen T; Cho, Er-Chieh;

    2008-01-01

    Activation of the p53 tumour suppressor protein in response to DNA damage leads to apoptosis or cell-cycle arrest. Enzymatic modifications are widely believed to affect and regulate p53 activity. We describe here a level of post-translational control that has an important functional consequence on...... of p53. Furthermore, PRMT5 depletion triggers p53-dependent apoptosis. Thus, methylation on arginine residues is an underlying mechanism of control during the p53 response....

  5. Rotations of the 2B Sub-domain of E. coli UvrD Helicase/Translocase Coupled to Nucleotide and DNA Binding

    OpenAIRE

    Jia, Haifeng; Korolev, Sergey; Niedziela-Majka, Anita; Maluf, Nasib K.; Gauss, George H.; Myong, Sua; Ha, Taekjip; Waksman, Gabriel; Lohman, Timothy M.

    2011-01-01

    E. coli UvrD is a superfamily 1 (SF1) DNA helicase and single stranded (ss) DNA translocase that functions in DNA repair, plasmid replication and as an anti-recombinase by removing RecA protein from ssDNA. UvrD couples ATP binding and hydrolysis to unwind double-stranded DNA (dsDNA) and translocate along ssDNA with 3′ to 5′ directionality. Although a UvrD monomer is able to translocate along ssDNA rapidly and processively, DNA helicase activity in vitro requires a minimum of a UvrD dimer. Pre...

  6. Níveis de proteína e de arginina digestível na ração pré-inicial de frangos de corte Protein and digestible arginine levels in pre-starter broiler rations

    Directory of Open Access Journals (Sweden)

    Mônica Schaitl Thon

    2010-05-01

    Full Text Available O experimento foi conduzido para avaliar níveis de proteína bruta e arginina digestível na ração pré-inicial de frangos de corte e seus efeitos no desempenho das aves dos 7 aos 21 dias de idade. Foram utilizados 600 pintos da linhagem Cobb 500, distribuídos em delineamento de blocos casualizados em esquema fatorial 4 × 2, composto de quatro níveis de arginina digestível (1,363; 1,463; 1,563 e 1,663% e dois níveis de proteína bruta (20 e 22%, totalizando oito tratamentos, cada um com cinco repetições de 15 aves. Avaliaram-se o ganho de peso, o consumo de ração, o índice de conversão alimentar, a biometria dos órgãos do trato gastrintestinal e a digestibilidade e retenção de matéria seca e nitrogênio. O maior ganho de peso na fase de 1 a 14 dias de idade foi obtido com a ração com 22% de proteína bruta. Os níveis de arginina digestível tiveram efeito quadrático na conversão alimentar na fase de 1 a 10 dias de idade. O peso do esôfago e inglúvio foi maior nas aves alimentadas com a ração com 20% de proteína bruta, no entanto, houve efeito quadrático dos níveis de arginina digestível sobre o comprimento do intestino aos 10 dias de idade e sobre o peso do esôfago + inglúvio aos 3 dias de idade. Houve interação entre os níveis de proteína bruta e arginina digestível para o peso relativo do fígado aos 14 dias, que respondeu de forma quadrática ao nível de 20% de proteína bruta, e para o comprimento de intestino, cujo maior valor foi obtido com os níveis de 22% de proteína bruta e 1,603% de arginina digestível. O balanço e a retenção de nitrogênio foram maiores no nível de 22% de proteína bruta. O nível de 1,363% de arginina digestível atende às exigências nutricionais dos frangos de corte na fase pré-inicial.This experiment was carried out to evaluate levels of crude protein and digestible arginine in pre-starter broiler ration and their effects on the performance of the broilers from 7 to

  7. Arginine Supplementation Recovered the IFN-γ-Mediated Decrease in Milk Protein and Fat Synthesis by Inhibiting the GCN2/eIF2α Pathway, Which Induces Autophagy in Primary Bovine Mammary Epithelial Cells

    Science.gov (United States)

    Xia, Xiaojing; Che, Yanyi; Gao, Yuanyuan; Zhao, Shuang; Ao, Changjin; Yang, Hongjian; Liu, Juxiong; Liu, Guowen; Han, Wenyu; Wang, Yuping; Lei, Liancheng

    2016-01-01

    During the lactation cycle of the bovine mammary gland, autophagy is induced in bovine mammary epithelial cells (BMECs) as a cellular homeostasis and survival mechanism. Interferon gamma (IFN-γ) is an important antiproliferative and apoptogenic factor that has been shown to induce autophagy in multiple cell lines in vitro. However, it remains unclear whether IFN-γ can induce autophagy and whether autophagy affects milk synthesis in BMECs. To understand whether IFN-γ affects milk synthesis, we isolated and purified primary BMECs and investigated the effect of IFN-γ on milk synthesis in primary BMECs in vitro. The results showed that IFN-γ significantly inhibits milk synthesis and that autophagy was clearly induced in primary BMECs in vitro within 24 h. Interestingly, autophagy was observed following IFN-γ treatment, and the inhibition of autophagy can improve milk protein and milk fat synthesis. Conversely, upregulation of autophagy decreased milk synthesis. Furthermore, mechanistic analysis confirmed that IFN-γ mediated autophagy by depleting arginine and inhibiting the general control nonderepressible-2 kinase (GCN2)/eukaryotic initiation factor 2α (eIF2α) signaling pathway in BMECs. Then, it was found that arginine supplementation could attenuate IFN-γ-induced autophagy and recover milk synthesis to some extent. These findings may not only provide a novel measure for preventing the IFN-γ-induced decrease in milk quality but also a useful therapeutic approach for IFN-γ-associated breast diseases in other animals and humans. PMID:27025389

  8. Carnitine/acylcarnitine translocase and carnitine palmitoyltransferase 2 form a complex in the inner mitochondrial membrane.

    Science.gov (United States)

    Console, Lara; Giangregorio, Nicola; Indiveri, Cesare; Tonazzi, Annamaria

    2014-09-01

    Carnitine/acylcarnitine translocase and carnitine palmitoyltransferase 2 are members of the carnitine system, which are responsible of the regulation of the mitochondrial CoA/acyl-CoA ratio and of supplying substrates for the ß-oxidation to mitochondria. This study, using cross-Linking reagent, Blue native electrophoresis and immunoprecipitation followed by detection with immunoblotting, shows conclusive evidence about the interaction between carnitine palmitoyltransferase 2 and carnitine/acylcarnitine translocase supporting the channeling of acylcarnitines and carnitine at level of the inner mitochondrial membrane. PMID:24898781

  9. Arginine side chain interactions and the role of arginine as a gating charge carrier in voltage sensitive ion channels

    Science.gov (United States)

    Armstrong, Craig T.; Mason, Philip E.; Anderson, J. L. Ross; Dempsey, Christopher E.

    2016-02-01

    Gating charges in voltage-sensing domains (VSD) of voltage-sensitive ion channels and enzymes are carried on arginine side chains rather than lysine. This arginine preference may result from the unique hydration properties of the side chain guanidinium group which facilitates its movement through a hydrophobic plug that seals the center of the VSD, as suggested by molecular dynamics simulations. To test for side chain interactions implicit in this model we inspected interactions of the side chains of arginine and lysine with each of the 19 non-glycine amino acids in proteins in the protein data bank. The arginine guanidinium interacts with non-polar aromatic and aliphatic side chains above and below the guanidinium plane while hydrogen bonding with polar side chains is restricted to in-plane positions. In contrast, non-polar side chains interact largely with the aliphatic part of the lysine side chain. The hydration properties of arginine and lysine are strongly reflected in their respective interactions with non-polar and polar side chains as observed in protein structures and in molecular dynamics simulations, and likely underlie the preference for arginine as a mobile charge carrier in VSD.

  10. Dietary arginine and linear growth

    DEFF Research Database (Denmark)

    van Vught, Anneke J A H; Dagnelie, Pieter C; Arts, Ilja C W;

    2013-01-01

    The amino acid arginine is a well-known growth hormone (GH) stimulator and GH is an important modulator of linear growth. The aim of the present study was to investigate the effect of dietary arginine on growth velocity in children between 7 and 13 years of age. Data from the Copenhagen School...

  11. Carnitine-acylcarnitine translocase deficiency, clinical, biochemical and genetic aspects.

    Science.gov (United States)

    Rubio-Gozalbo, M E; Bakker, J A; Waterham, H R; Wanders, R J A

    2004-01-01

    The carnitine-acylcarnitine translocase (CACT) is one of the components of the carnitine cycle. The carnitine cycle is necessary to shuttle long-chain fatty acids from the cytosol into the intramitochondrial space where mitochondrial beta-oxidation of fatty acids takes place. The oxidation of fatty acids yields acetyl-coenzyme A (CoA) units, which may either be degraded to CO(2) and H(2)O in the citric acid cycle to produce ATP or converted into ketone bodies which occurs in liver and kidneys. Metabolic consequences of a defective CACT are hypoketotic hypoglycaemia under fasting conditions, hyperammonemia, elevated creatine kinase and transaminases, dicarboxylic aciduria, very low free carnitine and an abnormal acylcarnitine profile with marked elevation of the long-chain acylcarnitines. Clinical signs and symptoms in CACT deficient patients, are a combination of energy depletion and endogenous toxicity. The predominantly affected organs are brain, heart and skeletal muscle, and liver, leading to neurological abnormalities, cardiomyopathy and arrythmias, skeletal muscle damage and liver dysfunction. Most patients become symptomatic in the neonatal period with a rapidly progressive deterioration and a high mortality rate. However, presentations at a later age with a milder phenotype have also been reported. The therapeutic approach is the same as in other long-chain fatty acid disorders and includes intravenous glucose (+/- insulin) administration to maximally inhibit lipolysis and subsequent fatty acid oxidation during the acute deterioration, along with other measures such as ammonia detoxification, depending on the clinical features. Long-term strategy consists of avoidance of fasting with frequent meals and a special diet with restriction of long-chain fatty acids. Due to the extremely low free carnitine concentrations, carnitine supplementation is often needed. Acylcarnitine profiling in plasma is the assay of choice for the diagnosis at a metabolite level

  12. NCBI nr-aa BLAST: CBRC-TTRU-01-1371 [SEVENS

    Lifescience Database Archive (English)

    Full Text Available ndent protein export [Candidatus Hamiltonella defensa 5AT (Acyrthosiphon pisum)] gb|ACQ68339.1| twin-arginine... translocase subunit, sec-independent protein export [Candidatus Hamiltonella defensa 5AT (Acyrthosiphon pisum)] YP_002924487.1 0.89 24% ... ...CBRC-TTRU-01-1371 ref|YP_002924487.1| twin-arginine translocase subunit, sec-indepe

  13. PGC-1β regulates mouse carnitine–acylcarnitine translocase through estrogen-related receptor α

    International Nuclear Information System (INIS)

    Highlights: ► The Cact gene is induced in mouse skeletal muscle after 24 h of fasting. ► The Cact gene contains a functional consensus sequence for ERR. ► This sequence binds ERRα both in vivo and in vitro. ► This ERRE is required for the activation of Cact expression by the PGC-1/ERR axis. ► Our results add Cact as a genuine gene target of these transcriptional regulators. -- Abstract: Carnitine/acylcarnitine translocase (CACT) is a mitochondrial-membrane carrier proteins that mediates the transport of acylcarnitines into the mitochondrial matrix for their oxidation by the mitochondrial fatty acid-oxidation pathway. CACT deficiency causes a variety of pathological conditions, such as hypoketotic hypoglycemia, cardiac arrest, hepatomegaly, hepatic dysfunction and muscle weakness, and it can be fatal in newborns and infants. Here we report that expression of the Cact gene is induced in mouse skeletal muscle after 24 h of fasting. To gain insight into the control of Cact gene expression, we examine the transcriptional regulation of the mouse Cact gene. We show that the 5′-flanking region of this gene is transcriptionally active and contains a consensus sequence for the estrogen-related receptor (ERR), a member of the nuclear receptor family of transcription factors. This sequence binds ERRαin vivo and in vitro and is required for the activation of Cact expression by the peroxisome proliferator-activated receptor gamma coactivator (PGC)-1/ERR axis. We also demonstrate that XTC790, the inverse agonist of ERRα, specifically blocks Cact activation by PGC-1β in C2C12 cells.

  14. PGC-1{beta} regulates mouse carnitine-acylcarnitine translocase through estrogen-related receptor {alpha}

    Energy Technology Data Exchange (ETDEWEB)

    Gacias, Mar; Perez-Marti, Albert; Pujol-Vidal, Magdalena; Marrero, Pedro F. [Department of Biochemistry and Molecular Biology, School of Pharmacy and the Institute of Biomedicine of the University of Barcelona (IBUB) (Spain); Haro, Diego, E-mail: dharo@ub.edu [Department of Biochemistry and Molecular Biology, School of Pharmacy and the Institute of Biomedicine of the University of Barcelona (IBUB) (Spain); Relat, Joana [Department of Biochemistry and Molecular Biology, School of Pharmacy and the Institute of Biomedicine of the University of Barcelona (IBUB) (Spain)

    2012-07-13

    Highlights: Black-Right-Pointing-Pointer The Cact gene is induced in mouse skeletal muscle after 24 h of fasting. Black-Right-Pointing-Pointer The Cact gene contains a functional consensus sequence for ERR. Black-Right-Pointing-Pointer This sequence binds ERR{alpha} both in vivo and in vitro. Black-Right-Pointing-Pointer This ERRE is required for the activation of Cact expression by the PGC-1/ERR axis. Black-Right-Pointing-Pointer Our results add Cact as a genuine gene target of these transcriptional regulators. -- Abstract: Carnitine/acylcarnitine translocase (CACT) is a mitochondrial-membrane carrier proteins that mediates the transport of acylcarnitines into the mitochondrial matrix for their oxidation by the mitochondrial fatty acid-oxidation pathway. CACT deficiency causes a variety of pathological conditions, such as hypoketotic hypoglycemia, cardiac arrest, hepatomegaly, hepatic dysfunction and muscle weakness, and it can be fatal in newborns and infants. Here we report that expression of the Cact gene is induced in mouse skeletal muscle after 24 h of fasting. To gain insight into the control of Cact gene expression, we examine the transcriptional regulation of the mouse Cact gene. We show that the 5 Prime -flanking region of this gene is transcriptionally active and contains a consensus sequence for the estrogen-related receptor (ERR), a member of the nuclear receptor family of transcription factors. This sequence binds ERR{alpha}in vivo and in vitro and is required for the activation of Cact expression by the peroxisome proliferator-activated receptor gamma coactivator (PGC)-1/ERR axis. We also demonstrate that XTC790, the inverse agonist of ERR{alpha}, specifically blocks Cact activation by PGC-1{beta} in C2C12 cells.

  15. 5'-Single-stranded/duplex DNA junctions are loading sites for E. coli UvrD translocase.

    Science.gov (United States)

    Tomko, Eric J; Jia, Haifeng; Park, Jeehae; Maluf, Nasib K; Ha, Taekjip; Lohman, Timothy M

    2010-11-17

    Escherichia coli UvrD is a 3'-5' superfamily 1A helicase/translocase involved in a variety of DNA metabolic processes. UvrD can function either as a helicase or only as an single-stranded DNA (ssDNA) translocase. The switch between these activities is controlled in vitro by the UvrD oligomeric state; a monomer has ssDNA translocase activity, whereas at least a dimer is needed for helicase activity. Although a 3'-ssDNA partial duplex provides a high-affinity site for a UvrD monomer, here we show that a monomer also binds with specificity to DNA junctions possessing a 5'-ssDNA flanking region and can initiate translocation from this site. Thus, a 5'-ss-duplex DNA junction can serve as a high-affinity loading site for the monomeric UvrD translocase, whereas a 3'-ss-duplex DNA junction inhibits both translocase and helicase activity of the UvrD monomer. Furthermore, the 2B subdomain of UvrD is important for this junction specificity. This highlights a separation of helicase and translocase function for UvrD and suggests that a monomeric UvrD translocase can be loaded at a 5'-ssDNA junction when translocation activity alone is needed. PMID:20877334

  16. Homology modeling of major intrinsic proteins in rice, maize and Arabidopsis: comparative analysis of transmembrane helix association and aromatic/arginine selectivity filters

    OpenAIRE

    Sankararamakrishnan Ramasubbu; Bansal Anjali

    2007-01-01

    Abstract Background The major intrinsic proteins (MIPs) facilitate the transport of water and neutral solutes across the lipid bilayers. Plant MIPs are believed to be important in cell division and expansion and in water transport properties in response to environmental conditions. More than 30 MIP sequences have been identified in Arabidopsis thaliana, maize and rice. Plasma membrane intrinsic proteins (PIPs), tonoplast intrinsic proteins (TIPs), Nod26-like intrinsic protein (NIPs) and small...

  17. An Arginine Deprivation Response Pathway Is Induced in Leishmania during Macrophage Invasion.

    Directory of Open Access Journals (Sweden)

    Adele Goldman-Pinkovich

    2016-04-01

    Full Text Available Amino acid sensing is an intracellular function that supports nutrient homeostasis, largely through controlled release of amino acids from lysosomal pools. The intracellular pathogen Leishmania resides and proliferates within human macrophage phagolysosomes. Here we describe a new pathway in Leishmania that specifically senses the extracellular levels of arginine, an amino acid that is essential for the parasite. During infection, the macrophage arginine pool is depleted due to its use to produce metabolites (NO and polyamines that constitute part of the host defense response and its suppression, respectively. We found that parasites respond to this shortage of arginine by up-regulating expression and activity of the Leishmania arginine transporter (LdAAP3, as well as several other transporters. Our analysis indicates the parasite monitors arginine levels in the environment rather than the intracellular pools. Phosphoproteomics and genetic analysis indicates that the arginine-deprivation response is mediated through a mitogen-activated protein kinase-2-dependent signaling cascade.

  18. An Arginine Deprivation Response Pathway Is Induced in Leishmania during Macrophage Invasion.

    Science.gov (United States)

    Goldman-Pinkovich, Adele; Balno, Caitlin; Strasser, Rona; Zeituni-Molad, Michal; Bendelak, Keren; Rentsch, Doris; Ephros, Moshe; Wiese, Martin; Jardim, Armando; Myler, Peter J; Zilberstein, Dan

    2016-04-01

    Amino acid sensing is an intracellular function that supports nutrient homeostasis, largely through controlled release of amino acids from lysosomal pools. The intracellular pathogen Leishmania resides and proliferates within human macrophage phagolysosomes. Here we describe a new pathway in Leishmania that specifically senses the extracellular levels of arginine, an amino acid that is essential for the parasite. During infection, the macrophage arginine pool is depleted due to its use to produce metabolites (NO and polyamines) that constitute part of the host defense response and its suppression, respectively. We found that parasites respond to this shortage of arginine by up-regulating expression and activity of the Leishmania arginine transporter (LdAAP3), as well as several other transporters. Our analysis indicates the parasite monitors arginine levels in the environment rather than the intracellular pools. Phosphoproteomics and genetic analysis indicates that the arginine-deprivation response is mediated through a mitogen-activated protein kinase-2-dependent signaling cascade. PMID:27043018

  19. Characterization of arginine decarboxylase from Dianthus caryophyllus.

    Science.gov (United States)

    Ha, Byung Hak; Cho, Ki Joon; Choi, Yu Jin; Park, Ky Young; Kim, Kyung Hyun

    2004-04-01

    Arginine decarboxylase (ADC, EC 4.1.1.9) is a key enzyme in the biosynthesis of polyamines in higher plants, whereas ornithine decarboxylase represents the sole pathway of polyamine biosynthesis in animals. Previously, we characterized a genomic clone from Dianthus caryophyllus, in which the deduced polypeptide of ADC was 725 amino acids with a molecular mass of 78 kDa. In the present study, the ADC gene was subcloned into the pGEX4T1 expression vector in combination with glutathione S-transferase (GST). The fusion protein GST-ADC was water-soluble and thus was purified by sequential GSTrap-arginine affinity chromatography. A thrombin-mediated on-column cleavage reaction was employed to release free ADC from GST. Hiload superdex gel filtration FPLC was then used to obtain a highly purified ADC. The identity of the ADC was confirmed by immunoblot analysis, and its specific activity with respect to (14)C-arginine decarboxylation reaction was determined to be 0.9 CO(2) pkat mg(-1) protein. K(m) and V(max) of the reaction between ADC and the substrate were 0.077 +/- 0.001 mM and 6.0 +/- 0.6 pkat mg(-1) protein, respectively. ADC activity was reduced by 70% in the presence of 0.1 mM Cu(2+) or CO(2+), but was only marginally affected by Mg(2+), or Ca(2+) at the same concentration. Moreover, spermine at 1 mM significantly reduced its activity by 30%. PMID:15120115

  20. Inhibition of Nonsmall Cell Lung Cancer Cell Migration by Protein Arginine Methyltransferase 1-small Hairpin RNA Through Inhibiting Epithelial-mesenchymal Transition, Extracellular Matrix Degradation, and Src Phosphorylation In Vitro

    Directory of Open Access Journals (Sweden)

    Ting Zhang

    2015-01-01

    Full Text Available Background: Protein arginine methyltransferases 1 (PRMT1 is over-expressed in a variety of cancers, including lung cancer, and is correlated with a poor prognosis of tumor development. This study aimed to investigate the role of PRMT1 in nonsmall cell lung cancer (NSCLC migration in vitro. Methods: In this study, PRMT1 expression in the NSCLC cell line A549 was silenced using lentiviral vector-mediated short hairpin RNAs. Cell migration was measured using both scratch wound healing and transwell cell migration assays. The mRNA expression levels of matrix metalloproteinase 2 (MMP-2 and tissue inhibitor of metalloproteinase 1, 2 (TIMP1, 2 were measured using quantitative real-time reverse transcription-polymerase chain reaction. The expression levels of protein markers for epithelial-mesenchymal transition (EMT (E-cadherin, N-cadherin, focal adhesion kinase (FAK, Src, AKT, and their corresponding phosphorylated states were detected by Western blot. Results: Cell migration was significantly inhibited in the PRMT1 silenced group compared to the control group. The mRNA expression of MMP-2 decreased while TIMP1 and TIMP2 increased significantly. E-cadherin mRNA expression also increased while N-cadherin decreased. Only phosphorylated Src levels decreased in the silenced group while FAK or AKT remained unchanged. Conclusions: PRMT1-small hairpin RNA inhibits the migration abilities of NSCLC A549 cells by inhibiting EMT, extracellular matrix degradation, and Src phosphorylation in vitro.

  1. Identification of protein secretion systems and novel secreted proteins in Rhizobium leguminosarum bv. viciae

    Directory of Open Access Journals (Sweden)

    Krehenbrink Martin

    2008-01-01

    Full Text Available Abstract Background Proteins secreted by bacteria play an important role in infection of eukaryotic hosts. Rhizobia infect the roots of leguminous plants and establish a mutually beneficial symbiosis. Proteins secreted during the infection process by some rhizobial strains can influence infection and modify the plant defence signalling pathways. The aim of this study was to systematically analyse protein secretion in the recently sequenced strain Rhizobium leguminosarum bv. viciae 3841. Results Similarity searches using defined protein secretion systems from other Gram-negative bacteria as query sequences revealed that R. l. bv. viciae 3841 has ten putative protein secretion systems. These are the general export pathway (GEP, a twin-arginine translocase (TAT secretion system, four separate Type I systems, one putative Type IV system and three Type V autotransporters. Mutations in genes encoding each of these (except the GEP were generated, but only mutations affecting the PrsDE (Type I and TAT systems were observed to affect the growth phenotype and the profile of proteins in the culture supernatant. Bioinformatic analysis and mass fingerprinting of tryptic fragments of culture supernatant proteins identified 14 putative Type I substrates, 12 of which are secreted via the PrsDE, secretion system. The TAT mutant was defective for the symbiosis, forming nodules incapable of nitrogen fixation. Conclusion None of the R. l. bv. viciae 3841 protein secretion systems putatively involved in the secretion of proteins to the extracellular space (Type I, Type IV, Type V is required for establishing the symbiosis with legumes. The PrsDE (Type I system was shown to be the major route of protein secretion in non-symbiotic cells and to secrete proteins of widely varied size and predicted function. This is in contrast to many Type I systems from other bacteria, which typically secrete specific substrates encoded by genes often localised in close proximity to

  2. Identification and characterization of two temperature-induced surface-associated proteins of Streptococcus suis with high homologies to members of the arginine deiminase system of Streptococcus pyogenes

    NARCIS (Netherlands)

    Winterhoff, N.; Goethe, R.; Gruening, P.; Rohde, M.; Kalisz, H.; Smith, H.E.; Valentin-Weigand, P.

    2002-01-01

    The present study was performed to identify stress-induced putative virulence proteins of Streptococcus suis. For this, protein expression patterns of streptococci grown at 32, 37, and 42°C were compared by one- and two-dimensional gel electrophoresis. Temperature shifts from 32 and 37 to 42°C induc

  3. Identification of oligonucleotide sequences that direct the movement of the Escherichia coli FtsK translocase

    OpenAIRE

    Levy, Oren; Ptacin, Jerod L.; Pease, Paul J.; Gore, Jeff; Eisen, Michael B.; Bustamante, Carlos; Cozzarelli, Nicholas R.

    2005-01-01

    FtsK from Escherichia coli is a fast and sequence-directed DNA translocase with roles in chromosome dimer resolution, segregation, and decatenation. From the movement of single FtsK particles on defined DNA substrates and an analysis of skewed DNA sequences in bacteria, we identify GNGNAGGG, its complement, or both as a sequence motif that controls translocation directionality. GNGNAGGG is skewed so that it is predominantly on the leading strand of chromosomal replication. Translocation acros...

  4. Two Proteins with Ornithine Acetyltransferase Activity Show Different Functions in Streptomyces clavuligerus: Oat2 Modulates Clavulanic Acid Biosynthesis in Response to Arginine

    OpenAIRE

    de la Fuente, A.; Martín, J F; Rodríguez-García, A.; Liras, P

    2004-01-01

    The oat2 gene, located in the clavulanic acid gene cluster in Streptomyces clavuligerus, is similar to argJ, which encodes N-acetylornithine:glutamic acid acetyltransferase activity. Purified proteins obtained by expression in Escherichia coli of the argJ and oat2 genes of S. clavuligerus posses N-acetyltransferase activity. The kinetics and substrate specificities of both proteins are very similar. Deletion of the oat2 gene did not affect the total N-acetylornithine transferase activity and ...

  5. Dietary arginine and linear growth: the Copenhagen School Child Intervention Study.

    Science.gov (United States)

    van Vught, Anneke J A H; Dagnelie, Pieter C; Arts, Ilja C W; Froberg, Karsten; Andersen, Lars B; El-Naaman, Bianca; Bugge, Anna; Nielsen, Birgit M; Heitman, Berit L

    2013-03-28

    The amino acid arginine is a well-known growth hormone (GH) stimulator and GH is an important modulator of linear growth. The aim of the present study was to investigate the effect of dietary arginine on growth velocity in children between 7 and 13 years of age. Data from the Copenhagen School Child Intervention Study during 2001-2 (baseline), and at 3-year and 7-year follow-up, were used. Arginine intake was estimated via a 7 d precoded food diary at baseline and 3-year follow-up. Data were analysed in a multilevel structure in which children were embedded within schools. Random intercept and slopes were defined to estimate the association between arginine intake and growth velocity, including the following covariates: sex; age; baseline height; energy intake; puberty stage at 7-year follow-up and intervention/control group. The association between arginine intake and growth velocity was significant for the third and fourth quintile of arginine intake (2.5-2.8 and 2.8-3.2 g/d, respectively) compared with the first quintile ( < 2.2 g/d) (P for trend = 0.04). Protein intake (excluding arginine) was significantly associated with growth velocity; however, the association was weaker than the association between arginine intake and growth velocity (P for trend = 0.14). The results of the present study suggest a dose-dependent physiological role of habitual protein intake, and specifically arginine intake, on linear growth in normally growing children. However, since the study was designed in healthy children, we cannot firmly conclude whether arginine supplementation represents a relevant clinical strategy. Further research is needed to investigate whether dietary arginine may represent a nutritional strategy potentially advantageous for the prevention and treatment of short stature. PMID:23046689

  6. An arginine-rich motif of ring finger protein 4 (RNF4) oversees the recruitment and degradation of the phosphorylated and SUMOylated Krüppel-associated box domain-associated protein 1 (KAP1)/TRIM28 protein during genotoxic stress.

    Science.gov (United States)

    Kuo, Ching-Ying; Li, Xu; Kong, Xiang-Qian; Luo, Cheng; Chang, Che-Chang; Chung, Yiyin; Shih, Hsiu-Ming; Li, Keqin Kathy; Ann, David K

    2014-07-25

    Krüppel-associated box domain-associated protein 1 (KAP1) is a universal transcriptional corepressor that undergoes multiple posttranslational modifications (PTMs), including SUMOylation and Ser-824 phosphorylation. However, the functional interplay of KAP1 PTMs in regulating KAP1 turnover during DNA damage response remains unclear. To decipher the role and cross-talk of multiple KAP1 PTMs, we show here that DNA double strand break-induced KAP1 Ser-824 phosphorylation promoted the recruitment of small ubiquitin-like modifier (SUMO)-targeted ubiquitin E3 ligase, ring finger protein 4 (RNF4), and subsequent RNF4-mediated, SUMO-dependent degradation. Besides the SUMO interacting motif (SIM), a previously unrecognized, but evolutionarily conserved, arginine-rich motif (ARM) in RNF4 acts as a novel recognition motif for selective target recruitment. Results from combined mutagenesis and computational modeling studies suggest that RNF4 utilizes concerted bimodular recognition, namely SIM for Lys-676 SUMOylation and ARM for Ser(P)-824 of simultaneously phosphorylated and SUMOylated KAP1 (Ser(P)-824-SUMO-KAP1). Furthermore, we proved that arginines 73 and 74 within the ARM of RNF4 are required for efficient recruitment to KAP1 or accelerated degradation of promyelocytic leukemia protein (PML) under stress. In parallel, results of bimolecular fluorescence complementation assays validated the role of the ARM in recognizing Ser(P)-824 in living cells. Taken together, we establish that the ARM is required for RNF4 to efficiently target Ser(P)-824-SUMO-KAP1, conferring ubiquitin Lys-48-mediated proteasomal degradation in the context of double strand breaks. The conservation of such a motif may possibly explain the requirement for timely substrate selectivity determination among a myriad of SUMOylated proteins under stress conditions. Thus, the ARM dynamically regulates the SIM-dependent recruitment of targets to RNF4, which could be critical to dynamically fine-tune the

  7. Co- and post-translational translocation through the protein-conducting channel : analogous mechanisms at work?

    NARCIS (Netherlands)

    Mitra, Kakoli; Frank, Joachim; Driessen, Arnold

    2006-01-01

    Many proteins are translocated across, or integrated into, membranes. Both functions are fulfilled by the 'translocon/translocase', which contains a membrane-embedded proteinconducting channel (PCC) and associated soluble factors that drive translocation and insertion reactions using nucleotide trip

  8. Inhibition of Nonsmall Cell Lung Cancer Cell Migration by Protein Arginine Methyltransferase 1-small Hairpin RNA Through Inhibiting Epithelial-mesenchymal Transition,Extracellular Matrix Degradation, and Src Phosphorylation In Vitro

    Institute of Scientific and Technical Information of China (English)

    Ting Zhang; Ge Cui; Yun-Liang Yao; Yue Guo; Qi-Chun Wang; Xi-Ning Li; Wen-Ming Feng

    2015-01-01

    Background:Protein arginine methyltransferases 1 (PRMT1) is over-expressed in a variety of cancers,including lung cancer,and is correlated with a poor prognosis of tumor development.This study aimed to investigate the role of PRMT1 in nonsmall cell lung cancer (NSCLC) migration in vitro.Methods:In this study,PRMT1 expression in the NSCLC cell line A549 was silenced using lentiviral vector-mediated short hairpin RNAs.Cell migration was measured using both scratch wound healing and transwell cell migration assays.The mRNA expression levels of matrix metalloproteinase 2 (MMP-2) and tissue inhibitor ofmetalloproteinase 1,2 (TIMP l,2) were measured using quantitative real-time reverse transcription-polymerase chain reaction.The expression levels of protein markers for epithelial-mesenchymal transition (EMT) (E-cadherin,N-cadherin),focal adhesion kinase (FAK),Src,AKT,and their corresponding phosphorylated states were detected by Western blot.Results:Cell migration was significantly inhibited in the PRMT1 silenced group compared to the control group.The mRNA expression of MMP-2 decreased while TIMP 1 and TIMP2 increased significantly.E-cadherin mRNA expression also increased while N-cadherin decreased.Only phosphorylated Src levels decreased in the silenced group while FAK or AKT remained unchanged.Conclusions:PRMT1-small hairpin RNA inhibits the migration abilities of NSCLC A549 cells by inhibiting EMT,extracellular matrix degradation,and Src phosphorylation in vitro.

  9. Environmental salinity determines the specificity and need for Tat-dependent secretion of the YwbN protein in Bacillus subtilis.

    Directory of Open Access Journals (Sweden)

    René van der Ploeg

    Full Text Available Twin-arginine protein translocation (Tat pathways are required for transport of folded proteins across bacterial, archaeal and chloroplast membranes. Recent studies indicate that Tat has evolved into a mainstream pathway for protein secretion in certain halophilic archaea, which thrive in highly saline environments. Here, we investigated the effects of environmental salinity on Tat-dependent protein secretion by the Gram-positive soil bacterium Bacillus subtilis, which encounters widely differing salt concentrations in its natural habitats. The results show that environmental salinity determines the specificity and need for Tat-dependent secretion of the Dyp-type peroxidase YwbN in B. subtilis. Under high salinity growth conditions, at least three Tat translocase subunits, namely TatAd, TatAy and TatCy, are involved in the secretion of YwbN. Yet, a significant level of Tat-independent YwbN secretion is also observed under these conditions. When B. subtilis is grown in medium with 1% NaCl or without NaCl, the secretion of YwbN depends strictly on the previously described "minimal Tat translocase" consisting of the TatAy and TatCy subunits. Notably, in medium without NaCl, both tatAyCy and ywbN mutants display significantly reduced exponential growth rates and severe cell lysis. This is due to a critical role of secreted YwbN in the acquisition of iron under these conditions. Taken together, our findings show that environmental conditions, such as salinity, can determine the specificity and need for the secretion of a bacterial Tat substrate.

  10. Kinetics of precursor interactions with the bacterial Tat translocase detected by real-time FRET.

    Science.gov (United States)

    Whitaker, Neal; Bageshwar, Umesh K; Musser, Siegfried M

    2012-03-30

    The Escherichia coli twin-arginine translocation (Tat) system transports fully folded and assembled proteins across the inner membrane into the periplasmic space. Traditionally, in vitro protein translocation studies have been performed using gel-based transport assays. This technique suffers from low time resolution, and often, an inability to distinguish between different steps in a continuously occurring translocation process. To address these limitations, we have developed an in vitro FRET-based assay that reports on an early step in the Tat translocation process in real-time. The natural Tat substrate pre-SufI was labeled with Alexa532 (donor), and the fluorescent protein mCherry (acceptor) was fused to the C terminus of TatB or TatC. The colored Tat proteins were easily visible during purification, enabling identification of a highly active inverted membrane vesicle (IMV) fraction yielding transport rates with NADH almost an order of magnitude faster than previously reported. When pre-SufI was bound to the translocon, FRET was observed for both Tat proteins. FRET was diminished upon addition of nonfluorescent pre-SufI, indicating that the initial binding step is reversible. When the membranes were energized with NADH, the FRET signal was lost after a short delay. These data suggest a model in which a Tat cargo initially associates with the TatBC complex, and an electric field gradient is required for the cargo to proceed to the next stage of transport. This cargo migration away from the TatBC complex requires a significant fraction of the total transport time. PMID:22315217

  11. Structural organization of the rat gene for the arginine vasopressin-neurophysin precursor

    OpenAIRE

    Schmale, H.; Heinsohn, S; Richter, D

    1983-01-01

    The rat arginine vasopressin-neurophysin precursor gene has been isolated from a genomic library cloned in lambda phage Charon 4A. Restriction mapping and nucleotide sequence analysis demonstrated that the gene is 1.85 kilobase pairs long and contains two intervening sequences located in the protein coding region. Exon A encodes a putative signal peptide, the hormone arginine vasopressin and the variable N terminus of the carrier protein neurophysin, exon B encodes the highly conserved middle...

  12. Membrane-Bound Dynamic Structure of an Arginine-Rich Cell-Penetrating Peptide, the Protein Transduction Domain of HIV TAT, from Solid-State NMR

    OpenAIRE

    Su, Yongchao; Alan J Waring; Ruchala, Piotr; Hong, Mei

    2010-01-01

    The protein transduction domain of HIV-1 TAT, TAT(48-60), is an efficient cell-penetrating peptide (CPP) that diffuses across the lipid membranes of cells despite eight cationic Arg and Lys residues. To understand its mechanism of membrane translocation against the free energy barrier, we have conducted solid-state NMR experiments to determine the site-specific conformation, dynamics, and lipid interaction of the TAT peptide in anionic lipid bilayers. We found that TAT(48-60) is a highly dyna...

  13. Dietary arginine affects energy metabolism through polyamine turnover in juvenile Atlantic salmon (Salmo salar).

    Science.gov (United States)

    Andersen, Synne M; Holen, Elisabeth; Aksnes, Anders; Rønnestad, Ivar; Zerrahn, Jens-Erik; Espe, Marit

    2013-12-14

    In the present study, quadruplicate groups of juvenile Atlantic salmon (Salmo salar) were fed plant protein-based diets with increasing arginine inclusions (range 28·8-37·4 g/kg DM) to investigate whether arginine supplementation affects growth and lipid accumulation through an elevated polyamine turnover. Dietary lysine was held at a constant concentration, just below the requirement. All other amino acids were balanced and equal in the diets. Arginine supplementation increased protein and fat accretion, without affecting the hepatosomatic or visceralsomatic indices. Dietary arginine correlated with putrescine in the liver (R 0·78, P= 0·01) and with ornithine in the muscle, liver and plasma (P= 0·0002, 0·003 and 0·0002, respectively). The mRNA of ornithine decarboxylase, the enzyme producing putrescine, was up-regulated in the white adipose tissue of fish fed the high-arginine inclusion compared with those fed the low-arginine diet. Concomitantly, spermidine/spermine-(N1)-acetyltransferase, the rate-limiting enzyme for polyamine turnover that consumes acetyl-CoA, showed an increased activity in the liver of fish fed the arginine-supplemented diets. In addition, lower acetyl-CoA concentrations were observed in the liver of fish fed the high-arginine diet, while ATP, which is used in the process of synthesising spermidine and spermine, did not show a similar trend. Gene expression of the rate-limiting enzyme for β-oxidation of long-chain fatty acids, carnitine palmitoyl transferase-1, was up-regulated in the liver of fish fed the high-arginine diet. Taken together, the data support that increased dietary arginine activates polyamine turnover and β-oxidation in the liver of juvenile Atlantic salmon and may act to improve the metabolic status of the fish. PMID:23656796

  14. Comparative analysis of serine/arginine-rich proteins across 27 eukaryotes: insights into sub-family classification and extent of alternative splicing.

    Directory of Open Access Journals (Sweden)

    Dale N Richardson

    Full Text Available Alternative splicing (AS of pre-mRNA is a fundamental molecular process that generates diversity in the transcriptome and proteome of eukaryotic organisms. SR proteins, a family of splicing regulators with one or two RNA recognition motifs (RRMs at the N-terminus and an arg/ser-rich domain at the C-terminus, function in both constitutive and alternative splicing. We identified SR proteins in 27 eukaryotic species, which include plants, animals, fungi and "basal" eukaryotes that lie outside of these lineages. Using RNA recognition motifs (RRMs as a phylogenetic marker, we classified 272 SR genes into robust sub-families. The SR gene family can be split into five major groupings, which can be further separated into 11 distinct sub-families. Most flowering plants have double or nearly double the number of SR genes found in vertebrates. The majority of plant SR genes are under purifying selection. Moreover, in all paralogous SR genes in Arabidopsis, rice, soybean and maize, one of the two paralogs is preferentially expressed throughout plant development. We also assessed the extent of AS in SR genes based on a splice graph approach (http://combi.cs.colostate.edu/as/gmap_SRgenes. AS of SR genes is a widespread phenomenon throughout multiple lineages, with alternative 3' or 5' splicing events being the most prominent type of event. However, plant-enriched sub-families have 57%-88% of their SR genes experiencing some type of AS compared to the 40%-54% seen in other sub-families. The SR gene family is pervasive throughout multiple eukaryotic lineages, conserved in sequence and domain organization, but differs in gene number across lineages with an abundance of SR genes in flowering plants. The higher number of alternatively spliced SR genes in plants emphasizes the importance of AS in generating splice variants in these organisms.

  15. The Protein Arginine Methylase 5 (PRMT5/SKB1) Gene Is Required for the Maintenance of Root Stem Cells in Response to DNA Damage.

    Science.gov (United States)

    Li, Qiuling; Zhao, Yan; Yue, Minghui; Xue, Yongbiao; Bao, Shilai

    2016-04-20

    Plant root stem cells and their surrounding microenvironment, namely the stem cell niche, are hypersensitive to DNA damage. However, the molecular mechanisms that help maintain the genome stability of root stem cells remain elusive. Here we show that the root stem cells in the skb1 (Shk1 kinase binding protein 1) mutant undergoes DNA damage-induced cell death, which is enhanced when combined with a lesion of the Ataxia-telangiectasia mutated (ATM) or the ATM/RAD3-related (ATR) genes, suggesting that the SKB1 plays a synergistically effect with ATM and ATR in DNA damage pathway. We also provide evidence that SKB1 is required for the maintenance of quiescent center (QC), a root stem cell niche, under DNA damage treatments. Furthermore, we report decreased and ectopic expression of SHORTROOT (SHR) in response to DNA damage in the skb1 root tips, while the expression of SCARECROW (SCR) remains unaffected. Our results uncover a new mechanism of plant root stem cell maintenance under DNA damage conditions that requires SKB1. PMID:27090604

  16. The role of the arginine metabolome in pain: implications for sickle cell disease

    Directory of Open Access Journals (Sweden)

    Bakshi N

    2016-03-01

    Full Text Available Nitya Bakshi,1–2 Claudia R Morris3–6 1Division of Pediatric Hematology-Oncology, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, USA; 2Aflac Cancer and Blood Disorders Center, Children’s Healthcare of Atlanta, Atlanta, GA, USA; 3Division of Pediatric Emergency Medicine, Department of Pediatrics, Emory University School of Medicine, Atlanta, GA, USA; 4Department of Emergency Medicine, Emory University School of Medicine, Atlanta, GA, USA; 5Emory-Children’s Center for Cystic Fibrosis and Airways Disease Research, Emory University School of Medicine, Atlanta, GA, USA; 6Pediatric Emergency Medicine, Children’s Healthcare of Atlanta, Atlanta, GA, USA Abstract: Sickle cell disease (SCD is the most common hemoglobinopathy in the US, affecting approximately 100,000 individuals in the US and millions worldwide. Pain is the hallmark of SCD, and a subset of patients experience pain virtually all of the time. Of interest, the arginine metabolome is associated with several pain mechanisms highlighted in this review. Since SCD is an arginine deficiency syndrome, the contribution of the arginine metabolome to acute and chronic pain in SCD is a topic in need of further attention. Normal arginine metabolism is impaired in SCD through various mechanisms that contribute to endothelial dysfunction, vaso-occlusion, pulmonary complications, risk of leg ulcers, and early mortality. Arginine is a semiessential amino acid that serves as a substrate for protein synthesis and is the precursor to nitric oxide (NO, polyamines, proline, glutamate, creatine, and agmatine. Since arginine is involved in multiple metabolic processes, a deficiency of this amino acid has the potential to disrupt many cellular and organ functions. NO is a potent vasodilator that is depleted in SCD and may contribute to vaso-occlusive pain. As the obligate substrate for NO production, arginine also plays a mechanistic role in SCD-related pain, although its

  17. An adenine nucleotide translocase (ANT) gene from Apostichopus japonicus; molecular cloning and expression analysis in response to lipopolysaccharide (LPS) challenge and thermal stress.

    Science.gov (United States)

    Liu, Qiu-Ning; Chai, Xin-Yue; Tu, Jie; Xin, Zhao-Zhe; Li, Chao-Feng; Jiang, Sen-Hao; Zhou, Chun-Lin; Tang, Bo-Ping

    2016-02-01

    The adenine nucleotide translocases (ANTs) play a vital role in energy metabolism via ADP/ATP exchange in eukaryotic cells. Apostichopus japonicus (Echinodermata: Holothuroidea) is an important economic species in China. Here, a cDNA representing an ANT gene of A. japonicus was isolated and characterized from respiratory tree and named AjANT. The full-length AjANT cDNA is 1924 bp, including a 5'-untranslated region (UTR) of 38 bp, 3'-UTR of 980 bp and an open reading frame (ORF) of 906 bp encoding a polypeptide of 301 amino acids. The protein contains three homologous repeat Mito_carr domains (Pfam00153). The deduced AjANT protein sequence has 49-81% in comparison to ANT proteins from other individuals. The predicted tertiary structure of AjANT protein is highly similar to animal ANT proteins. Phylogenetic analysis showed that the AjANT is closely related to Holothuroidea ANT genes. Real-time quantitative reverse transcription-PCR (qPCR) analysis showed that AjANT expression is higher in the respiratory tree than in other examined tissues. After thermal stress or LPS challenge, expression of AjANT was significantly fluctuant compared to the control. These results suggested that changes in the expression of ANT gene might be involved in immune defense and in protecting A. japonicus against thermal stress. PMID:26706223

  18. Arginase and Arginine Dysregulation in Asthma

    OpenAIRE

    Renée C. Benson; Hardy, Karen A.; Morris, Claudia R.

    2011-01-01

    In recent years, evidence has accumulated indicating that the enzyme arginase, which converts L-arginine into L-ornithine and urea, plays a key role in the pathogenesis of pulmonary disorders such as asthma through dysregulation of L-arginine metabolism and modulation of nitric oxide (NO) homeostasis. Allergic asthma is characterized by airway hyperresponsiveness, inflammation, and remodeling. Through substrate competition, arginase decreases bioavailability of L-arginine for nitric oxide syn...

  19. The RNA binding motif protein 15B (RBM15B/OTT3) is a functional competitor of serine-arginine (SR) proteins and antagonizes the positive effect of the CDK11p110-cyclin L2α complex on splicing.

    Science.gov (United States)

    Loyer, Pascal; Busson, Adeline; Trembley, Janeen H; Hyle, Judith; Grenet, Jose; Zhao, Wei; Ribault, Catherine; Montier, Tristan; Kidd, Vincent J; Lahti, Jill M

    2011-01-01

    Here, we report the identification of the RNA binding motif protein RBM15B/OTT3 as a new CDK11(p110) binding partner that alters the effects of CDK11 on splicing. RBM15B was initially identified as a binding partner of the Epstein-Barr virus mRNA export factor and, more recently, as a cofactor of the nuclear export receptor NXF1. In this study, we found that RBM15B co-elutes with CDK11(p110), cyclin L2α, and serine-arginine (SR) proteins, including SF2/ASF, in a large nuclear complex of ∼1-MDa molecular mass following size exclusion chromatography. Using co-immunoprecipitation experiments and in vitro pulldown assays, we mapped two distinct domains of RBM15B that are essential for its direct interaction with the N-terminal extension of CDK11(p110), cyclin L2α, and SR proteins such as 9G8 and SF2/ASF. Finally, we established that RBM15B is a functional competitor of the SR proteins SF2/ASF and 9G8, inhibits formation of the functional spliceosomal E complex, and antagonizes the positive effect of the CDK11(p110)-cyclin L2α complex on splicing both in vitro and in vivo. PMID:21044963

  20. The RNA Binding Motif Protein 15B (RBM15B/OTT3) Is a Functional Competitor of Serine-Arginine (SR) Proteins and Antagonizes the Positive Effect of the CDK11p110-Cyclin L2α Complex on Splicing*

    Science.gov (United States)

    Loyer, Pascal; Busson, Adeline; Trembley, Janeen H.; Hyle, Judith; Grenet, Jose; Zhao, Wei; Ribault, Catherine; Montier, Tristan; Kidd, Vincent J.; Lahti, Jill M.

    2011-01-01

    Here, we report the identification of the RNA binding motif protein RBM15B/OTT3 as a new CDK11p110 binding partner that alters the effects of CDK11 on splicing. RBM15B was initially identified as a binding partner of the Epstein-Barr virus mRNA export factor and, more recently, as a cofactor of the nuclear export receptor NXF1. In this study, we found that RBM15B co-elutes with CDK11p110, cyclin L2α, and serine-arginine (SR) proteins, including SF2/ASF, in a large nuclear complex of ∼1-MDa molecular mass following size exclusion chromatography. Using co-immunoprecipitation experiments and in vitro pulldown assays, we mapped two distinct domains of RBM15B that are essential for its direct interaction with the N-terminal extension of CDK11p110, cyclin L2α, and SR proteins such as 9G8 and SF2/ASF. Finally, we established that RBM15B is a functional competitor of the SR proteins SF2/ASF and 9G8, inhibits formation of the functional spliceosomal E complex, and antagonizes the positive effect of the CDK11p110-cyclin L2α complex on splicing both in vitro and in vivo. PMID:21044963

  1. Influence of in ovo injection of L-arginine on productive and physiological performance of quail

    Directory of Open Access Journals (Sweden)

    W. K. Al–Hayani,

    2011-07-01

    Full Text Available This study evaluated the influence of inoculation of different levels of L–arginine into eggs of 0-day-old quail embryos. On 0 day of incubation, 480 eggs (120 for each treatment group were injected with 0% arginine (C group; 1% arginine (T1; 2% arginine (T2; or 3% arginine (T3. After hatching, 336 quail chicks (84 chicks produced from each in ovo injection treatment were placed in an experimental quail house and distributed into 4 treatment groups of 3 replicates each with 16 quail chicks for each replicate. Traits determined in this study were hatchability rate, initial body weight (7 days of age, final body weight (42 days old, feed intake, weight gain, feed conversion ratio, proportional weights of carcass, breast, legs, back bone, wings, neck, abdominal fat, liver, heart, and gizzard, blood serum glucose, protein, cholesterol, total lipids, triglycerides, calcium and phosphorus and Results revealed that in ovo injection with different levels of L–arginine on 0 day of incubation resulted in significant increase (P≤0.05 in hatchability rate, initial body weight, final body weight, feed conversion ratio and serum glucose, protein, total protein, calcium, phosphorus and proportional weights of carcass, breast, legs, liver, heart, and gizzard and significant decrease (P≤0.05 in serum cholesterol, total lipids, triglycerides and proportional weight of back bone, wings and abdominal fat. In conclusion, the inoculation of different levels of L–arginine into eggs of 0–day–old quail embryos especially at the levels of 2% and 3% resulted in significant improvement in productive and physiological performance of quail. Hence in ovo injection with L–arginine could be used as a beneficial tool for enhance productive performance of quail.

  2. Mechanism of arginine regulation of acetylglutamate synthase, the first enzyme of arginine synthesis.

    Science.gov (United States)

    Sancho-Vaello, Enea; Fernández-Murga, María L; Rubio, Vicente

    2009-01-01

    N-acetyl-L-glutamate synthase (NAGS), the first enzyme of arginine biosynthesis in bacteria/plants and an essential urea cycle activator in animals, is, respectively, arginine-inhibited and activated. Arginine binds to the hexameric ring-forming amino acid kinase (AAK) domain of NAGS. We show that arginine inhibits Pseudomonas aeruginosa NAGS by altering the functions of the distant, substrate binding/catalytic GCN5-related N-acetyltransferase (GNAT) domain, increasing K(m)(Glu), decreasing V(max) and triggering substrate inhibition by AcCoA. These effects involve centrally the interdomain linker, since we show that linker elongation or two-residue linker shortening hampers and mimics, respectively, arginine inhibition. We propose a regulatory mechanism in which arginine triggers the expansion of the hexameric NAGS ring, altering AAK-GNAT domain interactions, and the modulation by these interactions of GNAT domain functions, explaining arginine regulation. PMID:19084009

  3. PRMT1-mediated arginine methylation controls ATXN2L localization

    International Nuclear Information System (INIS)

    Arginine methylation is a posttranslational modification that is of importance in diverse cellular processes. Recent proteomic mass spectrometry studies reported arginine methylation of ataxin-2-like (ATXN2L), the paralog of ataxin-2, a protein that is implicated in the neurodegenerative disorder spinocerebellar ataxia type 2. Here, we investigated the methylation state of ATXN2L and its significance for ATXN2L localization. We first confirmed that ATXN2L is asymmetrically dimethylated in vivo, and observed that the nuclear localization of ATXN2L is altered under methylation inhibition. We further discovered that ATXN2L associates with the protein arginine-N-methyltransferase 1 (PRMT1). Finally, we showed that neither mutation of the arginine–glycine-rich motifs of ATXN2L nor methylation inhibition alters ATXN2L localization to stress granules, suggesting that methylation of ATXN2L is probably not mandatory. - Highlights: • ATXN2L is asymmetrically dimethylated in vivo. • ATXN2L interacts with PRMT1 under normal and stress conditions. • PRMT1-mediated dimethylation of ATXN2L controls its nuclear localization. • ATXN2L localization to stress granules appears independent of its methylation state

  4. PRMT1-mediated arginine methylation controls ATXN2L localization

    Energy Technology Data Exchange (ETDEWEB)

    Kaehler, Christian; Guenther, Anika; Uhlich, Anja; Krobitsch, Sylvia, E-mail: krobitsc@molgen.mpg.de

    2015-05-15

    Arginine methylation is a posttranslational modification that is of importance in diverse cellular processes. Recent proteomic mass spectrometry studies reported arginine methylation of ataxin-2-like (ATXN2L), the paralog of ataxin-2, a protein that is implicated in the neurodegenerative disorder spinocerebellar ataxia type 2. Here, we investigated the methylation state of ATXN2L and its significance for ATXN2L localization. We first confirmed that ATXN2L is asymmetrically dimethylated in vivo, and observed that the nuclear localization of ATXN2L is altered under methylation inhibition. We further discovered that ATXN2L associates with the protein arginine-N-methyltransferase 1 (PRMT1). Finally, we showed that neither mutation of the arginine–glycine-rich motifs of ATXN2L nor methylation inhibition alters ATXN2L localization to stress granules, suggesting that methylation of ATXN2L is probably not mandatory. - Highlights: • ATXN2L is asymmetrically dimethylated in vivo. • ATXN2L interacts with PRMT1 under normal and stress conditions. • PRMT1-mediated dimethylation of ATXN2L controls its nuclear localization. • ATXN2L localization to stress granules appears independent of its methylation state.

  5. L-Arginine and L-Glutamine as Immunonutrients and Modulating Agents for Erysipelothrix rusiopathiae Infection

    Directory of Open Access Journals (Sweden)

    Haijun Wang

    2012-01-01

    Full Text Available L-arginine and L-glutamine were not only building blocks of proteins and polypeptides but also important regulators of key metabolic pathways that were necessary for maintenance, growth, reproduction and immunity in organisms. These compelling findings convinced us that L-arginine and L-glutamine play a vital role in virus and bacterium infection. However, scientific literature about its role on Erysipelothrix rhusiopathiae (E. rhusiopathiae infection was unavailable. Thus, this study was conduct to research the effect of dietary L-arginine and L-glutamine supplementation on E. rhusiopathiae infection. According to the exciting results, researchers concluded that dietary L-arginine and L-glutamine supplementation ameliorated the cytokines profile and blood parameters and delayed the development process of E. rhusiopathiae infection in mouse model.

  6. Large-Scale Identification of the Arginine Methylome by Mass Spectrometry

    DEFF Research Database (Denmark)

    Sylvestersen, Kathrine B; Nielsen, Michael L

    2016-01-01

    The attachment of one or more methylation groups to the side chain of arginine residues is a regulatory mechanism for cellular proteins. Recent advances in mass spectrometry-based characterization allow comprehensive identification of arginine methylation sites by peptide-level enrichment...... strategies. Described in this unit is a 4-day protocol for enrichment of arginine-methylated peptides and subsequent identification of thousands of distinct sites by mass spectrometry. Specifically, the protocol explains step-by-step sample preparation, enrichment using commercially available antibodies......, prefractionation using strong cation exchange, and identification using liquid chromatography coupled to tandem mass spectrometry. A strategy for relative quantification is described using stable isotope labeling by amino acids in cell culture (SILAC). Approaches for analysis of arginine methylation site occupancy...

  7. Cadmium-induced crystallization of proteins: II. Crystallization of the Salmonella typhimurium histidine-binding protein in complex with L-histidine, L-arginine, or L-lysine.

    Science.gov (United States)

    Trakhanov, S; Kreimer, D I; Parkin, S; Ames, G F; Rupp, B

    1998-03-01

    To further investigate favorable effects of divalent cations on the formation of protein crystals, three complexes of Salmonella typhimurium histidine-binding protein were crystallized with varying concentrations of cadmium salts. For each of the three histidine-binding protein complexes, cadmium cations were found to promote or improve crystallization. The optimal cadmium concentration is ligand specific and falls within a narrow concentration range. In each case, crystals grown in the presence of cadmium diffract to better than 2.0 angstroms resolution and belong to the orthorhombic space group P2(1)2(1)2(1). From our results and from the analysis of cadmium sites in well-refined protein structures, we propose that cadmium addition provides a generally useful technique to modify crystal morphology and to improve diffraction quality. PMID:9541391

  8. The crystal structure of the complex of PII and acetylglutamate kinase reveals how PII controls the storage of nitrogen as arginine

    OpenAIRE

    Llácer, José L.; Contreras, Asunción; Forchhammer, Karl; Marco-Marín, Clara; Gil-Ortiz, Fernando; Maldonado, Rafael; Fita, Ignacio; Rubio, Vicente

    2007-01-01

    Photosynthetic organisms can store nitrogen by synthesizing arginine, and, therefore, feedback inhibition of arginine synthesis must be relieved in these organisms when nitrogen is abundant. This relief is accomplished by the binding of the PII signal transduction protein to acetylglutamate kinase (NAGK), the controlling enzyme of arginine synthesis. Here, we describe the crystal structure of the complex between NAGK and PII of Synechococcus elongatus, at 2.75-Å resolution. We prove the physi...

  9. Cationic amino acid transporters and Salmonella Typhimurium ArgT collectively regulate arginine availability towards intracellular Salmonella growth.

    Directory of Open Access Journals (Sweden)

    Priyanka Das

    Full Text Available Cationic amino acid transporters (mCAT1 and mCAT2B regulate the arginine availability in macrophages. How in the infected cell a pathogen can alter the arginine metabolism of the host remains to be understood. We reveal here a novel mechanism by which Salmonella exploit mCAT1 and mCAT2B to acquire host arginine towards its own intracellular growth within antigen presenting cells. We demonstrate that Salmonella infected bone marrow derived macrophages and dendritic cells show enhanced arginine uptake and increased expression of mCAT1 and mCAT2B. We show that the mCAT1 transporter is in close proximity to Salmonella containing vacuole (SCV specifically by live intracellular Salmonella in order to access the macrophage cytosolic arginine pool. Further, Lysosome associated membrane protein 1, a marker of SCV, also was found to colocalize with mCAT1 in the Salmonella infected cell. The intra vacuolar Salmonella then acquire the host arginine via its own arginine transporter, ArgT for growth. The argT knockout strain was unable to acquire host arginine and was attenuated in growth in both macrophages and in mice model of infection. Together, these data reveal survival strategies by which virulent Salmonella adapt to the harsh conditions prevailing in the infected host cells.

  10. Differential role of arginine mutations on the structure and functions of α-crystallin☆

    Science.gov (United States)

    Panda, Alok Kumar; Nandi, Sandip Kumar; Chakraborty, Ayon; Nagaraj, Ram H.; Biswas, Ashis

    2016-01-01

    Background α-Crystallin is a major protein of the eye lens in vertebrates. It is composed of two subunits, αA- and αB-crystallin. α-Crystallin is an oligomeric protein having these two subunits in 3:1 ratio. It belongs to small heat shock protein family and exhibits molecular chaperone function, which plays an important role in maintaining the lens transparency. Apart from chaperone function, both subunits also exhibit anti-apoptotic property. Comparison of their primary sequences reveals that αA- and αB-crystallin posses 13 and 14 arginine residues, respectively. Several of them undergo mutations which eventually lead to various eye diseases such as congenital cataract, juvenile cataract, and retinal degeneration. Interestingly, many arginine residues of these subunits are modified during glycation and even some are truncated during aging. All these facts indicate the importance of arginine residues in α-crystallin. Scope of review In this review, we will emphasize the recent in vitro and in vivo findings related to congenital cataract causing arginine mutations in α-crystallin. Major conclusions Congenital cataract causing arginine mutations alters the structure and decreases the chaperone function of α-crystallin. These mutations also affect the lens morphology and phenotypes. Interestingly, non-natural arginine mutations (generated for mimicking the glycation and truncation environment) improve the chaperone function of α-crystallin which may play an important role in maintaining the eye lens transparency during aging. General significance The neutralization of positive charge on the guanidino group of arginine residues is not always detrimental to the functionality of α-crystallin. PMID:26080000

  11. Relationship of arginine with lysine in diets for laying Japanese quails

    Directory of Open Access Journals (Sweden)

    Renata de Souza Reis

    2012-01-01

    Full Text Available To determine the relationship of arginine with lysine for Japanese quails during the period of production, an experiment was conducted using 360 subspecies of Japanese quails (Coturnix coturnix japonica with 162 days of age, distributed in a completely randomized design. Diets were formulated with corn, soybean meal, sorghum and wheat bran containing 20.0% crude protein and 2,800 kcal ME/kg. The basal diet contained suboptimal level of lysine equal to 1% and was supplemented with five levels of L-arginine 99% (0.032; 0.083; 0.134; 0.185 and 0.236% to replace the glutamic acid, corresponding to the relationship of arginine with digestible lysine of 1.16, 1.21, 1.26, 1.31 and 1.36. The parameters studied were: feed intake, egg production per hen/day, egg production per hen housed, commercial egg production, egg weight, egg mass, feed conversion by egg mass, feed conversion per dozen eggs, weight and percentage of components of the eggs (yolk, albumen and shell and specific gravity. There was no significant effect on the relationship of arginine with digestible lysine in the diet of Japanese quails for any of the parameters examined. The arginine/lysine ratio of 1.16, which corresponds to a daily intake of 288.84 mg of arginine, provides satisfactory performance and egg quality of Japanese quails.

  12. Genetic and biochemical characterization of arginine biosynthesis in Sinorhizobium meliloti 1021.

    Science.gov (United States)

    Hernández, Victor M; Girard, Lourdes; Hernández-Lucas, Ismael; Vázquez, Alejandra; Ortíz-Ortíz, Catalina; Díaz, Rafael; Dunn, Michael F

    2015-08-01

    L-Ornithine production in the alfalfa microsymbiont Sinorhizobium meliloti occurs as an intermediate step in arginine biosynthesis. Ornithine is required for effective symbiosis but its synthesis in S. meliloti has been little studied. Unlike most bacteria, S. meliloti 1021 is annotated as encoding two enzymes producing ornithine: N-acetylornithine (NAO) deacetylase (ArgE) hydrolyses NAO to acetate and ornithine, and glutamate N-acetyltransferase (ArgJ) transacetylates l-glutamate with the acetyl group from NAO, forming ornithine and N-acetylglutamate (NAG). NAG is the substrate for the second step of arginine biosynthesis catalysed by NAG kinase (ArgB). Inactivation of argB in strain 1021 resulted in arginine auxotrophy. The activity of purified ArgB was significantly inhibited by arginine but not by ornithine. The purified ArgJ was highly active in NAO deacetylation/glutamate transacetylation and was significantly inhibited by ornithine but not by arginine. The purified ArgE protein (with a 6His-Sumo affinity tag) was also active in deacetylating NAO. argE and argJ single mutants, and an argEJ double mutant, are arginine prototrophs. Extracts of the double mutant contained aminoacylase (Ama) activity that deacetylated NAO to form ornithine. The purified products of three candidate ama genes (smc00682 (hipO1), smc02256 (hipO2) and smb21279) all possessed NAO deacetylase activity. hipO1 and hipO2, but not smb21279, expressed in trans functionally complemented an Escherichia coli ΔargE : : Km mutant. We conclude that Ama activity accounts for the arginine prototrophy of the argEJ mutant. Transcriptional assays of argB, argE and argJ, fused to a promoterless gusA gene, showed that their expression was not significantly affected by exogenous arginine or ornithine. PMID:26271664

  13. Arginine regulation of gramicidin S biosynthesis.

    OpenAIRE

    Poirier, A.; Demain, A L

    1981-01-01

    Several amino acids are known to affect the gramicidin S producer Bacillus brevis ATCC 9999 with respect ot growth, soluble gramicidin S synthetase formation, antibiotic production, or a combination of these. Our studies confirmed that arginine has paradoxical effects on the B. brevis fermentation; it markedly increased growth and antibiotic production, yet decreased the soluble heavy gramicidin S synthetase activity. We found that arginine did not repress heavy gramicidin S synthetase. The a...

  14. Intravenous Selenium Modulates L-Arginine-Induced Experimental Acute Pancreatitis

    Directory of Open Access Journals (Sweden)

    Jonathan Hardman

    2005-09-01

    Full Text Available Context Oxidative stress is understood to have a critical role in the development of acinar injury in experimental acute pancreatitis. We have previously demonstrated that compound multiple antioxidant therapy ameliorates end-organ damage in the intra-peritoneal L-arginine rat model. As the principal co-factor for glutathione, selenium is a key constituent of multiple antioxidant preparations. Objective The intention of this study was to investigate the effect of selenium on pancreatic and remote organ injury in a wellvalidated experimental model of acute pancreatitis. Methods Male Sprague-Dawley rats were randomly allocated to one of 3 groups (n=5/group and sacrificed at 72 hours. Acute pancreatitis was induced by 250 mg per 100 g body weight of 20% L-arginine hydrochloride in 0.15 mol/L sodium chloride. Group allocations were: Group 1, control; Group 2, acute pancreatitis; Group 3, selenium. Main outcome measures Serum amylase, anti-oxidant levels, bronchoalveolar lavage protein, lung myeloperoxidase activity, and histological assessment of pancreatic injury. Results L-arginine induced acute pancreatitis characterised by oedema, neutrophil infiltration, acinar cell degranulation and elevated serum amylase. Selenium treatment was associated with reduced pancreatic oedema and inflammatory cell infiltration. Acinar degranulation and dilatation were completely absent. A reduction in bronchoalveolar lavage protein content was also demonstrated. Conclusion Intravenous selenium given 24 hours after induction of experimental acute pancreatitis was associated with a reduction in the histological stigmata of pancreatic injury and a dramatic reduction in broncho-alveolar lavage protein content. Serum selenium fell during the course of experimental acute pancreatitis and this effect was not reversed by exogenous selenium supplementation.

  15. Proteomic analysis of arginine methylation sites in human cells reveals dynamic regulation during transcriptional arrest

    DEFF Research Database (Denmark)

    Sylvestersen, Kathrine B; Horn, Heiko; Jungmichel, Stephanie;

    2014-01-01

    contain regulated functions on their own. Collectively, we present a site-specific MMA dataset in human cells and demonstrate for the first time that MMA is a dynamic post-translational modification regulated during transcriptional arrest by a hitherto uncharacterized arginine demethylase....... mono-methylation (MMA) sites. We thereby identify 1,027 site-specific MMA sites on 494 human proteins, discovering numerous novel mono-methylation targets and confirming the majority of currently known MMA substrates. Nuclear RNA-binding proteins involved in RNA processing, RNA localization......, transcription, and chromatin remodeling are predominantly found modified with MMA. Despite this, MMA sites prominently are located outside RNA-binding domains as compared to the proteome-wide distribution of arginine residues. Quantification of arginine methylation in cells treated with Actinomycin D uncovers...

  16. Remodeling and Control of Homologous Recombination by DNA Helicases and Translocases that Target Recombinases and Synapsis.

    Science.gov (United States)

    Northall, Sarah J; Ivančić-Baće, Ivana; Soultanas, Panos; Bolt, Edward L

    2016-01-01

    Recombinase enzymes catalyse invasion of single-stranded DNA (ssDNA) into homologous duplex DNA forming "Displacement loops" (D-loops), a process called synapsis. This triggers homologous recombination (HR), which can follow several possible paths to underpin DNA repair and restart of blocked and collapsed DNA replication forks. Therefore, synapsis can be a checkpoint for controlling whether or not, how far, and by which pathway, HR proceeds to overcome an obstacle or break in a replication fork. Synapsis can be antagonized by limiting access of a recombinase to ssDNA and by dissociation of D-loops or heteroduplex formed by synapsis. Antagonists include DNA helicases and translocases that are identifiable in eukaryotes, bacteria and archaea, and which target synaptic and pre-synaptic DNA structures thereby controlling HR at early stages. Here we survey these events with emphasis on enabling DNA replication to be resumed from sites of blockage or collapse. We also note how knowledge of anti-recombination activities could be useful to improve efficiency of CRISPR-based genome editing. PMID:27548227

  17. Detection of a novel arginine vasopression defect by dideoxy fingerprinting

    Energy Technology Data Exchange (ETDEWEB)

    Krishnamani, M.R.S.; Phillips, J.A. III; Copeland, K.C. (Vanderbilt Univ. School of Medicine, Nashville, TN (United States) Univ. of Vermont College of Medicine, Burlington, VT (United States))

    1993-09-01

    Autosomal dominant neurohypophyseal diabetes insipidus is a familial form of diabetes insipidus. This disorder is associated with variable levels of arginine vasopressin (AVP) and diabetes insipidus of varying severity, which responds to exogenous AVP. To determine the molecular basis of autosomal dominant neurohypophyseal diabetes insipidus, the AVP genes of members of a large kindred were analyzed. A new method, called dideoxy fingerprinting, was used to detect an AVP mutation that was characterized by DNA sequencing. The novel defect found changes the last codon of the AVP signal peptide from alanine to threonine, which should perturb cleavage of mature AVP from its precursor protein and inhibit its secretion or action. 18 refs., 3 figs.

  18. Rosiglitazone increases fatty acid oxidation and fatty acid translocase (FAT/CD36) but not carnitine palmitoyltransferase I in rat muscle mitochondria

    Science.gov (United States)

    Benton, Carley R; Holloway, Graham P; Campbell, S E; Yoshida, Yuko; Tandon, Narendra N; Glatz, Jan F C; Luiken, Joost J J F P; Spriet, Lawrence L; Bonen, Arend

    2008-01-01

    Peroxisome proliferator-activated receptors (PPARs) alter the expression of genes involved in regulating lipid metabolism. Rosiglitazone, a PPARγ agonist, induces tissue-specific effects on lipid metabolism; however, its mode of action in skeletal muscle remains unclear. Since fatty acid translocase (FAT/CD36) was recently identified as a possible regulator of skeletal muscle fatty acid transport and mitochondrial fatty acid oxidation, we examined in this tissue the effects of rosiglitazone infusion (7 days, 1 mg day−1) on FAT/CD36 mRNA and protein, its plasmalemmal content and fatty acid transport. In addition, in isolated subsarcolemmal (SS) and intermyofibrillar (IMF) mitochondria we examined rates of fatty acid oxidation, FAT/CD36 and carnitine palmitoyltransferase I (CPTI) protein, and CPTI and β-hydroxyacyl CoA dehydrogenase (β-HAD) activities. Rosiglitazone did not alter FAT/CD36 mRNA or protein expression, FAT/CD36 plasmalemmal content, or the rate of fatty acid transport into muscle (P > 0.05). In contrast, rosiglitazone increased the rates of fatty acid oxidation in both SS (+21%) and IMF mitochondria (+36%). This was accompanied by concomitant increases in FAT/CD36 in subsarcolemmal (SS) (+43%) and intermyofibrillar (IMF) mitochondria (+46%), while SS and IMF CPTI protein content, and CPTI submaximal and maximal activities (P > 0.05) were not altered. Similarly, citrate synthase (CS) and β-HAD activities were also not altered by rosiglitazone in SS and IMF mitochondria (P > 0.05). These studies provide another example whereby changes in mitochondrial fatty oxidation are associated with concomitant changes in mitochondrial FAT/CD36 independent of any changes in CPTI. Moreover, these studies identify for the first time a mechanism by which rosiglitazone stimulates fatty acid oxidation in skeletal muscle, namely the chronic, subcellular relocation of FAT/CD36 to mitochondria. PMID:18238811

  19. Selection of Arginine-Rich Anti-Gold Antibodies Engineered for Plasmonic Colloid Self-Assembly

    CERN Document Server

    Jain, Purvi; Narayanan, S Shankara; Sharma, Jadab; Girard, Christian; Dujardin, Erik; Nizak, Clément

    2014-01-01

    Antibodies are affinity proteins with a wide spectrum of applications in analytical and therapeutic biology. Proteins showing specific recognition for a chosen molecular target can be isolated and their encoding sequence identified in vitro from a large and diverse library by phage display selection. In this work, we show that this standard biochemical technique rapidly yields a collection of antibody protein binders for an inorganic target of major technological importance: crystalline metallic gold surfaces. 21 distinct anti-gold antibody proteins emerged from a large random library of antibodies and were sequenced. The systematic statistical analysis of all the protein sequences reveals a strong occurrence of arginine in anti-gold antibodies, which corroborates recent molecular dynamics predictions on the crucial role of arginine in protein/gold interactions. Once tethered to small gold nanoparticles using histidine tag chemistry, the selected antibodies could drive the self-assembly of the colloids onto t...

  20. Arginine Adjunctive Therapy in Active Tuberculosis

    Directory of Open Access Journals (Sweden)

    Aliasghar Farazi

    2015-01-01

    Full Text Available Background. Dietary supplementation has been used as a mechanism to augment the immune system. Adjunctive therapy with L-arginine has the potential to improve outcomes in active tuberculosis. Methods. In a randomized clinical trial 63 participants with smear-positive pulmonary tuberculosis in Markazi Province of Iran were given arginine or placebo for 4 weeks in addition to conventional chemotherapy. The final treatment success, sputum conversion, weight gain, and clinical symptoms after one and two months were considered as primary outcomes and secondary outcomes were ESR, CRP, and Hg. Data were collected and analyzed with SPSS software (ver. 18. Results. Arginine supplementation reduced constitutional symptoms (P=0.032 in patients with smear-positive TB at the end of the first month of treatment. Arginine treated patients had significantly increased BMI at the end of the first and second months of treatment (P=0.032 and P=0.04 and a reduced CRP at the end of the first month of treatment (P=0.03 versus placebo group. Conclusion. Arginine is useful as an adjunctive therapy in patients with active tuberculosis, in which the effects are more likely mediated by the increased production of nitric oxide and improved constitutional symptoms and weight gain. This trial is registered with Clinical Trials Registry of Iran: IRCT201211179855N2.

  1. Arginase and Arginine Dysregulation in Asthma

    Directory of Open Access Journals (Sweden)

    Renée C. Benson

    2011-01-01

    Full Text Available In recent years, evidence has accumulated indicating that the enzyme arginase, which converts L-arginine into L-ornithine and urea, plays a key role in the pathogenesis of pulmonary disorders such as asthma through dysregulation of L-arginine metabolism and modulation of nitric oxide (NO homeostasis. Allergic asthma is characterized by airway hyperresponsiveness, inflammation, and remodeling. Through substrate competition, arginase decreases bioavailability of L-arginine for nitric oxide synthase (NOS, thereby limiting NO production with subsequent effects on airway tone and inflammation. By decreasing L-arginine bioavailability, arginase may also contribute to the uncoupling of NOS and the formation of the proinflammatory oxidant peroxynitrite in the airways. Finally, arginase may play a role in the development of chronic airway remodeling through formation of L-ornithine with downstream production of polyamines and L-proline, which are involved in processes of cellular proliferation and collagen deposition. Further research on modulation of arginase activity and L-arginine bioavailability may reveal promising novel therapeutic strategies for asthma.

  2. Arginine, scurvy and Cartier's "tree of life"

    Directory of Open Access Journals (Sweden)

    Durzan Don J

    2009-02-01

    Full Text Available Abstract Several conifers have been considered as candidates for "Annedda", which was the source for a miraculous cure for scurvy in Jacques Cartier's critically ill crew in 1536. Vitamin C was responsible for the cure of scurvy and was obtained as an Iroquois decoction from the bark and leaves from this "tree of life", now commonly referred to as arborvitae. Based on seasonal and diurnal amino acid analyses of candidate "trees of life", high levels of arginine, proline, and guanidino compounds were also probably present in decoctions prepared in the severe winter. The semi-essential arginine, proline and all the essential amino acids, would have provided additional nutritional benefits for the rapid recovery from scurvy by vitamin C when food supply was limited. The value of arginine, especially in the recovery of the critically ill sailors, is postulated as a source of nitric oxide, and the arginine-derived guanidino compounds as controlling factors for the activities of different nitric oxide synthases. This review provides further insights into the use of the candidate "trees of life" by indigenous peoples in eastern Canada. It raises hypotheses on the nutritional and synergistic roles of arginine, its metabolites, and other biofactors complementing the role of vitamin C especially in treating Cartier's critically ill sailors.

  3. [Antioxidant effects of L-arginine in the rat heart in experimental rhabdomyolysis].

    Science.gov (United States)

    Filimonenko, V P; Nikitchenko, I V; Kaliman, P A

    2009-01-01

    The glycerol administration in a dose of 1 ml of 50% water solution/100 g b. w. was found to cause considerable accumulation of the total heme in the rat blood serum that is accompanied by an increase of TBA-reactive products and protein carbonyl derivates contents and by changes of protein level. Heme entering in the heart tissue is observed in the first hours after glycerol injection. The breaches of heart antioxidant-prooxidant balance are noted in twenty-four hours: TBA-reactive products and protein carbonyl derivates accumulation, heme oxygenase and catalase activation, superoxide dismutase activity lowering and reduction of glutathione content elevation. Pretreatment by L-arginine (0.5 h before glycerol administration) almost did not affect the blood serum changes caused by glycerol injection. However in the rat heart L-arginine administration prevents from TBA-reactive products and protein carbonyl derivates accumulation and the breaches of superoxide dismutase and catalase activities. Besides L-arginine causes the ealier heme oxygenase induction. Possible mechanisms of L-arginine protective action in the rat heart under experimental rhabdomyolysis are discussed. PMID:19877424

  4. Inversion of allosteric effect of arginine on N-acetylglutamate synthase, a molecular marker for evolution of tetrapods

    Directory of Open Access Journals (Sweden)

    Cabrera-Luque Juan

    2008-09-01

    Full Text Available Abstract Background The efficient conversion of ammonia, a potent neurotoxin, into non-toxic metabolites was an essential adaptation that allowed animals to move from the aquatic to terrestrial biosphere. The urea cycle converts ammonia into urea in mammals, amphibians, turtles, snails, worms and many aquatic animals and requires N-acetylglutamate (NAG, an essential allosteric activator of carbamylphosphate synthetase I (CPSI in mammals and amphibians, and carbamylphosphate synthetase III (CPSIII in fish and invertebrates. NAG-dependent CPSI and CPSIII catalyze the formation of carbamylphosphate in the first and rate limiting step of ureagenesis. NAG is produced enzymatically by N-acetylglutamate synthase (NAGS, which is also found in bacteria and plants as the first enzyme of arginine biosynthesis. Arginine is an allosteric inhibitor of microbial and plant NAGS, and allosteric activator of mammalian NAGS. Results Information from mutagenesis studies of E. coli and P. aeruginosa NAGS was combined with structural information from the related bacterial N-acetylglutamate kinases to identify four residues in mammalian NAGS that interact with arginine. Substitutions of these four residues were engineered in mouse NAGS and into the vertebrate-like N-acetylglutamate synthase-kinase (NAGS-K of Xanthomonas campestris, which is inhibited by arginine. All mutations resulted in arginine losing the ability to activate mouse NAGS, and inhibit X. campestris NAGS-K. To examine at what point in evolution inversion of arginine effect on NAGS occur, we cloned NAGS from fish and frogs and examined the arginine response of their corresponding proteins. Fish NAGS were partially inhibited by arginine and frog NAGS were activated by arginine. Conclusion Difference in arginine effect on bacterial and mammalian NAGS most likely stems from the difference in the type of conformational change triggered by arginine binding to these proteins. The change from arginine

  5. Atualização da proteína ideal para frangos de corte: arginina, isoleucina, valina e triptofano Updating of the ideal protein for broilers: arginine, isoleucine, valine and tryptophan

    Directory of Open Access Journals (Sweden)

    Anastácia Maria de Araújo Campos

    2012-02-01

    69, 76 e 18%, respectivamente.Two experiments were carried out to evaluate digestible arginine:lysine, isoleucine:lysine, valine:lysine and tryptophan:lysine ratios for male broilers in two periods: 7 - 21 (starter and 28 - 40 (finisher days of age. A total of 1800 starter and 1440 finisher broilers were distributed in a completely randomized experimental design with 9 treatments, eight replicates of 25 and 20 birds per experimental unit in the starter and the finisher periods, respectively. The ratios used in the starter period were: 100, 105 and 110 arginine:lysine; 60, 65 and 70 isoleucine:lysine; 70, 75 and 80 valine:lysine; 15, 16 and 17 tryptophan:lysine, and in the finisher: 95, 105 and 115 arginine:lysine; 58, 67 and 76 isoleucine:lysine; 71,5, 77 and 82,5 valine:lysine; 14, 17 and 20 tryptophan:lysine. Diets were formulated to meet or exceed the nutritional requirements in both periods, except for digestible lysine (1.08% for the starter period and 0.98% for the finisher. At the end of each experiment, weight gain, feed intake, feed conversion and breast and breast fillet weight and yield were determined. In the starter period, the arginine:lysine and tryptophan:lysine ratios did not affect the evaluated parameters, but there were linear effects of isoleucine:lysine and valine:lysine ratios on birds weight gain and feed conversion. In the finisher period, the arginine:lysine ratios influenced linearlly weight gain and feed conversion. There was a quadratic effect of isoleucine:lysine, valine:lysine, and tryptophan:lysine on weight gain. The increase in the isoleucine:lysine and valine:lysine ratios results in better performance of the broilers from 7 to 21 days of age. The recommended isoleucine:lysine, valine:lysine and tryptophan:lysine ratios for broilers from 28 to 40 days of age are 69, 76 and 18%, respectively.

  6. Mechanism of Allosteric Inhibition of N-Acetyl-L-glutamate Synthase by L-Arginine

    Energy Technology Data Exchange (ETDEWEB)

    Min, Li; Jin, Zhongmin; Caldovic, Ljubica; Morizono, Hiroki; Allewell, Norma M.; Tuchman, Mendel; Shi, Dashuang (GUW); (Maryland); (GWU); (Georgia)

    2010-01-07

    N-Acetylglutamate synthase (NAGS) catalyzes the first committed step in L-arginine biosynthesis in plants and micro-organisms and is subject to feedback inhibition by L-arginine. This study compares the crystal structures of NAGS from Neisseria gonorrhoeae (ngNAGS) in the inactive T-state with L-arginine bound and in the active R-state complexed with CoA and L-glutamate. Under all of the conditions examined, the enzyme consists of two stacked trimers. Each monomer has two domains: an amino acid kinase (AAK) domain with an AAK-like fold but lacking kinase activity and an N-acetyltransferase (NAT) domain homologous to other GCN5-related transferases. Binding of L-arginine to the AAK domain induces a global conformational change that increases the diameter of the hexamer by {approx}10 {angstrom} and decreases its height by {approx}20{angstrom}. AAK dimers move 5{angstrom} outward along their 2-fold axes, and their tilt relative to the plane of the hexamer decreases by {approx}4{sup o}. The NAT domains rotate {approx}109{sup o} relative to AAK domains enabling new interdomain interactions. Interactions between AAK and NAT domains on different subunits also change. Local motions of several loops at the L-arginine-binding site enable the protein to close around the bound ligand, whereas several loops at the NAT active site become disordered, markedly reducing enzymatic specific activity.

  7. Mechanism of allosteric inhibition of N-acetyl-L-glutamate synthase by L-arginine.

    Science.gov (United States)

    Min, Li; Jin, Zhongmin; Caldovic, Ljubica; Morizono, Hiroki; Allewell, Norma M; Tuchman, Mendel; Shi, Dashuang

    2009-02-20

    N-Acetylglutamate synthase (NAGS) catalyzes the first committed step in l-arginine biosynthesis in plants and micro-organisms and is subject to feedback inhibition by l-arginine. This study compares the crystal structures of NAGS from Neisseria gonorrhoeae (ngNAGS) in the inactive T-state with l-arginine bound and in the active R-state complexed with CoA and l-glutamate. Under all of the conditions examined, the enzyme consists of two stacked trimers. Each monomer has two domains: an amino acid kinase (AAK) domain with an AAK-like fold but lacking kinase activity and an N-acetyltransferase (NAT) domain homologous to other GCN5-related transferases. Binding of l-arginine to the AAK domain induces a global conformational change that increases the diameter of the hexamer by approximately 10 A and decreases its height by approximately 20A(.) AAK dimers move 5A outward along their 2-fold axes, and their tilt relative to the plane of the hexamer decreases by approximately 4 degrees . The NAT domains rotate approximately 109 degrees relative to AAK domains enabling new interdomain interactions. Interactions between AAK and NAT domains on different subunits also change. Local motions of several loops at the l-arginine-binding site enable the protein to close around the bound ligand, whereas several loops at the NAT active site become disordered, markedly reducing enzymatic specific activity. PMID:19095660

  8. L-Arginine Intake Effect on Adenine Nucleotide Metabolism in Rat Parenchymal and Reproductive Tissues

    Directory of Open Access Journals (Sweden)

    G. Kocic

    2012-01-01

    Full Text Available L-arginine is conditionally essetcial amino acid, required for normal cell growth, protein synthesis, ammonia detoxification, tissue growth and general performance, proposed in the treatment of men sterility and prevention of male impotence. The aim of the present paper was to estimate the activity of the enzymes of adenine nucleotide metabolism: 5′-nucleotidase (5′-NU, adenosine deaminase (ADA, AMP deaminase, and xanthine oxidase (XO, during dietary intake of L-arginine for a period of four weeks of male Wistar rats. Adenosine concentration in tissues is maintained by the relative activities of the adenosine-producing enzyme, 5′-NU and the adenosine-degrading enzyme-ADA adenosine deaminase. Dietary L-arginine intake directed adenine nucleotide metabolism in liver, kidney, and testis tissue toward the activation of adenosine production, by increased 5′-NU activity and decreased ADA activity. Stimulation of adenosine accumulation could be of importance in mediating arginine antiatherosclerotic, vasoactive, immunomodulatory, and antioxidant effects. Assuming that the XO activity reflects the rate of purine catabolism in the cell, while the activity of AMP deaminase is of importance in ATP regeneration, reduced activity of XO, together with the increased AMP-deaminase activity, may suggest that adenine nucleotides are presumably directed to the ATP regenerating process during dietary L-arginine intake.

  9. Multiple proteins with essential mitochondrial functions have glycosylated isoforms

    OpenAIRE

    Burnham-Marusich, Amanda R.; Berninsone, Patricia M.

    2012-01-01

    Nucleocytosolic and secreted proteins are commonly glycosylated. However, reports of glycosylated mitochondrial proteins are rare. Using lectin chromatography on bovine heart, we detected low-abundance glycoforms of nuclear-encoded proteins with well-established mitochondrial function: pyruvate dehydrogenase E1α, NADH dehydrogenase [ubiquinone] iron-sulfur protein 3, ADP/ATP translocase, ATP synthase d and oligomycin sensitivity-conferring protein. Notably, the latter two have been previously...

  10. Role of membrane contact sites in protein import into mitochondria

    OpenAIRE

    Horvath, Susanne E.; Rampelt, Heike; Oeljeklaus, Silke; Warscheid, Bettina; van der Laan, Martin; Pfanner, Nikolaus

    2015-01-01

    Mitochondria import more than 1,000 different proteins from the cytosol. The proteins are synthesized as precursors on cytosolic ribosomes and are translocated by protein transport machineries of the mitochondrial membranes. Five main pathways for protein import into mitochondria have been identified. Most pathways use the translocase of the outer mitochondrial membrane (TOM) as the entry gate into mitochondria. Depending on specific signals contained in the precursors, the proteins are subse...

  11. Three ways in, one way out: water dynamics in the trans-membrane domains of the inner membrane translocase AcrB.

    Science.gov (United States)

    Fischer, Nadine; Kandt, Christian

    2011-10-01

    Powered by proton-motive force, the inner membrane translocase AcrB is the engine of the AcrAB-TolC efflux pump in Escherichia coli. As proton conduction in proteins occurs along hydrogen-bonded networks of polar residues and water molecules, knowledge of the protein-internal water distribution and water-interacting residues allows drawing conclusions to possible pathways of proton conduction. Here, we report a series of 6× 50 ns independent molecular dynamics simulations of asymmetric AcrB embedded in a phospholipid/water environment. Simulating each monomer in its proposed protonation state, we calculated for each trans-membrane domain the average water distribution, identified residues interacting with these waters and quantified each residue's frequency of water hydrogen bond contact. Combining this information we find three possible routes of proton transfer connecting a continuously hydrated region of known key residues in the TMD interior to bulk water by one cytoplasmic and up to three periplasm water channels in monomer B and A. We find that water access of the trans-membrane domains is regulated by four groups of residues in a combination of side chain re-orientations and shifts of trans-membrane helices. Our findings support a proton release event via Arg971 during the C intermediate or in the transition to A, and proton uptake occurring in the A or B state or during a so far unknown intermediate in between B and C where cytoplasmic water access is still possible. Our simulations suggest experimentally testable hypotheses, which have not been investigated so far. PMID:21905112

  12. Low plasma arginine:asymmetric dimethyl arginine ratios predict mortality after intracranial aneurysm rupture

    DEFF Research Database (Denmark)

    Staalsø, Jonatan Myrup; Bergström, Anita; Edsen, Troels; Weikop, Pia; Romner, Bertil; Olsen, Niels Vidiendal

    2013-01-01

    Asymmetrical dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide synthases, predicts mortality in cardiovascular disease and has been linked to cerebral vasospasm after aneurysmal subarachnoid hemorrhage (SAH). In this prospective study, we assessed whether circulating ADMA, arginine...

  13. Modulators of arginine metabolism support cancer immunosurveillance

    Directory of Open Access Journals (Sweden)

    Freschi Massimo

    2009-01-01

    Full Text Available Abstract Background Tumor-associated accrual of myeloid derived suppressor cells (MDSC in the blood, lymphoid organs and tumor tissues may lead to perturbation of the arginine metabolism and impairment of the endogenous antitumor immunity. The objective of this study was to evaluate whether accumulation of MDSC occurred in Th2 prone BALB/c and Th1 biased C57BL/6 mice bearing the C26GM colon carcinoma and RMA T lymphoma, respectively, and to investigate whether N(G nitro-L-arginine methyl ester (L-NAME and sildenafil, both modulators of the arginine metabolism, restored antitumor immunity. Results We report here that MDSC accumulate in the spleen and blood of mice irrespective of the mouse and tumor model used. Treatment of tumor-bearing mice with either the phosphodiesterase-5 inhibitor sildenafil or the nitric-oxide synthase (NOS inhibitor L-NAME significantly restrained tumor growth and expanded the tumor-specific immune response. Conclusion Our data emphasize the role of MDSC in modulating the endogenous tumor-specific immune response and underline the anti-neoplastic therapeutic potential of arginine metabolism modulators.

  14. Arginine-Rich Cationic Polypeptides Amplify Lipopolysaccharide-Induced Monocyte Activation

    OpenAIRE

    Bosshart, Herbert; Heinzelmann, Michael

    2002-01-01

    The human neutrophil-derived cationic protein CAP37, also known as azurocidin or heparin-binding protein, enhances the lipopolysaccharide (LPS)-induced release of tumor necrosis factor alpha (TNF-α) in isolated human monocytes. We measured the release of the proinflammatory cytokine interleukin-8 (IL-8) in human whole blood and found that in addition to CAP37, other arginine-rich cationic polypeptides, such as the small structurally related protamines, enhance LPS-induced monocyte activation....

  15. Dual role of arginine metabolism in establishing pathogenesis.

    Science.gov (United States)

    Gogoi, Mayuri; Datey, Akshay; Wilson, Keith T; Chakravortty, Dipshikha

    2016-02-01

    Arginine is an integral part of host defense when invading pathogens are encountered. The arginine metabolite nitric oxide (NO) confers antimicrobial properties, whereas the metabolite ornithine is utilized for polyamine synthesis. Polyamines are crucial to tissue repair and anti-inflammatory responses. iNOS/arginase balance can determine Th1/Th2 response. Furthermore, the host arginine pool and its metabolites are utilized as energy sources by various pathogens. Apart from its role as an immune modulator, recent studies have also highlighted the therapeutic effects of arginine. This article sheds light upon the roles of arginine metabolism during pathological conditions and its therapeutic potential. PMID:26610300

  16. Mass spectrometry-based identification and characterisation of lysine and arginine methylation in the human proteome.

    Science.gov (United States)

    Bremang, Michael; Cuomo, Alessandro; Agresta, Anna Maria; Stugiewicz, Magdalena; Spadotto, Valeria; Bonaldi, Tiziana

    2013-09-01

    Protein methylation is a post-translational modification (PTM) by which a variable number of methyl groups are transferred to lysine and arginine residues within proteins. Despite increased interest in this modification due to its reversible nature and its emerging role in a diverse set of biological pathways beyond chromatin, global identification of protein methylation has remained an unachieved goal. To characterise sites of lysine and arginine methylation beyond histones, we employed an approach that combines heavy methyl stable isotope labelling by amino acids in cell culture (hmSILAC) with high-resolution mass spectrometry-based proteomics. Through a broad evaluation of immuno-affinity enrichment and the application of two classical protein separation techniques prior to mass spectrometry, to nucleosolic and cytosolic fractions separately, we identified a total of 501 different methylation types, on 397 distinct lysine and arginine sites, present on 139 unique proteins. Our results considerably extend the number of known in vivo methylation sites and indicate their significant presence on several protein complexes involved at all stages of gene expression, from chromatin remodelling and transcription to splicing and translation. In addition, we describe the potential of the hmSILAC approach for accurate relative quantification of methylation levels between distinct functional states. PMID:23748837

  17. Structural bases of feed-back control of arginine biosynthesis, revealed by the structures of two hexameric N-acetylglutamate kinases, from Thermotoga maritima and Pseudomonas aeruginosa.

    Science.gov (United States)

    Ramón-Maiques, Santiago; Fernández-Murga, María Leonor; Gil-Ortiz, Fernando; Vagin, Alexei; Fita, Ignacio; Rubio, Vicente

    2006-02-24

    N-Acetylglutamate kinase (NAGK) catalyses the second step in the route of arginine biosynthesis. In many organisms this enzyme is inhibited by the final product of the route, arginine, and thus plays a central regulatory role. In addition, in photosynthetic organisms NAGK is the target of the nitrogen-signalling protein PII. The 3-D structure of homodimeric, arginine-insensitive, Escherichia coli NAGK, clarified substrate binding and catalysis but shed no light on arginine inhibition of NAGK. We now shed light on arginine inhibition by determining the crystal structures, at 2.75 A and 2.95 A resolution, of arginine-complexed Thermotoga maritima and arginine-free Pseudomonas aeruginosa NAGKs, respectively. Both enzymes are highly similar ring-like hexamers having a central orifice of approximately 30 A diameter. They are formed by linking three E.coli NAGK-like homodimers through the interlacing of an N-terminal mobile kinked alpha-helix, which is absent from E.coli NAGK. Arginine is bound in each subunit of T.maritima NAGK, flanking the interdimeric junction, in a site formed between the N helix and the C lobe of the subunit. This site is also present, in variable conformations, in P.aeruginosa NAGK, but is missing from E.coli NAGK. Arginine, by gluing the C lobe of each subunit to the inter-dimeric junction, may stabilize an enlarged active centre conformation, hampering catalysis. Acetylglutamate counters arginine inhibition by promoting active centre closure. The hexameric architecture justifies the observed sigmoidal arginine inhibition kinetics with a high Hill coefficient (N approximately 4), and appears essential for arginine inhibition and for NAGK-PII complex formation, since this complex may involve binding of NAGK and PII with their 3-fold axes aligned. The NAGK structures allow identification of diagnostic sequence signatures for arginine inhibition. These signatures are found also in the homologous arginine-inhibited enzyme NAG synthase. The findings

  18. Rotations of the 2B Sub-domain of E. coli UvrD Helicase/Translocase Coupled to Nucleotide and DNA Binding

    Energy Technology Data Exchange (ETDEWEB)

    Jia, Haifeng; Korolev, Sergey; Niedziela-Majka, Anita; Maluf, Nasib K.; Gauss, George H.; Myong, Sua; Ha, Taekjip; Waksman, Gabriel; Lohman, Timothy M. (UIUC); (St. Louis-MED); (WU-MED); (UCL)

    2011-11-02

    Escherichia coli UvrD is a superfamily 1 DNA helicase and single-stranded DNA (ssDNA) translocase that functions in DNA repair and plasmid replication and as an anti-recombinase by removing RecA protein from ssDNA. UvrD couples ATP binding and hydrolysis to unwind double-stranded DNA and translocate along ssDNA with 3'-to-5' directionality. Although a UvrD monomer is able to translocate along ssDNA rapidly and processively, DNA helicase activity in vitro requires a minimum of a UvrD dimer. Previous crystal structures of UvrD bound to a ssDNA/duplex DNA junction show that its 2B sub-domain exists in a 'closed' state and interacts with the duplex DNA. Here, we report a crystal structure of an apo form of UvrD in which the 2B sub-domain is in an 'open' state that differs by an {approx} 160{sup o} rotation of the 2B sub-domain. To study the rotational conformational states of the 2B sub-domain in various ligation states, we constructed a series of double-cysteine UvrD mutants and labeled them with fluorophores such that rotation of the 2B sub-domain results in changes in fluorescence resonance energy transfer. These studies show that the open and closed forms can interconvert in solution, with low salt favoring the closed conformation and high salt favoring the open conformation in the absence of DNA. Binding of UvrD to DNA and ATP binding and hydrolysis also affect the rotational conformational state of the 2B sub-domain, suggesting that 2B sub-domain rotation is coupled to the function of this nucleic acid motor enzyme.

  19. Rotations of the 2B sub-domain of E. coli UvrD helicase/translocase coupled to nucleotide and DNA binding.

    Science.gov (United States)

    Jia, Haifeng; Korolev, Sergey; Niedziela-Majka, Anita; Maluf, Nasib K; Gauss, George H; Myong, Sua; Ha, Taekjip; Waksman, Gabriel; Lohman, Timothy M

    2011-08-19

    Escherichia coli UvrD is a superfamily 1 DNA helicase and single-stranded DNA (ssDNA) translocase that functions in DNA repair and plasmid replication and as an anti-recombinase by removing RecA protein from ssDNA. UvrD couples ATP binding and hydrolysis to unwind double-stranded DNA and translocate along ssDNA with 3'-to-5' directionality. Although a UvrD monomer is able to translocate along ssDNA rapidly and processively, DNA helicase activity in vitro requires a minimum of a UvrD dimer. Previous crystal structures of UvrD bound to a ssDNA/duplex DNA junction show that its 2B sub-domain exists in a "closed" state and interacts with the duplex DNA. Here, we report a crystal structure of an apo form of UvrD in which the 2B sub-domain is in an "open" state that differs by an ∼160° rotation of the 2B sub-domain. To study the rotational conformational states of the 2B sub-domain in various ligation states, we constructed a series of double-cysteine UvrD mutants and labeled them with fluorophores such that rotation of the 2B sub-domain results in changes in fluorescence resonance energy transfer. These studies show that the open and closed forms can interconvert in solution, with low salt favoring the closed conformation and high salt favoring the open conformation in the absence of DNA. Binding of UvrD to DNA and ATP binding and hydrolysis also affect the rotational conformational state of the 2B sub-domain, suggesting that 2B sub-domain rotation is coupled to the function of this nucleic acid motor enzyme. PMID:21704638

  20. Noncanonical Myo9b-RhoGAP Accelerates RhoA GTP Hydrolysis by a Dual-Arginine-Finger Mechanism.

    Science.gov (United States)

    Yi, Fengshuang; Kong, Ruirui; Ren, Jinqi; Zhu, Li; Lou, Jizhong; Wu, Jane Y; Feng, Wei

    2016-07-31

    The GTP hydrolysis activities of Rho GTPases are stimulated by GTPase-activating proteins (GAPs), which contain a RhoGAP domain equipped with a characteristic arginine finger and an auxiliary asparagine for catalysis. However, the auxiliary asparagine is missing in the RhoGAP domain of Myo9b (Myo9b-RhoGAP), a unique motorized RhoGAP that specifically targets RhoA for controlling cell motility. Here, we determined the structure of Myo9b-RhoGAP in complex with GDP-bound RhoA and magnesium fluoride. Unexpectedly, Myo9b-RhoGAP contains two arginine fingers at its catalytic site. The first arginine finger resembles the one within the canonical RhoGAP domains and inserts into the nucleotide-binding pocket of RhoA, whereas the second arginine finger anchors the Switch I loop of RhoA and interacts with the nucleotide, stabilizing the transition state of GTP hydrolysis and compensating for the lack of the asparagine. Mutating either of the two arginine fingers impaired the catalytic activity of Myo9b-RhoGAP and affected the Myo9b-mediated cell migration. Our data indicate that Myo9b-RhoGAP accelerates RhoA GTP hydrolysis by a previously unknown dual-arginine-finger mechanism, which may be shared by other noncanonical RhoGAP domains lacking the auxiliary asparagine. PMID:27363609

  1. Arginine, scurvy and Cartier's "tree of life"

    OpenAIRE

    Durzan Don J

    2009-01-01

    Abstract Several conifers have been considered as candidates for "Annedda", which was the source for a miraculous cure for scurvy in Jacques Cartier's critically ill crew in 1536. Vitamin C was responsible for the cure of scurvy and was obtained as an Iroquois decoction from the bark and leaves from this "tree of life", now commonly referred to as arborvitae. Based on seasonal and diurnal amino acid analyses of candidate "trees of life", high levels of arginine, proline, and guanidino compoun...

  2. Protective Effects of Arginine on Saccharomyces cerevisiae Against Ethanol Stress

    Science.gov (United States)

    Cheng, Yanfei; Du, Zhaoli; Zhu, Hui; Guo, Xuena; He, Xiuping

    2016-01-01

    Yeast cells are challenged by various environmental stresses in the process of industrial fermentation. As the currently main organism for bio-ethanol production, Saccharomyces cerevisiae suffers from ethanol stress. Some amino acids have been reported to be related to yeast tolerance to stresses. Here the relationship between arginine and yeast response to ethanol stress was investigated. Marked inhibitions of ethanol on cell growth, expression of genes involved in arginine biosynthesis and intracellular accumulation of arginine were observed. Furthermore, extracellular addition of arginine can abate the ethanol damage largely. To further confirm the protective effects of arginine on yeast cells, yeast strains with different levels of arginine content were constructed by overexpression of ARG4 involved in arginine biosynthesis or CAR1 encoding arginase. Intracellular arginine was increased by 18.9% or 13.1% respectively by overexpression of ARG4 or disruption of CAR1, which enhanced yeast tolerance to ethanol stress. Moreover, a 41.1% decrease of intracellular arginine was observed in CAR1 overexpressing strain, which made yeast cells keenly sensitive to ethanol. Further investigations indicated that arginine protected yeast cells from ethanol damage by maintaining the integrity of cell wall and cytoplasma membrane, stabilizing the morphology and function of organellae due to low ROS generation. PMID:27507154

  3. The twin-arginine signal peptide of Bacillus subtilis YwbN can direct either Tat- or Sec-dependent secretion of different cargo proteins: secretion of active subtilisin via the B. subtilis Tat pathway.

    Science.gov (United States)

    Kolkman, Marc A B; van der Ploeg, René; Bertels, Michael; van Dijk, Maurits; van der Laan, Joop; van Dijl, Jan Maarten; Ferrari, Eugenio

    2008-12-01

    Proteins that are produced for commercial purposes in Bacillus subtilis are commonly secreted via the Sec pathway. Despite its high secretion capacity, the secretion of heterologous proteins via the Sec pathway is often unsuccessful. Alternative secretion routes, like the Tat pathway, are therefore of interest. Two parallel Tat pathways with distinct specificities have previously been discovered in B. subtilis. To explore the application potential of these Tat pathways, several commercially relevant or heterologous model proteins were fused to the signal peptides of the known B. subtilis Tat substrates YwbN and PhoD. Remarkably, the YwbN signal peptide directed secretion of active subtilisin, a typical Sec substrate, via the B. subtilis TatAyCy route. In contrast, the same signal peptide directed Tat-independent secretion of the Bacillus licheniformis alpha-amylase (AmyL). Moreover, the YwbN signal peptide directed secretion of SufI, an Escherichia coli Tat substrate, in a Tat-independent manner, most likely via Sec. Our results suggest that cytoplasmic protein folding prior to translocation is probably a major determinant of Tat-dependent protein secretion in B. subtilis, as is the case with E. coli. We conclude that future applications for the Tat system of B. subtilis will most likely involve commercially interesting proteins that are Sec incompatible. PMID:18931290

  4. [Effect of L-arginine on pro- and antioxidant status of the rat vessels and lungs in experimental rhabdomyolysis].

    Science.gov (United States)

    Fylymonenko, V P; Nikitchenko, I V; Kaliman, P A

    2009-01-01

    The glycerol administration was found to cause accumulation of the total heme in rat blood serum, vessels and lungs that are accompanied by increase of TBA-reactive products and protein carbonyl derivates contents. A decrease of superoxide dismutase activity and an increase of reduced glutathione in lung were observed. Heme entering the vessels and lungs is accompanied by elevation in heme oxygenase activity. Pretreatment by L-arginine (0.5 h before glycerol administration) didn't affect blood serum and vessels changes caused by glycerol injection. However, in lungs, L-arginine prevents TBA-reactive products and protein carbonyl derivates accumulation, the decrease ofsuperoxide dismutase activity and causes the ealier heme oxygenase induction. Prooxidant effects of heme in tissues studied and possible mechanisms of L-arginine protective action in lung under experimental rhabdomyolysis are discussed. PMID:20095386

  5. Dietary supplementation with L-arginine in patients with breast cancer (> 4 cm) receiving multimodality treatment: report of a feasibility study.

    OpenAIRE

    Brittenden, J.; Heys, S D; Miller, I.; Sarkar, T K; Hutcheon, A. W.; Needham, G.; Gilbert, F.; McKean, M.; Ah-See, A. K.; Eremin, O.

    1994-01-01

    L-Arginine has been shown, in human breast cancers, to increase protein synthesis and the number of cells in the growth phase of the cell cycle. L-Arginine, therefore, may potentiate the response of breast cancers to cell cycle-specific cytotoxic agents. This phase II pilot study assessed the clinical, radiological and pathological responses in 44 patients with breast cancers > 4 cm in diameter (46 tumours: T2, n = 6; T3, n = 22; T4, n = 19), who received oral L-arginine 30 g day-1 for 3 days...

  6. Methylation-controlled J-protein MCJ acts in the import of proteins into human mitochondria.

    Science.gov (United States)

    Schusdziarra, Christina; Blamowska, Marta; Azem, Abdussalam; Hell, Kai

    2013-04-01

    Loss of expression of the methylation-controlled J gene, MCJ (DNAJC15), is observed in cases of several tumors and plays a crucial role in the chemoresistance of ovarian cancer cells. Aside from the pathophysiological effects, almost nothing is known about the cellular function of MCJ. Here, we provide the first evidence that MCJ acts in the biogenesis of mitochondria. Our results demonstrate that MCJ is located in mitochondria. It is anchored in the mitochondrial inner membrane with the C-terminal J domain facing the matrix space. We show that MCJ forms a stable subcomplex with a component of the mitochondrial import motor, MAGMAS, a protein overexpressed in cells treated with granulocyte-macrophage colony-stimulating factor and in prostate carcinomas. In addition, MCJ and MAGMAS interact with the core components of the TIM23 pre-protein translocase. We demonstrate that the recombinant soluble MCJ domain stimulates the ATPase activity of the human mtHsp70 chaperone, mortalin, the central component of the import motor of the TIM23 translocase. This stimulation is counteracted by MAGMAS. Moreover, pre-protein import into mitochondria is impaired in the absence of MCJ. Interestingly, MCJ is able to take over the function of Tim14, the essential J co-chaperone of the mitochondrial protein import motor in yeast. In summary, our results show that MCJ functions as J co-chaperone of the human TIM23 pre-protein translocase, suggesting a link between mitochondrial pre-protein import and tumorigenesis. PMID:23263864

  7. Spectrophotometric Determination of Arginine in Grape Juice Using 8-Hydroquinoline

    Institute of Scientific and Technical Information of China (English)

    WANG Hua; LIANG Xin-hong; ZHAO Rui-xiang; FENG Li-dan; LI Hua

    2008-01-01

    Arginine in grape juice can be metabolized by wine yeasts and malolactic bacteria to precursors of ethyl carbamate, known as carcinogen. The aim of this study was to develop a simple, fast, and accurate method for determining arginine in grape juice with Sakaguchi reaction by separating arginine with strong cation-exchange resins. Parameters were optimized including the concentrations of 8-hydroquinoline and sodium hydrobromite. The color stability lasted for 4 min, which is sufficient to finish the measurement. The method is simple, reproducible and accurate, and can be applied for quick measurement of arginine in grape juice to take necessary measures for controlling the level of ethyl carbamate.

  8. Scanning mutagenesis of the I-II loop of the Cav2.2 calcium channel identifies residues Arginine 376 and Valine 416 as molecular determinants of voltage dependent G protein inhibition

    Directory of Open Access Journals (Sweden)

    Tedford Hugo W

    2010-02-01

    Full Text Available Abstract Direct interaction with the β subunit of the heterotrimeric G protein complex causes voltage-dependent inhibition of N-type calcium channels. To further characterize the molecular determinants of this interaction, we performed scanning mutagenesis of residues 372-387 and 410-428 of the N-type channel α1 subunit, in which individual residues were replaced by either alanine or cysteine. We coexpressed wild type Gβ1γ2 subunits with either wild type or point mutant N-type calcium channels, and voltage-dependent, G protein-mediated inhibition of the channels (VDI was assessed using patch clamp recordings. The resulting data indicate that Arg376 and Val416 of the α1 subunit, residues which are surface-exposed in the presence of the calcium channel β subunit, contribute significantly to the functional inhibition by Gβ1. To further characterize the roles of Arg376 and Val416 in this interaction, we performed secondary mutagenesis of these residues, coexpressing the resulting mutants with wild type Gβ1γ2 subunits and with several isoforms of the auxiliary β subunit of the N-type channel, again assessing VDI using patch clamp recordings. The results confirm the importance of Arg376 for G protein-mediated inhibition and show that a single amino acid substitution to phenylalanine drastically alters the abilities of auxiliary calcium channel subunits to regulate G protein inhibition of the channel.

  9. Surprising Arginine Biosynthesis: a Reappraisal of the Enzymology and Evolution of the Pathway in Microorganisms

    OpenAIRE

    Xu, Ying; Labedan, Bernard; Glansdorff, Nicolas

    2007-01-01

    Summary: Major aspects of the pathway of de novo arginine biosynthesis via acetylated intermediates in microorganisms must be revised in light of recent enzymatic and genomic investigations. The enzyme N-acetylglutamate synthase (NAGS), which used to be considered responsible for the first committed step of the pathway, is present in a limited number of bacterial phyla only and is absent from Archaea. In many Bacteria, shorter proteins related to the Gcn5-related N-acetyltransferase family ap...

  10. Methylation of CenH3 arginine 37 regulates kinetochore integrity and chromosome segregation

    OpenAIRE

    Samel, Anke; Cuomo, Alessandro; Bonaldi, Tiziana; Ehrenhofer-Murray, Ann E

    2012-01-01

    Centromeres of eukaryotic chromosomes mark the site for kinetochore formation and microtubule attachment and are essential for accurate chromosome segregation. Although centromere identity is defined by the presence of the histone H3 variant CenH3/centromere protein A (CENP-A), little is known about how epigenetic modifications on CenH3 might regulate kinetochore assembly and centromere function. Here we show that CENP-A from Saccharomyces cerevisiae, termed Cse4, is methylated on arginine 37...

  11. A mutation in the human phospholamban gene, deleting arginine 14, results in lethal, hereditary cardiomyopathy

    OpenAIRE

    Haghighi, Kobra; Kolokathis, Fotis; Gramolini, Anthony O.; Waggoner, Jason R.; Pater, Luke; Lynch, Roy A.; Fan, Guo-Chang; Tsiapras, Dimitris; Parekh, Rohan R.; Dorn, Gerald W., II; MacLennan, David H.; Kremastinos, Dimitrios Th; Kranias, Evangelia G.

    2006-01-01

    The sarcoplasmic reticulum Ca2+-cycling proteins are key regulators of cardiac contractility, and alterations in sarcoplasmic reticulum Ca2+-cycling properties have been shown to be causal of familial cardiomyopathies. Through genetic screening of dilated cardiomyopathy patients, we identified a previously uncharacterized deletion of arginine 14 (PLN-R14Del) in the coding region of the phospholamban (PLN) gene in a large family with hereditary heart failure. No homozygous individuals were ide...

  12. Arginine ADP-ribosylation mechanism based on structural snapshots of iota-toxin and actin complex

    OpenAIRE

    Tsurumura, Toshiharu; Tsumori, Yayoi; Qiu, Hao; Oda, Masataka; Sakurai, Jun; Nagahama, Masahiro; Tsuge, Hideaki

    2012-01-01

    Clostridium perfringens iota-toxin (Ia) mono-ADP ribosylates Arg177 of actin, leading to cytoskeletal disorganization and cell death. To fully understand the reaction mechanism of arginine-specific mono-ADP ribosyl transferase, the structure of the toxin-substrate protein complex must be characterized. Recently, we solved the crystal structure of Ia in complex with actin and the nonhydrolyzable NAD+ analog βTAD (thiazole-4-carboxamide adenine dinucleotide); however, the structures of the NAD+...

  13. Mitochondrial long chain fatty acid oxidation, fatty acid translocase/CD36 content and carnitine palmitoyltransferase I activity in human skeletal muscle during aerobic exercise

    Science.gov (United States)

    Holloway, Graham P; Bezaire, Veronic; Heigenhauser, George J F; Tandon, Narendra N; Glatz, Jan F C; Luiken, Joost J F P; Bonen, Arend; Spriet, Lawrence L

    2006-01-01

    Mitochondrial fatty acid transport is a rate-limiting step in long chain fatty acid (LCFA) oxidation. In rat skeletal muscle, the transport of LCFA at the level of mitochondria is regulated by carnitine palmitoyltransferase I (CPTI) activity and the content of malonyl-CoA (M-CoA); however, this relationship is not consistently observed in humans. Recently, fatty acid translocase (FAT)/CD36 was identified on mitochondria isolated from rat and human skeletal muscle and found to be involved in LCFA oxidation. The present study investigated the effects of exercise (120 min of cycling at ∼60% V̇O2peak) on CPTI palmitoyl-CoA and M-CoA kinetics, and on the presence and functional significance of FAT/CD36 on skeletal muscle mitochondria. Whole body fat oxidation rates progressively increased during exercise (P < 0.05), and concomitantly M-CoA inhibition of CPTI was progressively attenuated. Compared to rest, 120 min of cycling reduced (P < 0.05) the inhibition of 0.7, 2, 5 and 10 μm M-CoA by 16%, 21%, 30% and 34%, respectively. Whole body fat oxidation and palmitate oxidation rates in isolated mitochondria progressively increased (P < 0.05) during exercise, and were positively correlated (r = 0.78). Mitochondrial FAT/CD36 protein increased by 63% (P < 0.05) during exercise and was significantly (P < 0.05) correlated with mitochondrial palmitate oxidation rates at all time points (r= 0.41). However, the strongest (P < 0.05) correlation was observed following 120 min of cycling (r= 0.63). Importantly, the addition of sulfo-N-succimidyloleate, a specific inhibitor of FAT/CD36, reduced mitochondrial palmitate oxidation to ∼20%, indicating FAT/CD36 is functionally significant with respect to LCFA oxidation. We hypothesize that exercise-induced increases in fatty acid oxidation occur as a result of an increased ability to transport LCFA into mitochondria. We further suggest that decreased CPTI M-CoA sensitivity and increased mitochondrial FAT/CD36 protein are both

  14. Twin-arginine Translocation System (tat) Mutants of Salmonella are Attenuated Due to Envelope Defects, not Respiratory Defects

    OpenAIRE

    Craig, Maureen; Sadik, Adam Y.; Golubeva, Yekaterina A.; Tidhar, Avital; Slauch, James M.

    2013-01-01

    The twin-arginine translocation system (Tat) transports folded proteins across the cytoplasmic membrane and is critical to virulence in Salmonella and other pathogens. Experimental and bioinformatic data indicate that 30 proteins are exported via Tat in Salmonella Typhimurium. However, there are no data linking specific Tat substrates with virulence. We inactivated every Tat-exported protein and determined the virulence phenotype of mutant strains. Though a tat mutant is highly attenuated, no...

  15. Catabolism and safety of supplemental L-arginine in animals.

    Science.gov (United States)

    Wu, Zhenlong; Hou, Yongqing; Hu, Shengdi; Bazer, Fuller W; Meininger, Cynthia J; McNeal, Catherine J; Wu, Guoyao

    2016-07-01

    L-arginine (Arg) is utilized via multiple pathways to synthesize protein and low-molecular-weight bioactive substances (e.g., nitric oxide, creatine, and polyamines) with enormous physiological importance. Furthermore, Arg regulates cell signaling pathways and gene expression to improve cardiovascular function, augment insulin sensitivity, enhance lean tissue mass, and reduce obesity in humans. Despite its versatile roles, the use of Arg as a dietary supplement is limited due to the lack of data to address concerns over its safety in humans. Data from animal studies are reviewed to assess arginine catabolism and the safety of long-term Arg supplementation. The arginase pathway was responsible for catabolism of 76-85 and 81-96 % Arg in extraintestinal tissues of pigs and rats, respectively. Dietary supplementation with Arg-HCl or the Arg base [315- and 630-mg Arg/(kg BW d) for 91 d] had no adverse effects on male or female pigs. Similarly, no safety issues were observed for male or female rats receiving supplementation with 1.8- and 3.6-g Arg/(kg BW d) for at least 91 d. Intravenous administration of Arg-HCl to gestating sheep at 81 and 180 mg Arg/(kg BW d) is safe for at least 82 and 40 d, respectively. Animals fed conventional diets can well tolerate large amounts of supplemental Arg [up to 630-mg Arg/(kg BW d) in pigs or 3.6-g Arg/(kg BW d) in rats] for 91 d, which are equivalent to 573-mg Arg/(kg BW d) for humans. Collectively, these results can help guide studies to determine the safety of long-term oral administration of Arg in humans. PMID:27156062

  16. L-arginine inhibits isoproterenol-induced cardiac hypertrophy through nitric oxide and polyamine pathways.

    Science.gov (United States)

    Lin, Yan; Wang, Li-Na; Xi, Yu-Hui; Li, Hong-Zhu; Xiao, Feng-Gang; Zhao, Ya-Jun; Tian, Ye; Yang, Bao-Feng; Xu, Chang-Qing

    2008-08-01

    Polyamines (putrescine, spermidine and spermine) are essential for cell growth and differentiation. Nitric oxide exhibits antihypertrophic functions and inhibits cardiac remodelling. However, the metabolism of polyamines and the potential interactions with nitric oxide in cardiac hypertrophy remain unclear. We randomly divided Wistar rats into four treatment groups: controls, isoproterenol (ISO), ISO and L-arginine, and L-arginine. Isoproterenol (5 mg/kg/day, subcutaneously) and/or L-arginine (800 mg/kg/day, intraperitoneally) was administered once daily for 7 days. The expression of atrial natriuretic peptide mRNA was determined by reverse transcription-polymerase chain reaction, and fibrogenesis of heart was assessed by Van Gieson staining. Polyamines were measured with high-performance liquid chromatography, and plasma nitric oxide content and lactate dehydrogenase (LDH) activity were determined with a spectrophotometer. The expression levels of ornithine decarboxylase, spermidine/spermine N1-acetyltransferase (SSAT), endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS) were analysed by Western blot. Heart-to-body weight ratio, left ventricle-to-body weight ratio, atrial natriuretic peptide mRNA expression, collagen fibres and LDH activity were elevated, both ornithine decarboxylase and SSAT proteins were up-regulated, and total polyamines were increased in the group treated with ISO. Additionally, the expression of iNOS was up-regulated, eNOS was down-regulated, and nitric oxide levels were low. Notably, cotreatment with L-arginine reversed most of these changes except for SSAT expression,which was further up-regulated. We propose that increased polyamines and decreased nitric oxide are involved in cardiac hypertrophy induced by ISO and suggest that L-arginine pre-treatment can attenuate cardiac hypertrophy through the regulation of key enzymes of the polyamine and nitric oxide pathways. PMID:18816294

  17. Supplemental arginine above the requirement during suckling causes obesity and insulin resistance in rats.

    Science.gov (United States)

    Otani, Lila; Mori, Tomomi; Koyama, Ayaka; Takahashi, Shin-Ichiro; Kato, Hisanori

    2016-06-01

    Nutrition in early life is important in determining susceptibility to adult obesity, and arginine may promote growth acceleration in infants. We hypothesized that maternal arginine supplementation may promote growth in their pups and contribute to obesity and alteration of the metabolic system in later life. Dams and pups of Wistar rats were given a normal diet (15% protein) as a control (CN) or a normal diet with 2% arginine (ARG). Altered profiles of free amino acids in breast milk were observed in that the concentrations of threonine and glycine were lower in the ARG dams compared with the CN dams. The offspring of the CN and ARG dams were further subdivided into normal-diet (CN-CN and ARG-CN) groups and a high fat-diet groups (CN-HF and ARG-HF). In response to the high fat-diet feeding, the visceral fat deposits were significantly increased in the ARG-HF group (although not compared with the CN-HF group); no difference was observed between the CN-CN and ARG-CN groups. The blood glucose and insulin levels after glucose loading were significantly higher in the ARG-HF group compared with the CN-HF group. The results suggest that the offspring of dams supplemented with arginine during lactation acquired increased susceptibility to a high-fat diet, resulting in visceral obesity and insulin resistance. The lower supply of threonine and glycine to pups may be one of the contributing causes to the programming of lifelong obesity risk in offspring. Our findings also indicated that maternal arginine supplementation during suckling causes obesity and insulin resistance in rats. PMID:27188903

  18. The crucial role of L-arginine in macrophage activation: What you need to know about it

    Czech Academy of Sciences Publication Activity Database

    Pekarová, Michaela; Lojek, Antonín

    2015-01-01

    Roč. 137, SEP2015 (2015), s. 44-48. ISSN 0024-3205 R&D Projects: GA ČR GP13-40882P; GA MŠk(CZ) EE2.3.30.0030 Institutional support: RVO:68081707 Keywords : L-Arginine * Macrophages * G-protein-coupled receptor Subject RIV: BO - Biophysics Impact factor: 2.702, year: 2014

  19. Possible involvement of a plasmid in arginine auxotrophic mutation of Streptomyces kasugaensis.

    OpenAIRE

    Nakano, M M; Ozawa, K; Ogawara, H

    1980-01-01

    Streptomyces kasugaensis gave arginine auxotrophic mutants at high frequency, The coupled loss and reappearance of plasmid deoxyribonucleic acid with arginine auxotrophy suggested that the insertion of the plasmid into chromosomal deoxyribonucleic acid caused the arginine auxotrophy.

  20. The human neonatal small intestine has the potential for arginine synthesis; developmental changes in the expression of arginine-synthesizing and -catabolizing enzymes

    Directory of Open Access Journals (Sweden)

    Ruijter Jan M

    2008-11-01

    Full Text Available Abstract Background Milk contains too little arginine for normal growth, but its precursors proline and glutamine are abundant; the small intestine of rodents and piglets produces arginine from proline during the suckling period; and parenterally fed premature human neonates frequently suffer from hypoargininemia. These findings raise the question whether the neonatal human small intestine also expresses the enzymes that enable the synthesis of arginine from proline and/or glutamine. Carbamoylphosphate synthetase (CPS, ornithine aminotransferase (OAT, argininosuccinate synthetase (ASS, arginase-1 (ARG1, arginase-2 (ARG2, and nitric-oxide synthase (NOS were visualized by semiquantitative immunohistochemistry in 89 small-intestinal specimens. Results Between 23 weeks of gestation and 3 years after birth, CPS- and ASS-protein content in enterocytes was high and then declined to reach adult levels at 5 years. OAT levels declined more gradually, whereas ARG-1 was not expressed. ARG-2 expression increased neonatally to adult levels. Neurons in the enteric plexus strongly expressed ASS, OAT, NOS1 and ARG2, while varicose nerve fibers in the circular layer of the muscularis propria stained for ASS and NOS1 only. The endothelium of small arterioles expressed ASS and NOS3, while their smooth-muscle layer expressed OAT and ARG2. Conclusion The human small intestine acquires the potential to produce arginine well before fetuses become viable outside the uterus. The perinatal human intestine therefore resembles that of rodents and pigs. Enteral ASS behaves as a typical suckling enzyme because its expression all but disappears in the putative weaning period of human infants.

  1. Structures of Bacterial Biosynthetic Arginine Decarboxylases

    Energy Technology Data Exchange (ETDEWEB)

    F Forouhar; S Lew; J Seetharaman; R Xiao; T Acton; G Montelione; L Tong

    2011-12-31

    Biosynthetic arginine decarboxylase (ADC; also known as SpeA) plays an important role in the biosynthesis of polyamines from arginine in bacteria and plants. SpeA is a pyridoxal-5'-phosphate (PLP)-dependent enzyme and shares weak sequence homology with several other PLP-dependent decarboxylases. Here, the crystal structure of PLP-bound SpeA from Campylobacter jejuni is reported at 3.0 {angstrom} resolution and that of Escherichia coli SpeA in complex with a sulfate ion is reported at 3.1 {angstrom} resolution. The structure of the SpeA monomer contains two large domains, an N-terminal TIM-barrel domain followed by a {beta}-sandwich domain, as well as two smaller helical domains. The TIM-barrel and {beta}-sandwich domains share structural homology with several other PLP-dependent decarboxylases, even though the sequence conservation among these enzymes is less than 25%. A similar tetramer is observed for both C. jejuni and E. coli SpeA, composed of two dimers of tightly associated monomers. The active site of SpeA is located at the interface of this dimer and is formed by residues from the TIM-barrel domain of one monomer and a highly conserved loop in the {beta}-sandwich domain of the other monomer. The PLP cofactor is recognized by hydrogen-bonding, {pi}-stacking and van der Waals interactions.

  2. Arginine specific aminopeptidase from Lactobacillus brevis

    Directory of Open Access Journals (Sweden)

    Arya Nandan

    2010-12-01

    Full Text Available The proteolytic system of lactic acid bacteria contribute to the development of flavor during the ripening of cheese through the generation of short peptides and free amino acids, which directly or indirectly act as flavor precursors. Newly isolated lactic acid bacteria (LAB as well as those procured from culture collection centers were screened for the production of various substrate specific aminopeptidases. Among all the strains screened, L. brevis (NRRL B-1836 was found to produce quantifiable amount of intracellular arginine specific aminopeptidase (EC 3.4.11.6. The productivity of arginine aminopeptidase in 5 L fermentor was 36 IU/L/h. The Luedeking and Piret model was tested for intracellular production of aminopeptidase and the data seemed to fit well, as the correlation coefficient was 0.9964 for MRS. The αAP and βAP was 0.4865 and 0.0046, respectively in MRS medium indicating that the yield was predominantly depended on growth. The culture produced lactic acid and also tolerated pH 2.0-3.0 and 0.3-0.5% bile salts, the most important probiotic features.

  3. Arginine Deiminase Resistance in Melanoma Cells Is Associated with Metabolic Reprogramming, Glucose Dependence and Glutamine Addiction

    OpenAIRE

    Long, Yan; Tsai, Wen-Bin; Wangpaichitr, Medhi; Tsukamoto, Takashi; Savaraj, Niramol; Feun, Lynn G.; Kuo, Macus Tien

    2013-01-01

    Many malignant human tumors, including melanomas are auxotrophic for arginine due to reduced expression of argininosuccinate synthetase1 (ASS1), the rate-limiting enzyme for arginine biosynthesis. Pegylated arginine deiminase (ADI-PEG20), which degrades extracellular arginine resulting in arginine deprivation, has shown favorable results in clinical trials for treating arginine-auxotrophic tumors. Drug resistance is the major obstacle for effective ADI-PEG20 usage. To elucidate mechanisms of ...

  4. Effects of dietary salt intake on plasma arginine.

    Science.gov (United States)

    Kitiyakara, C; Chabrashvili, T; Jose, P; Welch, W J; Wilcox, C S

    2001-04-01

    Because L-arginine is degraded by hepatic arginase to ornithine and urea and is transported by the regulated 2A cationic amino acid y(+) transporter (CAT2A), hepatic transport may regulate plasma arginine concentration. Groups of rats (n = 6) were fed a diet of either low salt (LS) or high salt (HS) for 7 days to test the hypothesis that dietary salt intake regulates plasma arginine concentration and renal nitric oxide (NO) generation by measuring plasma arginine and ornithine concentrations, renal NO excretion, and expression of hepatic CAT2A, and arginase. LS rats had lower excretion of NO metabolites and cGMP, lower plasma arginine concentration (LS: 83 +/- 7 vs. HS: 165 +/- 10 micromol/l, P plasma ornithine concentration (LS: 82 +/- 6 vs. HS: 66 +/- 4 micromol/l, P plasma arginine concentration with increased plasma ornithine concentration and urea excretion during LS indicates increased arginine metabolism by arginase. This cannot be ascribed to changes in hepatic arginase expression but may be a consequence of increased hepatic arginine uptake via CAT2A. PMID:11247829

  5. Two distinct arginine methyltransferases are required for biogenesis of Sm-class ribonucleoproteins

    Science.gov (United States)

    Gonsalvez, Graydon B.; Tian, Liping; Ospina, Jason K.; Boisvert, François-Michel; Lamond, Angus I.; Matera, A. Gregory

    2007-01-01

    Small nuclear ribonucleoproteins (snRNPs) are core components of the spliceosome. The U1, U2, U4, and U5 snRNPs each contain a common set of seven Sm proteins. Three of these Sm proteins are posttranslationally modified to contain symmetric dimethylarginine (sDMA) residues within their C-terminal tails. However, the precise function of this modification in the snRNP biogenesis pathway is unclear. Several lines of evidence suggest that the methyltransferase protein arginine methyltransferase 5 (PRMT5) is responsible for sDMA modification of Sm proteins. We found that in human cells, PRMT5 and a newly discovered type II methyltransferase, PRMT7, are each required for Sm protein sDMA modification. Furthermore, we show that the two enzymes function nonredundantly in Sm protein methylation. Lastly, we provide in vivo evidence demonstrating that Sm protein sDMA modification is required for snRNP biogenesis in human cells. PMID:17709427

  6. Anti-aging effects of l-arginine

    Directory of Open Access Journals (Sweden)

    Mohamed Z. Gad

    2010-07-01

    Full Text Available l-Arginine is one of the most metabolically versatile amino acids. In addition to its role in the synthesis of nitric oxide, l-arginine serves as a precursor for the synthesis of polyamines, proline, glutamate, creatine, agmatine and urea. Several human and experimental animal studies have indicated that exogenous l-arginine intake has multiple beneficial pharmacological effects when taken in doses larger than normal dietary consumption. Such effects include reduction in the risk of vascular and heart diseases, reduction in erectile dysfunction, improvement in immune response and inhibition of gastric hyperacidity. This review summarises several positive studies and personal experiences of l-arginine. The demonstrated anti-aging benefits of l-arginine show greater potential than any pharmaceutical or nutraceutical agent ever previously discovered.

  7. Protective effect of L-arginine against necrosis and apoptosis induced by experimental ischemic and reperfusion in rat liver

    International Nuclear Information System (INIS)

    To study the effect of L-arginine on apoptosis and necrosis induced by 1-h ischemia followed by 3-h reperfusion. Adult Wistar rats underwent 60 min of partial liver ischemia followed by 3-h reperfusion. Eighteen Wistar rats were divided into sham-operated control group (1) ( n = 6), ischemia and reperfusion (I/R) group (0.9 % saline (5 ml/kg, orally) for 7 days) (2) ( n 6), and L-arginine-treated group (10 mg/kg body weight daily orally for 7 days before inducing ischemia-reperfusion maneuver) (3) ( n = 6). Apoptotic and necrotic hepatocytes, nitric oxide levels in hepatocytes, Bcl-2 mRNA, and Bcl-2 protein were measured. Liver injury was assessed by plasma alanine transaminases (ALT), aspartate transaminases (AST), liver histopathology, and electron microscopy. An ischemic and reperfusion hepatocellular injury occurred as was indicated by increased serum ALT, AST, histopathology, and electron microscopy. Apoptosis and necrosis associated marker gene Bcl-2 mRNA and protein expression were decreased in I/R group. Pretreatment with L-arginine significantly decreased serum ALT and AST level and apoptotic and necrotic cells after 1 h ischemia followed by 3 h of reperfusion. Nitric oxide production in hepatocytes was increased twofold by L-arginine treatment when compared with I/R group. Histopathology and transmission electron microscopy (TEM) studies showed markedly diminished hepatocellular injury in L-arginine-pretreated rats during the hepatic I/R. Thus, it may be concluded that L-arginine afforded significant protection from necrosis and apoptosis in I/R injury by upregulated Bcl-2 gene and nitric oxide production. (author)

  8. EPR, Endor and DFT Studies on X-Irradiated Single Crystals of L-Lysine HCl 2 H 2O and L-Arginine HCl H2O

    Science.gov (United States)

    Zhou, Yiying; Nelson, William H.

    2011-03-01

    When proteins and DNA interact, arginine and lysine are the two amino acids most often in close contact with the DNA. In order to understand the radiation damage to DNA in vivo, which is always associated with protein, it is important to learn the radiation chemistry of arginine and lysine independently, and then complexed to DNA. This work studied X-irradiated single crystals of L- lysine . HCl . 2 H2 O and L- arginine . HCl . H2 O with EPR, ENDOR techniques and DFT calculations. In both crystal types irradiated at 66K, the carboxyl anion radical and the decarboxylation radical were identified. Specifically, the calculations performed on the cluster models for the carboxyl anion radicals reproduced the proton transfers to the carboxyl group from the neighboring molecules through the hydrogen bonds. Moreover, computations supported the identification of one radical type within irradiated arginine as the guanidyl radical anion with an electron trapped by the guanidyl group. Based on the radicals detected in the crystal irradiated at 66K and at 298K, and the annealing experiments from the irradiation at 66K, the mechanisms of the irradiation damage on lysine and arginine were proposed, and the possible effects of irradiated arginine and lysine to the DNA within chromatin were analyzed.

  9. Arginine and proline applied as food additives stimulate high freeze tolerance in larvae of Drosophila melanogaster.

    Science.gov (United States)

    Koštál, Vladimír; Korbelová, Jaroslava; Poupardin, Rodolphe; Moos, Martin; Šimek, Petr

    2016-08-01

    The fruit fly Drosophila melanogaster is an insect of tropical origin. Its larval stage is evolutionarily adapted for rapid growth and development under warm conditions and shows high sensitivity to cold. In this study, we further developed an optimal acclimation and freezing protocol that significantly improves larval freeze tolerance (an ability to survive at -5°C when most of the freezable fraction of water is converted to ice). Using the optimal protocol, freeze survival to adult stage increased from 0.7% to 12.6% in the larvae fed standard diet (agar, sugar, yeast, cornmeal). Next, we fed the larvae diets augmented with 31 different amino compounds, administered in different concentrations, and observed their effects on larval metabolomic composition, viability, rate of development and freeze tolerance. While some diet additives were toxic, others showed positive effects on freeze tolerance. Statistical correlation revealed tight association between high freeze tolerance and high levels of amino compounds involved in arginine and proline metabolism. Proline- and arginine-augmented diets showed the highest potential, improving freeze survival to 42.1% and 50.6%, respectively. Two plausible mechanisms by which high concentrations of proline and arginine might stimulate high freeze tolerance are discussed: (i) proline, probably in combination with trehalose, could reduce partial unfolding of proteins and prevent membrane fusions in the larvae exposed to thermal stress (prior to freezing) or during freeze dehydration; (ii) both arginine and proline are exceptional among amino compounds in their ability to form supramolecular aggregates which probably bind partially unfolded proteins and inhibit their aggregation under increasing freeze dehydration. PMID:27489218

  10. Toxoplasma gondii lacks the enzymes required for de novo arginine biosynthesis and arginine starvation triggers cyst formation.

    Science.gov (United States)

    Fox, Barbara A; Gigley, Jason P; Bzik, David J

    2004-03-01

    Two separate carbamoyl phosphate synthetase activities are required for the de novo synthesis of pyrimidines and arginine in most eukaryotes. Toxoplasma gondii is novel in possessing a single carbamoyl phosphate synthetase II gene that corresponds to a glutamine-dependent form required for pyrimidine biosynthesis. We therefore examined arginine acquisition in T. gondii to determine whether the single carbamoyl phosphate synthetase II activity could provide both pyrimidine and arginine biosynthesis. We found that arginine deprivation efficiently blocks the replication of intracellular T. gondii, yet has little effect on long-term parasite viability. Addition of citrulline, but not ornithine, rescues the growth defect observed in the absence of exogenous arginine. This rescue with citrulline is ablated when parasites are cultured in a human citrullinemia fibroblast cell line that is deficient in argininosuccinate synthetase activity. These results reveal the absence of genes and activities of the arginine biosynthetic pathway and demonstrate that T. gondii is an arginine auxotroph. Arginine starvation was also found to efficiently trigger differentiation of replicative tachyzoites into bradyzoites contained within stable cyst-like structures. These same parasites expressing bradyzoite antigens can be efficiently switched back to rapidly proliferating tachyzoites several weeks after arginine starvation. We hypothesise that the absence of gene activities that are essential for the biosynthesis of arginine from carbamoyl phosphate confers a selective advantage by increasing bradyzoite switching during the host response to T. gondii infection. These findings are consistent with a model of host-parasite evolution that allowed host control of bradyzoite induction by trading off virulence for increased transmission. PMID:15003493

  11. Asymmetric Dimethyl Arginine in Hypothyroid Patients

    International Nuclear Information System (INIS)

    Thyroid diseases may lead to endothelial dysfunction, however, the mechanism underlying the endothelial dysfunction in thyroid disease is still not clear. Asymmetric dimethyl arginine (ADMA), a novel inhibitor of endothelial nitric oxide synthetase (eNOS), was reported to inhibit nitric oxide (NO) synthesis from L-arginine. The present study was carried out to investigate ADMA levels together with effects of dislipidemia in sub-clinical and overt hypothyroid females. There were significant increase in the levels of total cholesterol, low density lipoprotein-cholesterol (LDL-c), high density lipoprotein-cholesterol (HDL-c), thyroid stimulating hormone (TSH) and ADMA in hypothyroid females as compared to controls while the levels of NO and free T4 were significantly decreased than controls. Sub-clinical hypothyroid females had significant high TSH, LDL-c and non-significantly high ADMA levels and total cholesterol as compared to controls while they had significant decrease in NO, HDL-c and non-significant decrease in free T4 as compared to controls. There were significant negative correlations between NO and both ADMA (r2 = 0.84) and free T4 (r2 = 0.95) in overt hypothyroid group while significant positive correlation (r2 = 0.85) was detected between TSH and HDL-c in the same group. These results are highly suggestive that the decrease of nitric oxide secondary to accumulation of ADMA represent an important pathogenic factor together with dyslipidemia in endothelial dysfunction and increased cardiovascular risk especially in hypothyroid females

  12. Carnitine palmitoyltransferase 2 and carnitine/acylcarnitine translocase are involved in the mitochondrial synthesis and export of acylcarnitines.

    Science.gov (United States)

    Violante, Sara; Ijlst, Lodewijk; Te Brinke, Heleen; Tavares de Almeida, Isabel; Wanders, Ronald J A; Ventura, Fátima V; Houten, Sander M

    2013-05-01

    Acylcarnitines are commonly used in the diagnosis of mitochondrial fatty acid β-oxidation disorders (mFAODs). It is generally assumed that this plasma acylcarnitine profile reflects the mitochondrial accumulation of acyl-CoAs. The identity of the enzymes and the mitochondrial and plasmalemmal transporters involved in the synthesis and export of these metabolites have remained undefined. We used lentiviral shRNA to knock down the expression of medium-chain acyl-CoA dehydrogenase (MCAD) in control and carnitine palmitoyltransferase 2 (CPT2)-, carnitine/acylcarnitine translocase (CACT)-, and plasmalemmal carnitine transporter (OCTN2)-deficient human fibroblasts. These cell lines, including mock-transduced controls, were loaded with decanoic acid and carnitine, followed by the measurement of the acylcarnitine profile in the extracellular medium. In control fibroblasts, MCAD knockdown markedly increased the production of octanoylcarnitine (3-fold, P<0.01). OCTN2-deficient cell lines also showed extracellular accumulation of octanoylcarnitine (2.8-fold, P<0.01), suggesting that the cellular export of acylcarnitines does not depend on OCTN2. In contrast, in CPT2- and CACT-deficient cells, the accumulation of octanoylcarnitine in the medium did not significantly increase in the MCAD knockdown. Similar results were obtained using pharmacological inhibition of CPT2 in fibroblasts from MCAD-deficient individuals. This shows that CPT2 and CACT are crucial for mitochondrial acylcarnitine formation and export to the extracellular fluids in mFAOD. PMID:23322164

  13. Over-expression of Adenine Nucleotide Translocase 1 (ANT1) Induces Apoptosis and Tumor Regression in vivo

    International Nuclear Information System (INIS)

    Adenine nucleotide translocase (ANT) is located in the inner mitochondrial membrane and catalyzes the exchange of mitochondrial ATP for cytosolic ADP. ANT has been known to be a major component of the permeability transition pore complex of mitochondria and contributes to mitochondria-mediated apoptosis. Human ANT has four isoforms (ANT1, ANT2, ANT3, and ANT4), and the expression of the ANT isoforms is variable depending on the tissue and cell type, developmental stage, and proliferation status. Among the isoforms, ANT1 is highly expressed in terminally-differentiated tissues, but expressed in low levels in proliferating cells, such as cancer cells. In particular, over-expression of ANT1 induces apoptosis in cultured tumor cells. We applied an ANT1 gene transfer approach to induce apoptosis and to evaluate the anti-tumor effect of ANT1 in a nude mouse model. We demonstrated that ANT1 transfection induced apoptosis of MDA-MB-231 cells, inactivated NF-κB activity, and increased Bax expression. ANT1-inducing apoptosis was accompanied by the disruption of mitochondrial membrane potential, cytochrome c release and the activation of caspases-9 and -3. Moreover, ANT1 transfection significantly suppressed tumor growth in vivo. Our results suggest that ANT1 transfection may be a useful therapeutic modality for the treatment of cancer

  14. Over-expression of Adenine Nucleotide Translocase 1 (ANT1 Induces Apoptosis and Tumor Regression in vivo

    Directory of Open Access Journals (Sweden)

    Kim Chul-Woo

    2008-06-01

    Full Text Available Abstract Background Adenine nucleotide translocase (ANT is located in the inner mitochondrial membrane and catalyzes the exchange of mitochondrial ATP for cytosolic ADP. ANT has been known to be a major component of the permeability transition pore complex of mitochondria and contributes to mitochondria-mediated apoptosis. Human ANT has four isoforms (ANT1, ANT2, ANT3, and ANT4, and the expression of the ANT isoforms is variable depending on the tissue and cell type, developmental stage, and proliferation status. Among the isoforms, ANT1 is highly expressed in terminally-differentiated tissues, but expressed in low levels in proliferating cells, such as cancer cells. In particular, over-expression of ANT1 induces apoptosis in cultured tumor cells. Methods We applied an ANT1 gene transfer approach to induce apoptosis and to evaluate the anti-tumor effect of ANT1 in a nude mouse model. Results We demonstrated that ANT1 transfection induced apoptosis of MDA-MB-231 cells, inactivated NF-κB activity, and increased Bax expression. ANT1-inducing apoptosis was accompanied by the disruption of mitochondrial membrane potential, cytochrome c release and the activation of caspases-9 and -3. Moreover, ANT1 transfection significantly suppressed tumor growth in vivo. Conclusion Our results suggest that ANT1 transfection may be a useful therapeutic modality for the treatment of cancer.

  15. Arginine inhibits aggregation of α-lactalbumin but also decreases its stability: Calorimetric, spectroscopic, and molecular dynamics studies

    International Nuclear Information System (INIS)

    Highlights: • Arginine inhibits aggregation of α-lactalbumin, however, it destabilizes the protein. • ITC results show favorable interactions between α-lactalbumin and arginine. • Strong interactions of guanidinium group with α-lactalbumin leads to its destabilization. • Like charges repulsions decreases the severity of the destabilization. - Abstract: Arginine (Arg) has long been used to inhibit aggregation of proteins. Despite its frequent use in inhibition of aggregation, the exact mechanism of aggregation and the effect of Arg on the conformation and stability of proteins are still not well understood. In the present study, spectroscopic, calorimetric, and molecular dynamics methods have been used to understand the mechanism of inhibition of aggregation of α-lactalbumin (α-LA) by Arg along with its effect on stability of the protein. It is observed that although Arg inhibits aggregation of α-LA, it also decreases stability of the protein. The results suggest that strong favorable interactions between α-LA and guanidinium group of Arg decrease the stability of the protein. The results also suggest that the guanidinium group preferentially interacts with Gln, Asn and negatively charged residues through hydrogen bonding and electrostatic interactions

  16. Arginine affects appetite via nitric oxide in ducks.

    Science.gov (United States)

    Wang, C; Hou, S S; Huang, W; Xu, T S; Rong, G H; Xie, M

    2014-08-01

    The objective of the study was to investigate the mechanism by which arginine regulates feed intake in Pekin ducks. In experiment 1, one hundred forty-four 1-d-old male Pekin ducks were randomly allotted to 3 dietary treatments with 6 replicate pens of 8 birds per pen. Birds in each group were fed a corn-corn gluten meal diet containing 0.65, 0.95, and 1.45% arginine. Ducks fed the diet containing 0.65% arginine had lower feed intake and plasma nitric oxide level (P Pekin ducks. PMID:24902706

  17. Mammalian SRP receptor switches the Sec61 translocase from Sec62 to SRP-dependent translocation

    Czech Academy of Sciences Publication Activity Database

    Jadhav, B.; McKenna, M.; Johnson, Nicholas; High, S.; Sinning, I.; Pool, M. R.

    2015-01-01

    Roč. 6, Dec (2015), 10133/1-10133/11. ISSN 2041-1723 Institutional support: RVO:61388963 Keywords : signal recognition particle * endoplasmic reticulum membrane * tail-anchored protein Subject RIV: CE - Biochemistry Impact factor: 11.470, year: 2014 http://www.nature.com/ncomms/2015/151204/ncomms10133/full/ncomms10133.html

  18. Caries-free subjects have high levels of urease and arginine deiminase activity

    Directory of Open Access Journals (Sweden)

    Evelyn REYES

    2014-06-01

    Full Text Available Objectives: This study investigated the relationship between urease and arginine deiminase system (ADS activities and dental caries through a cross-sectional study. Material and Methods: Urease and ADS activities were measured in saliva and plaque samples from 10 caries-free subjects and 13 caries-active. Urease activity was obtained from the ammonia produced by incubation of plaque and saliva samples in urea. ADS activity was obtained from the ammonia generated by the arginine-HCl and Tris-maleate buffer. Specific activity was defined as micromoles of ammonia per minute per milligram of protein. Shapiro-Wilk statistical test was used to analyze the distribution of the data, and Mann-Whitney test was used to determine the significance of the data. Results: The specific urease activity in saliva and plaque was significantly higher in individuals with low DMFT scores. ADS activity in saliva (6.050 vs 1.350, p=0.0154 and plaque (8.830 vs 1.210, p=0.025 was also higher in individuals with low DMFT scores. Conclusions: Caries-free subjects had a higher ammonia generation activity by urease and arginine deiminase system for both saliva and plaque samples than low caries-active subjects. High levels of alkali production in oral environment were related to caries-free subjects.

  19. Evolution of phosphagen kinase V. cDNA-derived amino acid sequences of two molluscan arginine kinases from the chiton Liolophura japonica and the turbanshell Battilus cornutus.

    Science.gov (United States)

    Suzuki, T; Ban, T; Furukohri, T

    1997-06-20

    The cDNAs of arginine kinases from the chiton Liolophura japonica (Polyplacophora) and the turbanshell Battilus cornutus (Gastropoda) were amplified by polymerase chain reaction (PCR), and the complete nucleotide sequences of 1669 and 1624 bp, respectively, were determined. The open reading frame for Liolophura arginine kinase is 1050 nucleotides in length and encodes a protein with 349 amino acid residues, and that for Battilus is 1077 nucleotides and 358 residues. The validity of the cDNA-derived amino acid sequence was supported by chemical sequencing of internal tryptic peptides. The molecular masses were calculated to be 39,057 and 39,795 Da, respectively. The amino acid sequence of Liolophura arginine kinase showed 65-68% identity with those of Battilus and Nordotis (abalone) arginine kinases, and the homology between Battilus and Nordotis was 79%. Molluscan arginine kinases also show lower, but significant homology (38-43%) with rabbit creatine kinase. The sequences of arginine kinases could be used as a molecular clock to elucidate the phylogeny of Mollusca, one of the most diverse animal phyla. PMID:9217008

  20. Measurement of arginine metabolites: regulators of nitric oxide metabolism.

    Science.gov (United States)

    Augustine, Molly S; Rogers, Lynette K

    2013-01-01

    Arginine is the substrate for nitric oxide synthases (NOS), and arginine availability regulates the production of nitric oxide. Through the activity of methyltransferases, arginine can be methylated to form monomethylarginine (NMMA), asymmetrical dimethylarginine (ADMA), and symmetrical dimethylarginine (SDMA). NMMA and ADMA directly inhibit NOS, whereas SDMA inhibits the cellular import of arginine through the cationic amino acid transporter. Increased levels of methylarginine compounds have been associated with many diseases including atherosclerosis, renal failure, pulmonary hypertension, and preeclampsia. Previous HPLC methods to measure these molecules rely on derivatization with ortho-phthalaldehyde, which is unstable and requires immediate pre- or post-column reactions. We have identified a new fluorometric agent that is stable for at least 1 week and provides chromatographic properties that facilitate separation of these chemically similar compounds by reverse phase chromatography. PMID:24510541

  1. Characterization of the arcD Arginine: Ornithine Exchanger of Pseudomonas aeruginosa. Localization in the Cytoplasmic Membrane and a Topological Model

    OpenAIRE

    Bourdineaud, Jean-Paul; Heierli, Daniel; Gamper, Marianne; Verhoogt, Hans J.C.; Driessen, Arnold J. M.; Konings, Wil N.; Lazdunski, Claude; Haas, Dieter

    1993-01-01

    The arcDABC operon of Pseudomonas aeruginosa encodes the enzymes of the arginine deiminase pathway and is induced by oxygen limitation. The arcD gene specifies a 53-kDa protein with arginine: ornithine exchange activity. The ArcD protein of P. aeruginosa, like the LysI lysine transporter of Corynebacterium glutamicum, has 13 hydrophobic regions which could span the cytoplasmic membrane. Fusion of a Caa (colicin A) epitope to the N-terminal part of ArcD permitted the localization, by immunoblo...

  2. Cellular Mechanisms of L-arginine Induced Experimental Acute Pancreatitis

    OpenAIRE

    Masood, Omar

    2013-01-01

    AbstractThe University Of ManchesterOmar MasoodMD Thesis 2013Cellular Mechanisms of L-arginine Induced Experimental Acute Pancreatitis. IntroductionImpairment of cytosolic calcium ([Ca2+]i) signaling and in particular calcium overload has emerged as a possible unifying mechanism for precipitating acute pancreatitis (AP.)In the L-arginine (L-arg) experimental model of AP, nitric oxide (NO) has been implicated however the disease progression is largely unaffected by nitric oxide synthase (NOS) ...

  3. L-Arginine Pathway in COPD Patients with Acute Exacerbation

    DEFF Research Database (Denmark)

    Ruzsics, Istvan; Nagy, Lajos; Keki, Sandor; Sarosi, Veronika; Illes, Balazs; Illes, Zsolt; Horvath, Ildiko; Bogar, Lajos; Molnar, Tihamer

    BACKGROUND: Acute exacerbation of chronic obstructive pulmonary disease (AECOPD) remains a major cause of mortality. Clinical criteria of AECOPD are subjective. Biomarkers for AECOPD may aid in the initiation of early treatment. Increased production of asymmetric and symmetric dimethylarginine (A......-arginine, ADMA and SDMA serum levels. In patients with AECOPD, production of ADMA and SDMA are more pronounced presumably due to more severe hypoxic insult. Methylated arginine derivatives in the sera may help early recognition of AECOPD....

  4. L-Arginine ameliorates cardiac left ventricular oxidative stress by upregulating eNOS and Nrf2 target genes in alloxan-induced hyperglycemic rats

    Energy Technology Data Exchange (ETDEWEB)

    Ramprasath, Tharmarajan; Hamenth Kumar, Palani; Syed Mohamed Puhari, Shanavas; Senthil Murugan, Ponniah; Vasudevan, Varadaraj [Molecular Cardiology Unit, Department of Biochemistry, Center for Excellence in Genomic Sciences, School of Biological Sciences, Madurai Kamaraj University, Madurai 625 021, Tamilnadu (India); Selvam, Govindan Sadasivam, E-mail: drselvamgsbiochem@rediffmail.com [Molecular Cardiology Unit, Department of Biochemistry, Center for Excellence in Genomic Sciences, School of Biological Sciences, Madurai Kamaraj University, Madurai 625 021, Tamilnadu (India)

    2012-11-23

    Highlights: Black-Right-Pointing-Pointer L-Arginine treatment reduced the metabolic disturbances in diabetic animals. Black-Right-Pointing-Pointer Antioxidant marker proteins were found high in myocardium by L-arginine treatment. Black-Right-Pointing-Pointer Elevated antioxidant status, mediates the reduced TBA-reactivity in left ventricle. Black-Right-Pointing-Pointer L-Arginine treatment enhanced the Nrf2 and eNOS signaling in left ventricle. Black-Right-Pointing-Pointer Improved cell survival signaling by arginine, offers a novel tactic for targeting. -- Abstract: Hyperglycemia is independently related with excessive morbidity and mortality in cardiovascular disorders. L-Arginine-nitric oxide (NO) pathway and the involvement of NO in modulating nuclear factor-E2-related factor-2 (Nrf2) signaling were well established. In the present study we investigated, whether L-arginine supplementation would improve the myocardial antioxidant defense under hyperglycemia through activation of Nrf2 signaling. Diabetes was induced by alloxan monohydrate (90 mg kg{sup -1} body weight) in rats. Both non-diabetic and diabetic group of rats were divided into three subgroups and they were administered either with L-arginine (2.25%) or L-NAME (0.01%) in drinking water for 12 days. Results showed that L-arginine treatment reduced the metabolic disturbances in diabetic rats. Antioxidant enzymes and glutathione levels were found to be increased in heart left ventricles, thereby reduction of lipid peroxidation by L-arginine treatment. Heart histopathological analysis further validates the reversal of typical diabetic characteristics consisting of alterations in myofibers and myofibrillary degeneration. qRT-PCR studies revealed that L-arginine treatment upregulated the transcription of Akt and downregulated NF-{kappa}B. Notably, transcription of eNOS and Nrf2 target genes was also upregulated, which were accompanied by enhanced expression of Nrf2 in left ventricular tissue from diabetic

  5. L-Arginine ameliorates cardiac left ventricular oxidative stress by upregulating eNOS and Nrf2 target genes in alloxan-induced hyperglycemic rats

    International Nuclear Information System (INIS)

    Highlights: ► L-Arginine treatment reduced the metabolic disturbances in diabetic animals. ► Antioxidant marker proteins were found high in myocardium by L-arginine treatment. ► Elevated antioxidant status, mediates the reduced TBA-reactivity in left ventricle. ► L-Arginine treatment enhanced the Nrf2 and eNOS signaling in left ventricle. ► Improved cell survival signaling by arginine, offers a novel tactic for targeting. -- Abstract: Hyperglycemia is independently related with excessive morbidity and mortality in cardiovascular disorders. L-Arginine-nitric oxide (NO) pathway and the involvement of NO in modulating nuclear factor-E2-related factor-2 (Nrf2) signaling were well established. In the present study we investigated, whether L-arginine supplementation would improve the myocardial antioxidant defense under hyperglycemia through activation of Nrf2 signaling. Diabetes was induced by alloxan monohydrate (90 mg kg−1 body weight) in rats. Both non-diabetic and diabetic group of rats were divided into three subgroups and they were administered either with L-arginine (2.25%) or L-NAME (0.01%) in drinking water for 12 days. Results showed that L-arginine treatment reduced the metabolic disturbances in diabetic rats. Antioxidant enzymes and glutathione levels were found to be increased in heart left ventricles, thereby reduction of lipid peroxidation by L-arginine treatment. Heart histopathological analysis further validates the reversal of typical diabetic characteristics consisting of alterations in myofibers and myofibrillary degeneration. qRT-PCR studies revealed that L-arginine treatment upregulated the transcription of Akt and downregulated NF-κB. Notably, transcription of eNOS and Nrf2 target genes was also upregulated, which were accompanied by enhanced expression of Nrf2 in left ventricular tissue from diabetic and control rats. Under these findings, we suggest that targeting of eNOS and Nrf2 signaling by L-arginine supplementation could be

  6. Proteolytic processing of Escherichia coli twin-arginine signal peptides by LepB.

    Science.gov (United States)

    Lüke, Iris; Handford, Jennifer I; Palmer, Tracy; Sargent, Frank

    2009-12-01

    The twin-arginine translocation (Tat) apparatus is a protein targeting system found in the cytoplasmic membranes of many prokaryotes. Substrate proteins of the Tat pathway are synthesised with signal peptides bearing SRRxFLK 'twin-arginine' amino acid motifs. All Tat signal peptides have a common tripartite structure comprising a polar N-terminal region, followed by a hydrophobic region of variable length and a polar C-terminal region. In Escherichia coli, Tat signal peptides are proteolytically cleaved after translocation. The signal peptide C-terminal regions contain conserved AxA motifs, which are possible recognition sequences for leader peptidase I (LepB). In this work, the role of LepB in Tat signal peptide processing was addressed directly. Deliberate repression of lepB expression prevented processing of all Tat substrates tested, including SufI, AmiC, and a TorA-23K reporter protein. In addition, electron microscopy revealed gross defects in cell architecture and membrane integrity following depletion of cellular LepB protein levels. PMID:19809807

  7. Acellular matrix of bovine pericardium bound with L-arginine

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Hyo Joo [Department of Molecular Science and Technology, Ajou University, Suwon 443-749 (Korea, Republic of); Bae, Jin Woo [Department of Molecular Science and Technology, Ajou University, Suwon 443-749 (Korea, Republic of); Kim, Chun Ho [Laboratory of Tissue Engineering, Korea Cancer Center Hospital, Seoul 139-240 (Korea, Republic of); Lee, Jin Woo [Department of Orthopaedic Surgery, College of Medicine, Yonsei University, Seoul 120-749 (Korea, Republic of); Shin, Jung Woog [Department of Biomedical Engineering, Inje University, Gimhae 621-749 (Korea, Republic of); Park, Ki Dong [Department of Molecular Science and Technology, Ajou University, Suwon 443-749 (Korea, Republic of)

    2007-09-15

    Surface immobilization of bioactive molecules onto natural tissues has been interestingly studied for the development of new functional matrices for the replacement of lost or malfunctioning tissues. In this study, an acellular matrix of bovine pericardium (ABP) was chemically modified by the direct coupling of L-arginine after glutaraldehyde (GA) cross-linking. The effects of L-arginine coupling on durability and calcification were investigated and the biocompatibility was evaluated in vitro and in vivo. A four-step detergent and enzymatic extraction process has been utilized to remove cellular components from fresh bovine pericardium (BP). Microscopic observation confirmed that nearly all cellular constituents are removed. Thermal and mechanical properties showed that the durability of L-arginine-treated matrices increased as compared with control ABP and GA-treated ABP. Resistance to collagenase digestion revealed that modified matrices have greater resistance to enzyme digestion than control ABP and GA-treated ABP. The in vivo calcification study demonstrated much less calcium deposition on L-arginine-treated ABP than GA-treated one. In vitro cell viability results showed that ABP modified with L-arginine leads to a significant increase in attachment of human dermal fibroblasts. The obtained results attest to the usefulness of L-arginine-treated ABP matrices for cardiovascular bioprostheses.

  8. Unraveling the Fanconi anaemia-DNA repair connection through DNA helicase and translocase activities

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, L H

    2005-08-16

    How the Fanconi anaemia (FA) chromosome stability pathway functions to cope with interstrand crosslinks and other DNA lesions has been elusive, even after FANCD1 proved to be BRCA2, a partner of Rad51 in homologous recombination. The identification and characterization of two new Fanconi proteins having helicase motifs, FANCM and FANCJ/BRIP1/BACH1, implicates the FANC nuclear core complex as a participant in recognizing or processing damaged DNA, and the BRIP1 helicase as acting independently of this complex.

  9. Active displacement of RecA filaments by UvrD translocase activity

    OpenAIRE

    Petrova, Vessela; Chen, Stefanie H.; Molzberger, Eileen T.; Tomko, Eric; Chitteni-Pattu, Sindhu; Jia, Haifeng; Ordabayev, Yerdos; Lohman, Timothy M.; Cox, Michael M.

    2015-01-01

    The UvrD helicase has been implicated in the disassembly of RecA nucleoprotein filaments in vivo and in vitro. We demonstrate that UvrD utilizes an active mechanism to remove RecA from the DNA. Efficient RecA removal depends on the availability of DNA binding sites for UvrD and/or the accessibility of the RecA filament ends. The removal of RecA from DNA also requires ATP hydrolysis by the UvrD helicase but not by RecA protein. The RecA-removal activity of UvrD is slowed by RecA variants with ...

  10. Human Upf1 is a highly processive RNA helicase and translocase with RNP remodelling activities

    Science.gov (United States)

    Fiorini, Francesca; Bagchi, Debjani; Le Hir, Hervé; Croquette, Vincent

    2015-07-01

    RNA helicases are implicated in most cellular RNA-dependent events. In eukaryotes however, only few have been functionally characterized. Upf1 is a RNA helicase essential for nonsense-mediated mRNA decay (NMD). Here, using magnetic tweezers and bulk assays, we observe that human Upf1 is able to translocate slowly over long single-stranded nucleic acids with a processivity >10 kb. Upf1 efficiently translocates through double-stranded structures and protein-bound sequences, demonstrating that Upf1 is an efficient ribonucleoprotein complex remodeler. Our observation of processive unwinding by an eukaryotic RNA helicase reveals that Upf1, once recruited onto NMD mRNA targets, can scan the entire transcript to irreversibly remodel the mRNP, facilitating its degradation by the NMD machinery.

  11. L-Arginine Supplementation and Metabolism in Asthma

    Directory of Open Access Journals (Sweden)

    Angela Linderholm

    2011-01-01

    Full Text Available L-Arginine, the amino acid substrate for nitric oxide synthase, has been tested as a therapeutic intervention in a variety of chronic diseases and is commonly used as a nutritional supplement. In this study, we hypothesized that a subset of moderate to severe persistent asthma patients would benefit from supplementation with L-arginine by transiently increasing nitric oxide levels, resulting in bronchodilation and a reduction in inflammation. The pilot study consisted of a 3 month randomized, double-blind, placebo-controlled trial of L-arginine (0.05 g/kg twice daily in patients with moderate to severe asthma. We measured spirometry, exhaled breath nitric oxide, serum arginine metabolites, questionnaire scores, daily medication use and PEFR with the primary endpoint being the number of minor exacerbations at three months. Interim analysis of the 20 subjects showed no difference in the number of exacerbations, exhaled nitric oxide levels or lung function between groups, though participants in the L-arginine group had higher serum L-arginine at day 60 (2.0 ± 0.6 × 10−3 vs. 1.1 ± 0.2 × 10−3 µmol/L, p < 0.05, ornithine at day 30 (2.4 ± 0.9 vs. 1.2 ± 0.3 µmol/L serum, p < 0.05 and ADMA at day 30 (6.0 ± 1.5 × 10−1 vs. 2.6 ± 0.6 × 10−1 µmol/L serum, p < 0.05 on average compared to the placebo group. The study was terminated prematurely. Supplementing asthma subjects with L-arginine increases plasma levels; whether subgroups might benefit from such supplementation requires further study.

  12. The formation of argpyrimidine, a methylglyoxal-arginine adduct, in the nucleus of neural cells

    International Nuclear Information System (INIS)

    Methylglyoxal (MG) is an endogenous metabolite in glycolysis and forms stable adducts primarily with arginine residues of intracellular proteins. The biological role of this modification in cell function is not known. In the present study, we found that a MG-detoxification enzyme glyoxalase I (GLO1) is mainly expressed in the ventricular zone (VZ) at embryonic day 16 which neural stem and progenitor cells localize. Moreover, immunohistochemical analysis revealed that argpyrimidine, a major MG-arginine adduct, is predominantly produced in cortical plate neurons not VZ during cerebral cortex development and is exclusively located in the nucleus. Immunoblotting experiment showed that the formation of argpyrimidine occurs on some nuclear proteins of cortical neurons. To our knowledge, this is first report of the argpyrimidine formation in the nucleus of neuron. These findings suggest that GLO1, which is dominantly expressed in the embryonic VZ, reduces the intracellular level of MG and suppresses the formation of argpyrimidine in neural stem and progenitor cells. Argpyrimidine may contribute to the neural differentiation and/or the maintenance of the differentiated state via the modification of nuclear proteins.

  13. MTAP deletion confers enhanced dependency on the PRMT5 arginine methyltransferase in cancer cells | Office of Cancer Genomics

    Science.gov (United States)

    The discovery of cancer dependencies has the potential to inform therapeutic strategies and to identify putative drug targets. Integrating data from comprehensive genomic profiling of cancer cell lines and from functional characterization of cancer cell dependencies, we discovered that loss of the enzyme methylthioadenosine phosphorylase (MTAP) confers a selective dependence on protein arginine methyltransferase 5 (PRMT5) and its binding partner WDR77. MTAP is frequently lost due to its proximity to the commonly deleted tumor suppressor gene, CDKN2A.

  14. Identification of arginine- and lysine-methylation in the proteome of Saccharomyces cerevisiae and its functional implications

    Directory of Open Access Journals (Sweden)

    Gasteiger Elisabeth

    2010-02-01

    Full Text Available Abstract Background The methylation of eukaryotic proteins has been proposed to be widespread, but this has not been conclusively shown to date. In this study, we examined 36,854 previously generated peptide mass spectra from 2,607 Saccharomyces cerevisiae proteins for the presence of arginine and lysine methylation. This was done using the FindMod tool and 5 filters that took advantage of the high number of replicate analysis per protein and the presence of overlapping peptides. Results A total of 83 high-confidence lysine and arginine methylation sites were found in 66 proteins. Motif analysis revealed many methylated sites were associated with MK, RGG/RXG/RGX or WXXXR motifs. Functionally, methylated proteins were significantly enriched for protein translation, ribosomal biogenesis and assembly and organellar organisation and were predominantly found in the cytoplasm and ribosome. Intriguingly, methylated proteins were seen to have significantly longer half-life than proteins for which no methylation was found. Some 43% of methylated lysine sites were predicted to be amenable to ubiquitination, suggesting methyl-lysine might block the action of ubiquitin ligase. Conclusions This study suggests protein methylation to be quite widespread, albeit associated with specific functions. Large-scale tandem mass spectroscopy analyses will help to further confirm the modifications reported here.

  15. Transsulfuration pathway thiols and methylated arginines: the Hunter Community Study.

    Directory of Open Access Journals (Sweden)

    Arduino A Mangoni

    Full Text Available BACKGROUND: Serum homocysteine, when studied singly, has been reported to be positively associated both with the endogenous nitric oxide synthase inhibitor asymmetric dimethylarginine [ADMA, via inhibition of dimethylarginine dimethylaminohydrolase (DDAH activity] and with symmetric dimethylarginine (SDMA. We investigated combined associations between transsulfuration pathway thiols, including homocysteine, and serum ADMA and SDMA concentrations at population level. METHODS: Data on clinical and demographic characteristics, medication exposure, C-reactive protein, serum ADMA and SDMA (LC-MS/MS, and thiols (homocysteine, cysteine, taurine, glutamylcysteine, total glutathione, and cysteinylglycine; capillary electrophoresis were collected from a sample of the Hunter Community Study on human ageing [n = 498, median age (IQR = 64 (60-70 years]. RESULTS: REGRESSION ANALYSIS SHOWED THAT: a age (P = 0.001, gender (P = 0.03, lower estimated glomerular filtration rate (eGFR, P = 0.08, body mass index (P = 0.008, treatment with beta-blockers (P = 0.03, homocysteine (P = 0.02, and glutamylcysteine (P = 0.003 were independently associated with higher ADMA concentrations; and b age (P = 0.001, absence of diabetes (P = 0.001, lower body mass index (P = 0.01, lower eGFR (P<0.001, cysteine (P = 0.007, and glutamylcysteine (P < 0.001 were independently associated with higher SDMA concentrations. No significant associations were observed between methylated arginines and either glutathione or taurine concentrations. CONCLUSIONS: After adjusting for clinical, demographic, biochemical, and pharmacological confounders the combined assessment of transsulfuration pathway thiols shows that glutamylcysteine has the strongest and positive independent associations with ADMA and SDMA. Whether this reflects a direct effect of glutamylcysteine on DDAH activity (for ADMA and/or cationic amino acid transport requires further investigations.

  16. Hypoxia-induced nitric oxide production and tumour perfusion is inhibited by pegylated arginine deiminase (ADI-PEG20).

    Science.gov (United States)

    Burrows, Natalie; Cane, Gaelle; Robson, Mathew; Gaude, Edoardo; J Howat, William; Szlosarek, Peter W; Pedley, R Barbara; Frezza, Christian; Ashcroft, Margaret; Maxwell, Patrick H

    2016-01-01

    The hypoxic tumour microenvironment represents an aggressive, therapy-resistant compartment. As arginine is required for specific hypoxia-induced processes, we hypothesised that arginine-deprivation therapy may be useful in targeting hypoxic cancer cells. We explored the effects of the arginine-degrading agent ADI-PEG20 on hypoxia-inducible factor (HIF) activation, the hypoxia-induced nitric oxide (NO) pathway and proliferation using HCT116 and UMUC3 cells and xenografts. The latter lack argininosuccinate synthetase (ASS1) making them auxotrophic for arginine. In HCT116 cells, ADI-PEG20 inhibited hypoxic-activation of HIF-1α and HIF-2α, leading to decreased inducible-nitric oxide synthase (iNOS), NO-production, and VEGF. Interestingly, combining hypoxia and ADI-PEG20 synergistically inhibited ASS1. ADI-PEG20 inhibited mTORC1 and activated the unfolded protein response providing a mechanism for inhibition of HIF and ASS1. ADI-PEG20 inhibited tumour growth, impaired hypoxia-associated NO-production, and decreased vascular perfusion. Expression of HIF-1α/HIF-2α/iNOS and VEGF were reduced, despite an increased hypoxic tumour fraction. Similar effects were observed in UMUC3 xenografts. In summary, ADI-PEG20 inhibits HIF-activated processes in two tumour models with widely different arginine biology. Thus, ADI-PEG20 may be useful in the clinic to target therapy-resistant hypoxic cells in ASS1-proficient tumours and ASS1-deficient tumours. PMID:26972697

  17. Preliminary X-ray crystallographic studies of yeast mitochondrial protein Tom70p

    International Nuclear Information System (INIS)

    Tom70p is an important translocase of the outer membrane complex member and a major surface receptor of the protein-translocation machinery in the outer mitochondrial membrane. To investigate the mechanism by which Tom70p functions to deliver the mitochondrial protein precursors, the cytosolic fragment of yeast Tom70p (cTom70p) has been crystallized. Protein translocations across mitochondrial membranes play critical roles in mitochondrion biogenesis. Protein transport from the cell cytosol to the mitochondrial matrix is carried out by the translocase of the outer membrane (TOM) complex and the translocase of the inner membrane (TIM) complexes. Tom70p is an important TOM-complex member and a major surface receptor of the protein-translocation machinery in the outer mitochondrial membrane. To investigate the mechanism by which Tom70p functions to deliver the mitochondrial protein precursors, the cytosolic fragment of yeast Tom70p (cTom70p) was crystallized. The crystals diffract to 3.2 Å using a synchrotron X-ray source and belong to space group P21, with unit-cell parameters a = 44.89, b = 168.78, c = 83.41 Å, α = 90.00, β = 102.74, γ = 90.00°. There are two Tom70p molecules in one asymmetric unit, which corresponds to a solvent content of approximately 51%. Structure determination by MAD methods is under way

  18. Preliminary X-ray crystallographic studies of yeast mitochondrial protein Tom70p

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Yunkun [Department of Cell Biology, Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States); McCombs, Debbie; Nagy, Lisa; DeLucas, Lawrence [Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States); Sha, Bingdong, E-mail: bdsha@uab.edu [Department of Cell Biology, Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States)

    2006-03-01

    Tom70p is an important translocase of the outer membrane complex member and a major surface receptor of the protein-translocation machinery in the outer mitochondrial membrane. To investigate the mechanism by which Tom70p functions to deliver the mitochondrial protein precursors, the cytosolic fragment of yeast Tom70p (cTom70p) has been crystallized. Protein translocations across mitochondrial membranes play critical roles in mitochondrion biogenesis. Protein transport from the cell cytosol to the mitochondrial matrix is carried out by the translocase of the outer membrane (TOM) complex and the translocase of the inner membrane (TIM) complexes. Tom70p is an important TOM-complex member and a major surface receptor of the protein-translocation machinery in the outer mitochondrial membrane. To investigate the mechanism by which Tom70p functions to deliver the mitochondrial protein precursors, the cytosolic fragment of yeast Tom70p (cTom70p) was crystallized. The crystals diffract to 3.2 Å using a synchrotron X-ray source and belong to space group P2{sub 1}, with unit-cell parameters a = 44.89, b = 168.78, c = 83.41 Å, α = 90.00, β = 102.74, γ = 90.00°. There are two Tom70p molecules in one asymmetric unit, which corresponds to a solvent content of approximately 51%. Structure determination by MAD methods is under way.

  19. Structure and function of ubiquitin: evidence for differential interactions of arginine-74 with the activating enzyme and the proteases of ATP-dependent proteolysis

    International Nuclear Information System (INIS)

    Ubiquitin was modified with the anionic, arginine-specific reagent 4-(oxoacetyl)phenoxyacetic acid in order to study the relationship between structure and function of the molecule. Four different derivatives (A, B, C, and D) were purified from the reaction mixture by anion-exchange high-performance liquid chromatography and subjected to tryptic peptide mapping to determine the location of the modification(s). These derivatives were stable throughout the procedures required for purification, tryptic hydrolysis, and peptide mapping. Derivative A was modified at arginine-42, derivative B at arginine-72, derivative C at arginines-42 and -72, and derivative D at arginine-74. Modification of ubiquitin with 14C-labeled 4-(oxoacetyl)phenoxyacetic acid indicated that the reagent formed a stable, 1:1 complex with arginine residues of the protein. Native ubiquitin and each of the four derivatives were tested for their ability to stimulate 32P exchange between ATP and pyrophosphate, a reaction catalyzed by enzyme 1 of the ubiquitin-dependent proteolytic pathway. A and C were capable of promoting this exchange at a rate only 15% that of native ubiquitin, B stimulated the exchange to 25%, and D stimulated exchange to 60% of the native level. None of the derivative was capable of promoting a significant level of ubiquitin-dependent proteolysis. These results indicate that in this system, the integrity or arginines-42, -72, and -74 is essential for full function of ubiquitin and suggest that the ubiquitin activating enzyme (E1) and the protease(s) of the system recognize different regions or conformations of ubiquitin

  20. Comparison of the effect of topical versus systemic L-arginine on wound healing in acute incisional diabetic rat model

    Directory of Open Access Journals (Sweden)

    Alireza Zandifar

    2015-01-01

    Full Text Available Background: Diabetes is associated with endothelial dysfunction and impaired wound healing. The amino acid L-arginine is the only substrate for nitric oxide (NO synthesis. The purpose of this study was to compare the topical versus systemic L-arginine treatment on total nitrite (NO x and vascular endothelial growth factor (VEGF concentrations in wound fluid and rate of wound healing in an acute incisional diabetic wound model. Materials and Methods: A total of 56 Sprague-Dawley rats were used of which 32 were rendered diabetic. Animals underwent a dorsal skin incision. Dm-sys-arg group (N = 8, diabetic and Norm-sys-arg group (N = 8, normoglycemic were gavaged with L-arginine. Dm-sys-control group (N = 8, diabetic and Norm-sys-control group (N = 8, normoglycemic were gavaged with water. Dm-top-arg group (N = 8, diabetic and norm-top-arg group (N = 8, normoglycemic received topical L-arginine gel. Dm-top-control group (N = 8, diabetic received gel vehicle. On the day 5 the amount of NO x in wound fluid was measured by Griess reaction. VEGF/total protein in wound fluids was also measured on day 5 using enzyme-linked immunosorbent assay. All wound tissue specimens were fixed and stained to be evaluated for rate of healing. Data were analyzed using SPSS software (version 18.0, Chicago, IL, USA through One-way analysis of variance test and Tukey′s post-hoc. Results: In dm-sys-arg group, the level of NO x on day 5 was significantly more than dm-top-arg group (P < 0.05. VEGF content in L-arginine treated groups were significantly more than controls (P < 0.05. Rate of diabetic wound healing in dm-sys-arg group was significantly more than dm-top-arg group. Conclusion: Systemic L-arginine is more efficient than topical L-arginine in wound healing. This process is mediated at least in part, by increasing VEGF and NO in the wound fluid.

  1. Reduced arginine availability and nitric oxide synthesis in cancer is related to impaired endogenous arginine synthesis.

    Science.gov (United States)

    Engelen, Mariëlle P K J; Safar, Ahmed M; Bartter, Thaddeus; Koeman, Fari; Deutz, Nicolaas E P

    2016-07-01

    Reduced plasma arginine (ARG) concentrations are found in various types of cancer. ARG and its product nitric oxide (NO) are important mediators in the immune function and the defense against tumour cells. It remains unclear whether the diminished systemic ARG availability in cancer is related to insufficient endogenous ARG synthesis, negatively affecting NO synthesis, and whether a dietary amino acid mixture is able to restore this. In 13 patients with advanced non-small cell lung cancer (NSCLC) and 11 healthy controls, whole body ARG and CIT (citrulline) rates of appearance were measured by stable isotope methodology before and after intake of a mixture of amino acids as present in whey protein. The conversions of CIT to ARG (indicator of de novo ARG synthesis) and ARG to CIT (marker of NO synthesis), and ARG clearance (reflecting ARG disposal capacity) were calculated. Plasma isotopic enrichments and amino acid concentrations were measured by LC-MS/MS. Conversions of CIT to ARG and ARG to CIT (P<0.05), and CIT rate of appearance (P=0.07) were lower in NSCLC. ARG rate of appearance and clearance were comparable suggesting no enhanced systemic ARG production and disposal capacity in NSCLC. After intake of the mixture, ARG rate of appearance and concentration increased (P<0.001), and ARG to CIT conversion was restored in NSCLC. In conclusion, an impaired endogenous ARG synthesis plays a role in the reduced systemic ARG availability and NO synthesis in advanced NSCLC. Nutritional approaches may restore systemic ARG availability and NO synthesis in cancer, but the clinical implication remains unclear. PMID:27129191

  2. Characterization of conserved arginine residues on Cdt1 that affect licensing activity and interaction with Geminin or Mcm complex.

    Science.gov (United States)

    You, Zhiying; Ode, Koji L; Shindo, Mayumi; Takisawa, Haruhiko; Masai, Hisao

    2016-05-01

    All organisms ensure once and only once replication during S phase through a process called replication licensing. Cdt1 is a key component and crucial loading factor of Mcm complex, which is a central component for the eukaryotic replicative helicase. In higher eukaryotes, timely inhibition of Cdt1 by Geminin is essential to prevent rereplication. Here, we address the mechanism of DNA licensing using purified Cdt1, Mcm and Geminin proteins in combination with replication in Xenopus egg extracts. We mutagenized the 223th arginine of mouse Cdt1 (mCdt1) to cysteine or serine (R-S or R-C, respectively) and 342nd and 346th arginines constituting an arginine finger-like structure to alanine (RR-AA). The RR-AA mutant of Cdt1 could not only rescue the DNA replication activity in Cdt1-depleted extracts but also its specific activity for DNA replication and licensing was significantly increased compared to the wild-type protein. In contrast, the R223 mutants were partially defective in rescue of DNA replication and licensing. Biochemical analyses of these mutant Cdt1 proteins indicated that the RR-AA mutation disabled its functional interaction with Geminin, while R223 mutations resulted in ablation in interaction with the Mcm2∼7 complex. Intriguingly, the R223 mutants are more susceptible to the phosphorylation-induced inactivation or chromatin dissociation. Our results show that conserved arginine residues play critical roles in interaction with Geminin and Mcm that are crucial for proper conformation of the complexes and its licensing activity. PMID:26940553

  3. Fragment based discovery of Arginine isosteres through REPLACE: towards non-ATP competitive CDK inhibitors

    Science.gov (United States)

    Premnath, Padmavathy Nandha; Liu, Shu; Perkins, Tracy; Abbott, Jennifer; Anderson, Erin; McInnes, Campbell

    2013-01-01

    In order to develop non-ATP competitive CDK2/cyclin A inhibitors, the REPLACE strategy has been applied to generate fragment alternatives for the N-terminal tetrapeptide of the cyclin binding motif (HAKRRLIF) involved in substrate recruitment prior to phosphotransfer. The docking approach used for the prediction of small molecule mimics for peptide determinants was validated through reproduction of experimental binding modes of known inhibitors and provides useful information for evaluating binding to protein-protein interaction sites. Further to this, potential arginine isosteres predicted using the validated LigandFit docking method were ligated to the truncated C-terminal peptide, RLIF using solid phase synthesis and evaluated in a competitive binding assay. After testing, identified fragments were shown to represent not only appropriate mimics for a critical arginine residue but also to interact effectively with a minor hydrophobic pocket present in the binding groove. Further evaluation of binding modes was undertaken to optimize the potency of these compounds. Through further application of the REPLACE strategy in this study, peptide-small molecule hybrid CDK2 inhibitors were identified that are more drug-like and suitable for further optimization as anti-tumor therapeutics. PMID:24286762

  4. Arginine protection against ammonia toxicity in exhausted rat.

    Science.gov (United States)

    Krishna Mohan, P; Indira, K; Rajendra, W

    1987-01-01

    Arginine administration (5 m moles/kg/day) to albino rats for 7 days, revealed that this vital basic amino acid possesses latent potentiality for the accentuation of urea cycle or at least for arginase activity. The mitigation of ammonia toxicity was observed to be more effective in the case of gastrocnemius and red vastus as compared to white vastus. Further, ammonia and lactate levels were also decreased by arginine in blood and thereby delaying the onset of fatigue by preventing ammonotoxemia and lactic acidemia. PMID:3666875

  5. Glutamine Assimilation and Feedback Regulation of L-acetyl-N-glutamate Kinase Activity in Chlorella variabilis NC64A Results in Changes in Arginine Pools.

    Science.gov (United States)

    Minaeva, Ekaterina; Forchhammer, Karl; Ermilova, Elena

    2015-11-01

    Glutamine is a metabolite of central importance in nitrogen metabolism of microorganisms and plants. The Chlorella PII signaling protein controls, in a glutamine-dependent manner, the key enzyme of the ornithine/arginine biosynthesis pathway, N-acetyl-L-glutamate kinase (NAGK) that leads to arginine formation. We provide evidence that glutamine promotes effective growth of C. variabilis strain NC64A. The present study shows that externally supplied glutamine directly influences the internal pool of arginine in NC64A. Glutamine synthetase (GS) catalyzes the ATP-dependent conversion of glutamate and ammonium to glutamine. The results of this study demonstrate that glutamine acts as a negative effector of GS activity. These data emphasize the importance of glutamine-dependent coupling of metabolism and signaling as components of an efficient pathway allowing the maintenance of metabolic homeostasis and sustaining growth of Chlorella. PMID:26356535

  6. The Arginine/ADMA Ratio Is Related to the Prevention of Atherosclerotic Plaques in Hypercholesterolemic Rabbits When Giving a Combined Therapy with Atorvastatine and Arginine

    Directory of Open Access Journals (Sweden)

    Saskia J. H. Brinkmann

    2015-05-01

    Full Text Available Supplementation with arginine in combination with atorvastatin is more efficient in reducing the size of an atherosclerotic plaque than treatment with a statin or arginine alone in homozygous Watanabe heritable hyperlipidemic (WHHL rabbits. We evaluated the mechanism behind this feature by exploring the role of the arginine/asymmetric dimethylarginine (ADMA ratio, which is the substrate and inhibitor of nitric oxide synthase (NOS and thereby nitric oxide (NO, respectively. Methods: Rabbits were fed either an arginine diet (group A, n = 9, standard rabbit chow plus atorvastatin (group S, n = 8, standard rabbit chow plus an arginine diet with atorvastatin (group SA, n = 8 or standard rabbit chow (group C, n = 9 as control. Blood was sampled and the aorta was harvested for topographic and histological analysis. Plasma levels of arginine, ADMA, cholesterol and nitric oxide were determined and the arginine/ADMA ratio was calculated. Results: The decrease in ADMA levels over time was significantly correlated to fewer aortic lesions in the distal aorta and total aorta. The arginine/ADMA ratio was correlated to cholesterol levels and decrease in cholesterol levels over time in the SA group. A lower arginine/ADMA ratio was significantly correlated to lower NO levels in the S and C group. Discussion: A balance between arginine and ADMA is an important indicator in the prevention of the development of atherosclerotic plaques.

  7. Improvement of L-arginine production by overexpression of a bifunctional ornithine acetyltransferase in Corynebacterium crenatum.

    Science.gov (United States)

    Dou, Wenfang; Xu, Meijuan; Cai, Dongmei; Zhang, Xiaomei; Rao, Zhiming; Xu, Zhenghong

    2011-10-01

    Ornithine acetyltransferase (EC 2.3.1.35; OATase) gene (argJ) from the L-arginine-producing mutant Corynebacterium crenatum SYPA5-5 was cloned, sequenced, and expressed in Escherichia coli BL21 (DE3). Analysis of the argJ sequence revealed that the argJ coded a polypeptide of 388 amino acids with a calculated molecular weight of 39.7 kDa. In this study, the function of the OATase (argJ) of C. crenatum SYPA5-5 has been identified as a conserved ATML sequence for the autolysis of the protein to α- and β-subunits. When the argJ regions corresponding to the α- and β-subunits were cloned and expressed separately in E. coli BL21, OATase activities were abolished. At the same time, a functional study revealed that OATase from C. crenatum SYPA5-5 was a bifunctional enzyme with the functions of acetylglutamate synthase (EC 2.3.1.1, NAGS) and acetylornithine deacetylase (EC 3.5.1.16, AOase) activities. In order to investigate the effects of the overexpression of the argJ gene on L: -arginine production, the argJ gene was inserted into pJCtac to yield the recombinant shuttle plasmid pJCtac-argJ and then transformed into C. crenatum SYPA5-5. The results showed that the engineered strains could not only express more OATase (90.9%) but also increase the production of L: -arginine significantly (16.8%). PMID:21785983

  8. Effect of oral L-arginine administration on exhaled nitric oxide (no) concentration in healthy volunteers

    OpenAIRE

    Ogata, Hiroshi; Yatabe, Midori; Misaka, Shingen; Shikama, Yayoi; Sato, Suguru; Munakata, Mitsuru; Kimura, Junko

    2013-01-01

    We previously reported a case of pulmonary hypertension, where the symptoms were improved by oral L-arginine (arginine) administration. Arginine may increase nitric oxide (NO) production in the pulmonary artery. Exhaled NO may reflect pulmonary artery NO production. It has been demonstrated that exhaled NO concentration is higher in patients with allergic diseases, but whether oral arginine administration alters exhaled NO is unknown. Therefore, in this study, we investigated whether oral arg...

  9. Interplay among coactivator-associated arginine methyltransferase 1, CBP, and CIITA in IFN-gamma-inducible MHC-II gene expression.

    Science.gov (United States)

    Zika, Eleni; Fauquier, Lucas; Vandel, Laurence; Ting, Jenny P-Y

    2005-11-01

    Class II major histocompatibility (MHC-II) genes are prototype targets of IFN-gamma. IFN-gamma activates the expression of the non-DNA-binding master regulator of MHC-II, class II transactivator (CIITA), which is crucial for enhanceosome formation and gene activation. This report shows the importance of the histone methyltransferase, coactivator-associated arginine methyltransferase (CARM1/PRMT4), during IFN-gamma-induced MHC-II gene activation. It also demonstrates the coordinated regulation of CIITA, CARM1, and the acetyltransferase cyclic-AMP response element binding (CREB)-binding protein (CBP) during this process. CARM1 synergizes with CIITA in activating MHC-II transcription and synergy is abrogated when an arginine methyltransferase-defective CARM1 mutant is used. Protein-arginine methyltransferase 1 has much less effect on MHC-II transcription. Specific RNA interference reduced CARM1 expression as well as MHC-II expression. The recruitment of CARM1 to the promoter requires endogenous CIITA and results in methylation of histone H3-R17; hence, CIITA is an upstream regulator of histone methylation. Previous work has shown that CARM1 can methylate CBP at three arginine residues. Using wild-type CBP and a mutant of CBP lacking the CARM1-targeted arginine residues (R3A), we show that arginine methylation of CBP is required for IFN-gamma induction of MHC-II. A kinetic analysis shows that CIITA, CARM1, and H3-R17 methylation all precede CBP loading on the MHC-II promoter during IFN-gamma treatment. These results suggest functional and temporal relationships among CIITA, CARM1, and CBP for IFN-gamma induction of MHC-II. PMID:16254053

  10. Arginine Functionally Improves Clinically Relevant Human Galactose-1-Phosphate Uridylyltransferase (GALT) Variants Expressed in a Prokaryotic Model.

    Science.gov (United States)

    Coelho, Ana I; Trabuco, Matilde; Silva, Maria João; de Almeida, Isabel Tavares; Leandro, Paula; Rivera, Isabel; Vicente, João B

    2015-01-01

    Classic galactosemia is a rare genetic disease of the galactose metabolism, resulting from deficient activity of galactose-1-phosphate uridylyltransferase (GALT). The current standard of care is lifelong dietary restriction of galactose, which however fails to prevent the development of long-term complications. Structural-functional studies demonstrated that the most prevalent GALT mutations give rise to proteins with increased propensity to aggregate in solution. Arginine is a known stabilizer of aggregation-prone proteins, having already shown a beneficial effect in other inherited metabolic disorders.Herein we developed a prokaryotic model of galactose sensitivity that allows evaluating in a cellular context the mutations' impact on GALT function, as well as the potential effect of arginine in functionally rescuing clinically relevant variants.This study revealed that some hGALT variants, previously described to exhibit no detectable activity in vitro, actually present residual activity when determined in vivo. Furthermore, it revealed that arginine presents a mutation-specific beneficial effect, particularly on the prevalent p.Q188R and p.K285N variants, which led us to hypothesize that it might constitute a promising therapeutic agent in classic galactosemia. PMID:25814382

  11. Arginine Patch Predicts the RNA Annealing Activity of Hfq from Gram-Negative and Gram-Positive Bacteria.

    Science.gov (United States)

    Zheng, Amy; Panja, Subrata; Woodson, Sarah A

    2016-06-01

    The Sm-protein Hfq facilitates interactions between small non-coding RNA (sRNA) and target mRNAs. In enteric Gram-negative bacteria, Hfq is required for sRNA regulation, and hfq deletion results in stress intolerance and reduced virulence. By contrast, the role of Hfq in Gram-positive is less established and varies among species. The RNA binding and RNA annealing activity of Hfq from Escherichia coli, Pseudomonas aeruginosa, Listeria monocytogenes, Bacillus subtilis, and Staphylococcus aureus were compared using minimal RNAs and fluorescence spectroscopy. The results show that RNA annealing activity increases with the number of arginines in a semi-conserved patch on the rim of the Hfq hexamer and correlates with the previously reported requirement for Hfq in sRNA regulation. Thus, the amino acid sequence of the arginine patch can predict the chaperone function of Hfq in sRNA regulation in different organisms. PMID:27049793

  12. In vivo arginine production and intravascular nitric oxide synthesis in hypotensive sepsis

    Science.gov (United States)

    Arginine is important in the response to infections and is a precursor for the synthesis of the vasodilator nitric oxide (NO). Low plasma arginine is correlated with a worse prognosis in patients with sepsis, and increased NO has been implicated in the hypotension of sepsis. Data on in vivo arginine...

  13. Symmetric Allosteric Mechanism of Hexameric Escherichia coli Arginine Repressor Exploits Competition between L-Arginine Ligands and Resident Arginine Residues

    Czech Academy of Sciences Publication Activity Database

    Strawn, R.; Melicherčík, Milan; Green, M.; Stockner, T.; Carey, J.; Ettrich, Rüdiger

    2010-01-01

    Roč. 6, č. 6 (2010), s. 1-12. ISSN 1553-734X R&D Projects: GA MŠk(CZ) LC06010; GA ČR GAP207/10/1934 Institutional research plan: CEZ:AV0Z60870520 Keywords : molecular-dynamics simulations * free-energy calculations * structural basis * DNA-binding domain * bacillus -stearothermophilus * T4 lysozyme * proteins * hemoglobin * model * affinity Subject RIV: EH - Ecology, Behaviour Impact factor: 5.515, year: 2010

  14. Effect of iron, taurine and arginine on rat hepatic fibrosis

    International Nuclear Information System (INIS)

    Objective: The promotion role of iron on pathogenesis of hepatic fibrosis and the protective role of taurine and L-arginine against hepatic fibrosis were studied. Method: The model of rat radiation hepatic fibrosis was used. Experimental rats were divided into 0 Gy, 30 Gy, 30 Gy + iron, 30 Gy + taurine and 30 Gy + L-arginine groups. Serum iron, liver tissue hydroxyproline (Hyp) and malondialdehyde (MDA) were measured one and three months respectively after irradiation of hepatic tissue, production and distribution characteristics of hepatic tissue type I and III collagen were observed with a polarizing microscope. Results: Administration of iron agent could significantly increase hepatic tissue MDA content and serum iron concentration, one month after irradiation, hepatic tissue Hyp in 30 Gy + iron group began to increase, and collagen in hepatic tissue obviously increased. Taurine and L-arginine could reduce serum iron concentration and decrease production of hepatic fissure Hyp. Conclusion: Exogenous iron agent could promote early development of radiation hepatic fibrosis; taurine and arginine could diminish pathologic alteration of hepatic fibrosis to a certain extent

  15. Safety, efficacy and physiological actions of a lysine-free, arginine-rich formula to treat glutaryl-CoA dehydrogenase deficiency: focus on cerebral amino acid influx.

    Science.gov (United States)

    Strauss, Kevin A; Brumbaugh, Joan; Duffy, Alana; Wardley, Bridget; Robinson, Donna; Hendrickson, Christine; Tortorelli, Silvia; Moser, Ann B; Puffenberger, Erik G; Rider, Nicholas L; Morton, D Holmes

    2011-01-01

    Striatal degeneration from glutaryl-CoA dehydrogenase deficiency (glutaric aciduria type 1, GA1) is associated with cerebral formation and entrapment of glutaryl-CoA and its derivatives that depend on cerebral lysine influx. In 2006 we designed a lysine-free study formula enriched with arginine to selectively block lysine transport across cerebral endothelia and thereby limit glutaryl-CoA production by brain. Between 2006 and present, we treated twelve consecutive children with study formula (LYSx group) while holding all other treatment practices constant. Clinical and biochemical outcomes were compared to 25 GA1 patients (PROx group) treated between 1995 and 2005 with natural protein restriction (dietary lysine/arginine ratio of 1.7±0.3 mg:mg). We used published kinetic parameters of the y+and LAT1 blood-brain barrier transporters to model the influx of amino acids into the brain. Arginine fortification to achieve a mean dietary lysine/arginine ratio of 0.7±0.2 mg:mg was neuroprotective. All 12 LYSx patients are physically and neurologically healthy after 28 aggregate patient-years of follow up (current ages 28±21 months) and there were no adverse events related to formula use. This represents a 36% reduction of neurological risk (95% confidence interval 14-52%, p=0.018) that we can directly attribute to altered amino acid intake. During the first year of life, 20% lower lysine intake and two-fold higher arginine intake by LYSx patients were associated with 50% lower plasma lysine, 3-fold lower plasma lysine/arginine concentration ratio, 42% lower mean calculated cerebral lysine influx, 54% higher calculated cerebral arginine influx, 15-26% higher calculated cerebral influx of several anaplerotic precursors (isoleucine, threonine, methionine, and leucine), 50% less 3-hydroxyglutarate excretion, and a 3-fold lower hospitalization rate (0.8 versus 2.3 hospitalizations per patient per year). The relationship between arginine fortification and plasma lysine

  16. L-Arginine improves multiple physiological parameters in mice exposed to diet-induced metabolic disturbances

    DEFF Research Database (Denmark)

    Clemmensen, Christoffer; Madsen, Andreas Nygaard; Smajilovic, Sanela;

    2012-01-01

    L: -Arginine (L: -Arg) is a conditionally essential amino acid and a natural constituent of dietary proteins. Studies in obese rats and type 2 diabetic humans have indicated that dietary supplementation with L: -Arg can diminish gain in white adipose tissue (WAT) and improve insulin sensitivity....... However, the effects of L: -Arg on glucose homeostasis, body composition and energy metabolism remain unclear. In addition, no studies have, to our knowledge, examined whether L: -Arg has beneficial effects as a dietary supplement in the mouse model. In the present study, we investigated the effects of L......: -Arg supplementation to male C57BL/6 mice on an array of physiological parameters. L: -Arg supplemented mice were maintained on a low-protein diet and body composition, appetite regulation, glucose tolerance, insulin sensitivity and energy expenditure were evaluated. A significant reduction in...

  17. Secretory production of an FAD cofactor-containing cytosolic enzyme (sorbitol–xylitol oxidase from Streptomyces coelicolor) using the twin-arginine translocation (Tat) pathway of Corynebacterium glutamicum

    OpenAIRE

    Scheele, S.; Oertel, Dan; Bongaerts, Johannes; Evers, Stefan; Hellmuth, Hendrik; Maurer, Karl-Heinz; Bott, Michael; Freudl, Roland

    2012-01-01

    Carbohydrate oxidases are biotechnologically interesting enzymes that require a tightly or covalently bound cofactor for activity. Using the industrial workhorse Corynebacterium glutamicum as the expression host, successful secretion of a normally cytosolic FAD cofactor-containing sorbitol–xylitol oxidase from Streptomyces coelicolor was achieved by using the twin-arginine translocation (Tat) protein export machinery for protein translocation across the cytoplasmic membrane. Our results demon...

  18. Salmonella Type III Secretion-Associated Protein InvE Controls Translocation of Effector Proteins into Host Cells

    OpenAIRE

    Kubori, Tomoko; Galán, Jorge E.

    2002-01-01

    Salmonella enterica encodes a type III secretion system (TTSS) within a pathogenicity island located at centisome 63 (SPI-1), which is essential for its pathogenicity. This system mediates the transfer of a battery of bacterial proteins into the host cell with the capacity to modulate cellular functions. The transfer process is dependent on the function of protein translocases SipB, SipC, and SipD. We report here that Salmonella protein InvE, which is also encoded within SPI-1, is essential f...

  19. Data supporting the involvement of the adenine nucleotide translocase conformation in opening the Tl(+)-induced permeability transition pore in Ca(2+)-loaded rat liver mitochondria.

    Science.gov (United States)

    Korotkov, Sergey M

    2016-06-01

    There we made available information about the effects of the adenine nucleotide translocase (ANT) 'c' conformation fixers (phenylarsine oxide (PAO), tert-butylhydroperoxide (tBHP), and carboxyatractyloside) as well as thiol reagent (4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS)) on isolated rat liver mitochondria. We observed a decrease in A540 (mitochondrial swelling) and respiratory control rates (RCRADP [state 3/state 4] and RCRDNP [2,4-dinitrophenol-uncoupled state/basal state or state 4]), as well as an increase in Ca(2+)-induced safranin fluorescence (F485/590, arbitrary units), showed a dissipation in the inner membrane potential (ΔΨmito), in experiments with energized rat liver mitochondria, injected into the buffer containing 25-75 mM TlNO3, 125 mM KNO3, and 100 µM Ca(2+). The fixers and DIDS, in comparison to Ca(2+) alone, greatly increased A540 decline and the rate of Ca(2+)-induced ΔΨmito dissipation. These reagents also markedly decreased RCRADP and RCRDNP. The MPTP inhibitors (ADP, cyclosporin A, bongkrekic acid, and N-ethylmaleimide) fixing the ANT in 'm' conformation significantly hindered the above-mentioned effects of the fixers and DIDS. A more complete scientific analysis of these findings may be obtained from the manuscript "To involvement the conformation of the adenine nucleotide translocase in opening the Tl(+)-induced permeability transition pore in Ca(2+)-loaded rat liver mitochondria" (Korotkov et al., 2016 [1]). PMID:27054168

  20. Analysis of an Alanine/Arginine Mixture by Using TLC/FTIR Technique

    Directory of Open Access Journals (Sweden)

    Jun Liu

    2014-01-01

    Full Text Available We applied TLC/FTIR coupled with mapping technique to analyze an alanine/arginine mixture. Narrow band TLC plates prepared by using AgI as a stationary phase were used to separate alanine and arginine. The distribution of alanine and arginine spots was manifested by a 3D chromatogram. Alanine and arginine can be successfully separated by the narrow band TLC plate. In addition, the FTIR spectra of the separated alanine and arginine spots on the narrow band TLC plate are roughly the same as the corresponding reference IR spectra.

  1. Expression of arg genes of Escherichia coli during arginine limitation dependent upon stringent control of translation.

    OpenAIRE

    Williams, M.G.; Rogers, P

    1987-01-01

    The transcription and translation of operons for arginine biosynthetic enzymes after arginine removal (arginine down shift) were studied in relA and relA+ strains of Escherichia coli. After arginine down shift, derepression of synthesis of the arginine biosynthetic enzymes ornithine carbamoyltransferase (argF) and argininosuccinate lyase (argH) began at about 15 min in relA+ cells but was delayed in relA cells for more than 2 h. However, both relA+ and relA cells accumulated high levels of ar...

  2. Differential effects of arginine, glutamate and phosphoarginine on Ca(2+)-activation properties of muscle fibres from crayfish and rat.

    Science.gov (United States)

    Jame, David W; West, Jan M; Dooley, Philip C; Stephenson, D George

    2004-01-01

    The effects of two amino acids, arginine which has a positively charged side-chain and glutamate which has a negatively charged side-chain on the Ca2+-activation properties of the contractile apparatus were examined in four structurally and functionally different types of skeletal muscle; long- and short-sarcomere fibres from the claw muscle of the yabby (a freshwater decapod crustacean), and fast- and slow-twitch fibres from limb muscles of the rat. Single skinned fibres were activated in carefully balanced solutions of different pCa (-log10[Ca2+]) that either contained the test solute ("test") or not ("control"). The effect of phosphoarginine, a phosphagen that bears a nett negative charge, was also compared to the effects of arginine. Results show that (i) arginine (33-36 mmol l(-1)) significantly shifted the force-pCa curve by 0.08-0.13 pCa units in the direction of increased sensitivity to Ca2+-activated contraction in all fibre types; (ii) phosphoarginine (9-10 mmol l(-1)) induced a significant shift of the force-pCa curve by 0.18-0.24 pCa units in the direction of increased sensitivity to Ca2+ in mammalian fast- and slow-twitch fibres, but had no significant effects on the force-pCa relation in either long- or short-sarcomere crustacean fibres; (iii) glutamate (36-40 mmol l(-1)), like arginine affected the force-pCa relation of all fibre types investigated, but in the opposite direction, causing a significant decrease in the sensitivity to Ca2+-activated contraction by 0.08-0.19 pCa units; (iv) arginine, phosphoarginine and glutamate had little or no effect on the maximum Ca2+-activated force of crustacean and mammalian fibres. The results suggest that the opposing effects of glutamate and arginine are not related to simply their charge structure, but must involve complex interactions between these molecules, Ca2+ and the regulatory and other myofibrillar proteins. PMID:15711880

  3. A rare disease-associated mutation in the medium-chain acyl-CoA dehydrogenase (MCAD) gene changes a conserved arginine, previously shown to be functionally essential in short-chain acyl-CoA dehydrogenase (SCAD)

    DEFF Research Database (Denmark)

    Andresen, B S; Bross, P; Jensen, T G;

    1993-01-01

    . On the basis of knowledge about the three-dimensional structure of the MCAD protein, we suggest that the mutation destroys a salt bridge between arginine28 and glutamate86, thereby affecting the formation of enzymatically active protein. Twenty-two additional families with compound heterozygous patients were...

  4. Enthalpy-driven interactions with sulfated glycosaminoglycans promote cell membrane penetration of arginine peptides.

    Science.gov (United States)

    Takechi-Haraya, Yuki; Nadai, Ryo; Kimura, Hitoshi; Nishitsuji, Kazuchika; Uchimura, Kenji; Sakai-Kato, Kumiko; Kawakami, Kohsaku; Shigenaga, Akira; Kawakami, Toru; Otaka, Akira; Hojo, Hironobu; Sakashita, Naomi; Saito, Hiroyuki

    2016-06-01

    The first step of cell membrane penetration of arginine peptides is thought to occur via electrostatic interactions between positive charges of arginine residues and negative charges of sulfated glycosaminoglycans (GAGs) on the cell surface. However, the molecular interaction of arginine peptides with GAG still remains unclear. Here, we compared the interactions of several arginine peptides of Tat, R8, and Rev and their analogues with heparin in relation to the cell membrane penetration efficiency. The high-affinity binding of arginine peptides to heparin was shown to be driven by large favorable enthalpy contributions, possibly reflecting multidentate hydrogen bondings of arginine residues with sulfate groups of heparin. Interestingly, the lysine peptides in which all arginine residues are substituted with lysine residues exhibited negligible binding enthalpy despite of their considerable binding to heparin. In CHO-K1 cells, arginine peptides exhibited a great cell-penetrating ability whereas their corresponding lysine peptides did not penetrate into cells. The degree of cell penetration of arginine peptides markedly decreased by the chlorate treatment of cells which prevents the sulfation of GAG chains. Significantly, the cell penetration efficiency of arginine peptides was found to be correlated with the favorable enthalpy of binding to heparin. These results suggest that the enthalpy-driven strong interaction with sulfated GAGs such as heparan sulfate plays a critical role in the efficient cell membrane penetration of arginine peptides. PMID:27003128

  5. Effect of l-arginine therapy on plasma NO/sub 2/ and NO/sub 3/ levels, and blood pressure in uremic rabbits

    International Nuclear Information System (INIS)

    Background: Normal kidney function is regulated by Nitric oxide (NO) and Superoxide (O/sub 2/-) in the body, and consequently controls blood pressure. Nitric Oxide promotes natriuresis and diuresis, and therefore results in reduction of blood pressure. The objective of this study was to determine the effect of L-arginine supplementation on blood pressure, urinary protein, nitrite and nitrate in addition to blood urea, serum creatinine and creatinine clearance in uremic rabbits. Methods: This study was carried out in the Department of Biochemistry Basic Medical Sciences Institute, Jinnah Postgraduate Medical Centre, Karachi. A total of 48 rabbits were included in the study. Twenty-four of the rabbits on surgical intervention were prepared as uremic and so became hypertensive as well. Two groups were uremic, one group was given L-arginine and the other was remained untreated. Systolic and diastolic blood pressure was measured on week 0, week 2, week 4, and week 6, while blood and urine was collected on week 0 and week 6. Results: On supplementation with L-arginine to uremic rabbits systolic and diastolic blood pressures were decreased significantly. Nitrite/nitrate and urinary protein were corrected to some extent while blood urea and serum creatinine were unaffected. Conclusion: L-arginine has a beneficial role as blood pressure lowering agent in uremic rabbits. It corrects NO/sub 2/NO/sub 3/ plasma level and proteinuria which is indicator of renal failure. (author)

  6. The N-acetylglutamate synthase/N-acetylglutamate kinase metabolon of Saccharomyces cerevisiae allows co-ordinated feedback regulation of the first two steps in arginine biosynthesis.

    Science.gov (United States)

    Pauwels, Katia; Abadjieva, Agnes; Hilven, Pierre; Stankiewicz, Anna; Crabeel, Marjolaine

    2003-03-01

    In Saccharomyces cerevisiae, which uses the nonlinear pathway of arginine biosynthesis, the first two enzymes, N-acetylglutamate synthase (NAGS) and N-acetylglutamate kinase (NAGK), are controlled by feedback inhibition. We have previously shown that NAGS and NAGK associate in a complex, essential to synthase activity and protein level [Abadjieva, A., Pauwels, K., Hilven, P. & Crabeel, M. (2001) J. Biol. Chem.276, 42869-42880]. The NAGKs of ascomycetes possess, in addition to the catalytic domain that is shared by all other NAGKs and whose structure has been determined, a C-terminal domain of unknown function and structure. Exploring the role of these two domains in the synthase/kinase interaction, we demonstrate that the ascomycete-specific domain is required to maintain synthase activity and protein level. Previous results had suggested a participation of the third enzyme of the pathway, N-acetylglutamylphosphate reductase, in the metabolon. Here, genetic analyses conducted in yeast at physiological level, or in a heterologous background, clearly demonstrate that the reductase is dispensable for synthase activity and protein level. Most importantly, we show that the arginine feedback regulation of the NAGS and NAGK enzymes is mutually interdependent. First, the kinase becomes less sensitive to arginine feedback inhibition in the absence of the synthase. Second, and as in Neurospora crassa, in a yeast kinase mutant resistant to arginine feedback inhibition, the synthase becomes feedback resistant concomitantly. We conclude that the NAGS/NAGK metabolon promotes the co-ordination of the catalytic activities and feedback regulation of the first two, flux controlling, enzymes of the arginine pathway. PMID:12603335

  7. Protective effect of quercetin and/or l-arginine against nano-zinc oxide-induced cardiotoxicity in rats

    Energy Technology Data Exchange (ETDEWEB)

    Faddah, L. M.; Baky, Nayira A. Abdel [King Saud University, Pharmacology Department, Faculty of Pharmacy (Saudi Arabia); Mohamed, Azza M., E-mail: azzamohamed99@yahoo.com [King Abdulaziz University, Biochemistry Department, Faculty of Science for Girls (Saudi Arabia); Al-Rasheed, Nouf M.; Al-Rasheed, Nawal M. [King Saud University, Pharmacology Department, Faculty of Pharmacy (Saudi Arabia)

    2013-04-15

    The aim of this study was to investigate the protective role of quercetin and/or l-arginine against the cardiotoxic potency of zinc oxide nanoparticle (ZnO-NP)-induced cardiac infarction. ZnO-NPs (50 nm) were administered orally at either 600 mg or 1 g/kg body weight for 5 consecutive days. The results revealed that co-administration of quercetin and/or l-arginine (each 200 mg/kg body weight) daily for 3 weeks to rats intoxicated by either of the two doses markedly ameliorated increases in serum markers of cardiac infarction, including troponin T, creatine kinase-MB, and myoglobin, as well as increases in proinflammatory biomarkers, including tumor necrosis factor-{alpha}, interleukin-6, and C-reactive protein, compared with intoxicated, untreated rats. Each agent alone or in combination also successfully modulated the alterations in serum vascular endothelial growth factor, cardiac calcium concentration, and oxidative DNA damage as well as the increase in the apoptosis marker caspase 3 of cardiac tissue in response to ZnO-NP toxicity. In conclusion, early treatment with quercetin and l-arginine may protect cardiac tissue from infarction induced by the toxic effects of ZnO-NPs.

  8. Effects of nutritional supplementation with l-arginine on repair of injuries due to muscle strain: experimental study on rats

    Directory of Open Access Journals (Sweden)

    Lauren Izabel Medeiros Couto

    2015-08-01

    Full Text Available ABSTRACTOBJECTIVE: To evaluate the influence of oral supplementation with arginine on regeneration of injuries due to straining of the anterior tibial muscle of rats. METHODS: Twenty-four Wistar rats of weight 492.5 ± 50.45 g were used. Injuries were induced through straining the anterior tibial muscles. The rats were separated into three groups of eight rats each. In the untreated group (UTG, after induction of injuries, the rats were observed for 24 h. In the simulation group (SG and the arginine group (AG respectively, the rats received isotonic saline solution and arginine solution via direct gavage, over a seven-day period. At the end of the period, blood samples were collected for serum evaluations of creatine kinase (CK, lactic dehydrogenase (LDH, aspartate aminotransferase (AST and C-reactive protein (CRP. The right and left anterior tibial muscles were resected for histopathological evaluations on the muscle injuries, investigating edema, hemorrhage and disorganization or morphometric alteration of the muscle fibers. The tissue repair was investigated in terms of proliferation of adipose tissue, angiogenesis and collagen fibers. The ANOVA and Student's tmethods were used and p≤ 0.05 was taken to be statistically significant. RESULTS: In the serum evaluations, the AG showed lower CK assay values and higher AST values. In the histopathological evaluation, the UTG presented edema and hemorrhage compatible with injuries due to strain; the SG presented edema and hemorrhage with proliferation of adipose tissue and collagen fibers; and the AG presented not only the findings of the SG but also, especially, intense angiogenesis. CONCLUSION: Oral supplementation with arginine did not cause any significant metabolic alterations that would contraindicate its use and it induced angiogenesis during the repair of muscles injured due to strain.

  9. Mutation of the Erwinia amylovora argD gene causes arginine auxotrophy, nonpathogenicity in apples, and reduced virulence in pears.

    Science.gov (United States)

    Ramos, Laura S; Lehman, Brian L; Peter, Kari A; McNellis, Timothy W

    2014-11-01

    Fire blight is caused by Erwinia amylovora and is the most destructive bacterial disease of apples and pears worldwide. In this study, we found that E. amylovora argD(1000)::Tn5, an argD Tn5 transposon mutant that has the Tn5 transposon inserted after nucleotide 999 in the argD gene-coding region, was an arginine auxotroph that did not cause fire blight in apple and had reduced virulence in immature pear fruits. The E. amylovora argD gene encodes a predicted N-acetylornithine aminotransferase enzyme, which is involved in the production of the amino acid arginine. A plasmid-borne copy of the wild-type argD gene complemented both the nonpathogenic and the arginine auxotrophic phenotypes of the argD(1000)::Tn5 mutant. However, even when mixed with virulent E. amylovora cells and inoculated onto immature apple fruit, the argD(1000)::Tn5 mutant still failed to grow, while the virulent strain grew and caused disease. Furthermore, the pCR2.1-argD complementation plasmid was stably maintained in the argD(1000)::Tn5 mutant growing in host tissues without any antibiotic selection. Therefore, the pCR2.1-argD complementation plasmid could be useful for the expression of genes, markers, and reporters in E. amylovora growing in planta, without concern about losing the plasmid over time. The ArgD protein cannot be considered an E. amylovora virulence factor because the argD(1000)::Tn5 mutant was auxotrophic and had a primary metabolism defect. Nevertheless, these results are informative about the parasitic nature of the fire blight disease interaction, since they indicate that E. amylovora cannot obtain sufficient arginine from apple and pear fruit tissues or from apple vegetative tissues, either at the beginning of the infection process or after the infection has progressed to an advanced state. PMID:25172854

  10. Effects of non-surgical periodontal treatment on the L-arginine-nitric oxide pathway and oxidative status in platelets.

    Science.gov (United States)

    Siqueira, Mariana Alves de Sá; Fischer, Ricardo Guimarães; Pereira, Natália Rodrigues; Martins, Marcela Anjos; Moss, Monique Bandeira; Mendes-Ribeiro, Antônio Cláudio; Figueredo, Carlos Marcelo da Silva; Brunini, Tatiana Marlowe Cunha

    2013-06-01

    Several studies have suggested an increase of cardiovascular disease (CVD) risk on periodontitis patients. An enhancement has been demonstrated on both platelet activation and oxidative stress on periodontitis patients, which may contribute for this association. Therefore, the aim of this study was to evaluate the effects of non-surgical periodontal treatment on the l-arginine-nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) pathway and oxidative status in platelets. A total of eight periodontitis patients and eight controls were included in this study. Clinical, laboratory and experimental evaluations were performed on baseline and 90 days after periodontal treatment (except for western blot analysis). The clinical periodontal evaluation included measurements of probing pocket depth (PPD), clinical attachment loss (CAL), % of sites with plaque and % of sites with bleeding on probing. We evaluated: l-[(3)H]arginine influx; nitric oxide synthase (NOS) and arginase enzymes activity and expression; expression of guanylate cyclase and phosphodiesterase-5 enzymes; cGMP levels; platelet aggregation; oxidative status through superoxide dismutase (SOD) and catalase activities, and measurement of reactive oxygen species (ROS) levels and C-reactive protein (CRP) levels. The initial results showed an activation of both l-arginine influx and via system y (+ )L associated with reduced intraplatelet cGMP levels in periodontitis patients and increased systemic levels of CRP. After periodontal treatment, there was a significant reduction of the % of sites with PPD 4-5mm, % of sites with CAL 4-5 mm, and an enhancement in cGMP levels and SOD activity. Moreover, CRP levels were reduced after treatment. Therefore, alterations in the intraplatelet l-arginine-NO-cGMP pathway and oxidant-antioxidant balance associated with a systemic inflammatory response may lead to platelet dysfunction, which may contribute to a higher risk of CVD in periodontitis. PMID:23918883

  11. Prediction of twin-arginine signal peptides

    DEFF Research Database (Denmark)

    Bendtsen, Jannick Dyrløv; Nielsen, Henrik; Widdick, D.; Palmer, T.; Brunak, Søren

    2005-01-01

    publicly available method, TatP, for prediction of bacterial Tat signal peptides. Results: We have retrieved sequence data for Tat substrates in order to train a computational method for discrimination of Sec and Tat signal peptides. The TatP method is able to positively classify 91% of 35 known Tat signal...... complementary rule based prediction method. Conclusion: The method developed here is able to discriminate Tat signal peptides from cytoplasmic proteins carrying a similar motif, as well as from Sec signal peptides, with high accuracy. The method allows filtering of input sequences based on Perl syntax regular...... expressions, whereas hydrophobicity discrimination of Tat- and Sec- signal peptides is carried out by an artificial neural network. A potential cleavage site of the predicted Tat signal peptide is also reported. The TatP prediction server is available as a public web server at http://www.cbs.dtu.dk/services/TatP/....

  12. Gliclazide directly inhibits arginine-induced glucagon release

    DEFF Research Database (Denmark)

    Cejvan, Kenan; Coy, David H; Holst, Jens Juul;

    2002-01-01

    Arginine-stimulated insulin and somatostatin release is enhanced by the sulfonylurea gliclazide. In contrast, gliclazide inhibits the glucagon response. The aim of the present study was to investigate whether this inhibition of glucagon release was mediated by a direct suppressive effect of...... specific antagonist of type 2 somatostatin receptor, DC-41-33 (2 micro mol/l), which fully antagonizes the suppressive somatostatin effect on rat A cells. Gliclazide (30 micro mol/l) inhibited glucagon release by 54% in the perfusion experiments, whereas the somatostatin response was nearly abolished. In...... islet perifusions with DC-41-33, arginine-induced glucagon release was inhibited by 66%. We therefore concluded that gliclazide inhibits glucagon release by a direct action on the pancreatic A cell....

  13. Arginine Methyltransferases Are Regulated by Epstein-Barr Virus in B Cells and Are Differentially Expressed in Hodgkin’s Lymphoma

    Directory of Open Access Journals (Sweden)

    Sarah Leonard

    2012-09-01

    Full Text Available Although there is increasing evidence that aberrant expression of those enzymes which control protein arginine methylation contribute to carcinogenesis, their de-regulation by oncogenic viruses in primary cells has yet to be reported. We first show that the protein arginine methyltransferases, CARM1, PRMT1 and PRMT5 are strongly expressed in Hodgkin Reed-Sternberg (HRS cells, and up-regulated in Hodgkin's lymphoma (HL cell lines. Given that Epstein-Barr virus (EBV can be detected in approximately 50% of primary HL, we next examined how EBV infection of germinal centre (GC B cells, the presumptive precursors of HRS cells, modulated the expression of these proteins. EBV infection of GC B cells was followed by the up-regulation of CARM1, PRMT1 and PRMT5, and by the down-regulation of the arginine deiminase, PADI4. Latent membrane protein 1 (LMP1, the major EBV transforming gene was shown to induce PRMT1 in GC B cells and in a stably transfected B cell line. The recent development of compounds which inhibit PRMT-mediated reactions provides a compelling case for continuing to dissect the contribution of virus induced changes in these proteins to lymphomagenesis.

  14. Arginine vasopressin in septic shock: supplement or substitute for norepinephrine?

    OpenAIRE

    Rehberg, Sebastian; Enkhbaatar, Perenlei; Traber, Daniel L

    2009-01-01

    In the current issue of Critical Care, Simon and coworkers investigated the effects of first-line arginine vasopressin (AVP) on organ function and systemic metabolism compared with norepinephrine in a pig model of fecal peritonitis. AVP was titrated according to the mean arterial pressure suggesting a vasopressor rather than a hormone replacement therapy. The study provides some evidence for the safety of this therapeutic approach. It needs to be determined whether AVP is most beneficial as a...

  15. Characterization of casein and Poly-L-arginine Multilayer Films

    OpenAIRE

    Warszynska, Lilianna Szyk; Kilan, Katarzyna; Socha, Robert P.

    2013-01-01

    Thin films containing casein appear to be a promising material for coatings used in the medical area to promote biomineralization. alfa- and beta-casein and poly-L-arginine multilayer films were formed by the layer-by layer technique and their thickness and mass were analyzed by ellipsometry and quartz crystal microbalance with dissipation monitoring (QCM-D). We investigated the effect of the type of casein used for the film formation and of the polyethyleneimine anchoring layer on the thickn...

  16. Plasmid DNA delivery by arginine-rich cell-penetrating peptides containing unnatural amino acids.

    Science.gov (United States)

    Kato, Takuma; Yamashita, Hiroko; Misawa, Takashi; Nishida, Koyo; Kurihara, Masaaki; Tanaka, Masakazu; Demizu, Yosuke; Oba, Makoto

    2016-06-15

    Cell-penetrating peptides (CPPs) have been developed as drug, protein, and gene delivery tools. In the present study, arginine (Arg)-rich CPPs containing unnatural amino acids were designed to deliver plasmid DNA (pDNA). The transfection ability of one of the Arg-rich CPPs examined here was more effective than that of the Arg nonapeptide, which is the most frequently used CPP. The transfection efficiencies of Arg-rich CPPs increased with longer post-incubation times and were significantly higher at 48-h and 72-h post-incubation than that of the commercially available transfection reagent TurboFect. These Arg-rich CPPs were complexed with pDNA for a long time in cells and effectively escaped from the late endosomes/lysosomes into the cytoplasm. These results will be helpful for designing novel CPPs for pDNA delivery. PMID:27132868

  17. Systems pathway engineering of Corynebacterium crenatum for improved L-arginine production

    Science.gov (United States)

    Man, Zaiwei; Xu, Meijuan; Rao, Zhiming; Guo, Jing; Yang, Taowei; Zhang, Xian; Xu, Zhenghong

    2016-01-01

    L-arginine is an important amino acid in food and pharmaceutical industries. Until now, the main production method of L-arginine in China is the highly polluting keratin acid hydrolysis. The industrial level L-arginine production by microbial fermentation has become an important task. In previous work, we obtained a new L-arginine producing Corynebacterium crenatum (subspecies of Corynebacterium glutamicum) through screening and mutation breeding. In this work, we performed systems pathway engineering of C. crenatum for improved L-arginine production, involving amplification of L-arginine biosynthetic pathway flux by removal of feedback inhibition and overexpression of arginine operon; optimization of NADPH supply by modulation of metabolic flux distribution between glycolysis and pentose phosphate pathway; increasing glucose consumption by strengthening the preexisting glucose transporter and exploitation of new glucose uptake system; channeling excess carbon flux from glycolysis into tricarboxylic acid cycle to alleviate the glucose overflow metabolism; redistribution of carbon flux at α-ketoglutarate metabolic node to channel more flux into L-arginine biosynthetic pathway; minimization of carbon and cofactor loss by attenuation of byproducts formation. The final strain could produce 87.3 g L−1 L-arginine with yield up to 0.431 g L-arginine g−1 glucose in fed-batch fermentation. PMID:27338253

  18. Systems pathway engineering of Corynebacterium crenatum for improved L-arginine production.

    Science.gov (United States)

    Man, Zaiwei; Xu, Meijuan; Rao, Zhiming; Guo, Jing; Yang, Taowei; Zhang, Xian; Xu, Zhenghong

    2016-01-01

    L-arginine is an important amino acid in food and pharmaceutical industries. Until now, the main production method of L-arginine in China is the highly polluting keratin acid hydrolysis. The industrial level L-arginine production by microbial fermentation has become an important task. In previous work, we obtained a new L-arginine producing Corynebacterium crenatum (subspecies of Corynebacterium glutamicum) through screening and mutation breeding. In this work, we performed systems pathway engineering of C. crenatum for improved L-arginine production, involving amplification of L-arginine biosynthetic pathway flux by removal of feedback inhibition and overexpression of arginine operon; optimization of NADPH supply by modulation of metabolic flux distribution between glycolysis and pentose phosphate pathway; increasing glucose consumption by strengthening the preexisting glucose transporter and exploitation of new glucose uptake system; channeling excess carbon flux from glycolysis into tricarboxylic acid cycle to alleviate the glucose overflow metabolism; redistribution of carbon flux at α-ketoglutarate metabolic node to channel more flux into L-arginine biosynthetic pathway; minimization of carbon and cofactor loss by attenuation of byproducts formation. The final strain could produce 87.3 g L(-1) L-arginine with yield up to 0.431 g L-arginine g(-1) glucose in fed-batch fermentation. PMID:27338253

  19. Principles of protein targeting to the nucleolus.

    Science.gov (United States)

    Martin, Robert M; Ter-Avetisyan, Gohar; Herce, Henry D; Ludwig, Anne K; Lättig-Tünnemann, Gisela; Cardoso, M Cristina

    2015-01-01

    The nucleolus is the hallmark of nuclear compartmentalization and has been shown to exert multiple roles in cellular metabolism besides its main function as the place of rRNA synthesis and assembly of ribosomes. Nucleolar proteins dynamically localize and accumulate in this nuclear compartment relative to the surrounding nucleoplasm. In this study, we have assessed the molecular requirements that are necessary and sufficient for the localization and accumulation of peptides and proteins inside the nucleoli of living cells. The data showed that positively charged peptide entities composed of arginines alone and with an isoelectric point at and above 12.6 are necessary and sufficient for mediating significant nucleolar accumulation. A threshold of 6 arginines is necessary for peptides to accumulate in nucleoli, but already 4 arginines are sufficient when fused within 15 amino acid residues of a nuclear localization signal of a protein. Using a pH sensitive dye, we found that the nucleolar compartment is particularly acidic when compared to the surrounding nucleoplasm and, hence, provides the ideal electrochemical environment to bind poly-arginine containing proteins. In fact, we found that oligo-arginine peptides and GFP fusions bind RNA in vitro. Consistent with RNA being the main binding partner for arginines in the nucleolus, we found that the same principles apply to cells from insects to man, indicating that this mechanism is highly conserved throughout evolution. PMID:26280391

  20. The arginine methyltransferase Rmt2 is enriched in the nucleus and co-purifies with the nuclear porins Nup49, Nup57 and Nup100

    International Nuclear Information System (INIS)

    Arginine methylation is a post-translational modification of proteins implicated in RNA processing, protein compartmentalization, signal transduction, transcriptional regulation and DNA repair. In a screen for proteins associated with the nuclear envelope in the yeast Saccharomyces cerevisiae, we have identified the arginine methyltransferase Rmt2, previously shown to methylate the ribosomal protein L12. By indirect immunofluorescence and subcellular fractionations we demonstrate here that Rmt2 has nuclear and cytoplasmic localizations. Biochemical analysis of a fraction enriched in nuclei reveals that nuclear Rmt2 is resistant to extractions with salt and detergent, indicating an association with structural components. This was supported by affinity purification experiments with TAP-tagged Rmt2. Rmt2 was found to co-purify with the nucleoporins Nup49, Nup57 and Nup100, revealing a novel link between arginine methyltransferases and the nuclear pore complex. In addition, a genome-wide transcription study of the rmt2Δ mutant shows significant downregulation of the transcription of MYO1, encoding the Type II myosin heavy chain required for cytokinesis and cell separation

  1. Purification and characterization of an arginine ester hydrolase from the venom of Trimeresurus mucrosqumatus in Hunan province of China

    Institute of Scientific and Technical Information of China (English)

    YU Xiao-dong; LI Bo; YU Zheng-ping

    2005-01-01

    Objective: To study the physical and chemical properties of an arginine ester hydrolase from the venom of Trimeresurus mucrosqumatus in Hunan province of China. Methods :The arginine ester hydrolase (AEH) was isolated from the venom of Chinese Trimeresurus mucrosqumatus by a combination of ionexchange chromatography on DEAE-Sephadex A-50, CM-Sepharose Cl-6B and gel filtration on Sephadex G-100. Results: The purified protein named TM-AEH,a glycoprotein with carbohydrate content of 0.5 % neutral hexose and 0. 75 % sialic acid,a relative molecular mass of 29.0 kDa,and an isoelectric point (pI) of 5. 2. It shares with an extinction coefficient (E0.1%/cm) of 1.332 at 280 nm,consisted of 225 amino acid residues ,and migrated as a band under reduced or non-reduced condition in basic PAGE. TM-AEH was a highly thermostable protein and was stable to pH changes between 5 and 9. The optimum temperature and optimum pH were 55℃ and 8. 4 for its catalytic activity respectively,which was inhibited by Fe3+ and Cu2+. Conclusion:This protein can exhibit higher BAEE-hydrolysing activity and fibrinogenolytic activity as compared to that of whole venom.

  2. Expression pattern of a nuclear encoded mitochondrial arginine-ornithine translocator gene from Arabidopsis

    OpenAIRE

    Schneider Anja; Kunze Reinhard; Wipf Daniel; Hilpert Melanie; Desimone Marcelo; Catoni Elisabetta; Flügge Ulf-Ingo; Schumacher Karin; Frommer Wolf B

    2003-01-01

    Abstract Background Arginine and citrulline serve as nitrogen storage forms, but are also involved in biosynthetic and catabolic pathways. Metabolism of arginine, citrulline and ornithine is distributed between mitochondria and cytosol. For the shuttle of intermediates between cytosol and mitochondria transporters present on the inner mitochondrial membrane are required. Yeast contains a mitochondrial translocator for ornithine and arginine, Ort1p/Arg11p. Ort1p/Arg11p is a member of the mitoc...

  3. The arginine-ornithine antiporter ArcD contributes to biological fitness of Streptococcus suis

    Directory of Open Access Journals (Sweden)

    Marcus eFulde

    2014-08-01

    Full Text Available The arginine-ornithine antiporter (ArcD is part of the Arginine Deiminase System (ADS, a catabolic, energy-providing pathway found in a variety of different bacterial species, including the porcine zoonotic pathogen Streptococcus suis. The ADS has recently been shown to play a role in the pathogenicity of S. suis, in particular in its survival in host cells. The contribution of arginine and arginine transport mediated by ArcD, however, has yet to be clarified. In the present study, we showed by experiments using [U-13C6]arginine as a tracer molecule that S. suis is auxotrophic for arginine and that bacterial growth depends on the uptake of extracellular arginine. To further study the role of ArcD in arginine metabolism, we generated an arcD-specific mutant strain and characterized its growth compared to the wild-type (WT strain, a virulent serotype 2 strain. The mutant strain showed a markedly reduced growth rate in chemically defined media supplemented with arginine when compared to the WT strain, indicating that ArcD promotes arginine uptake. To further evaluate the in vivo relevance of ArcD, we studied the intracellular bacterial survival of the arcD mutant strain in an epithelial cell culture infection model. The mutant strain was substantially attenuated, and its reduced intracellular survival rate correlated with a lower ability to neutralize the acidified environment. Based on these results, we propose that ArcD, by its function as an arginine-ornithine antiporter, is important for supplying arginine as substrate of the ADS and, thereby, contributes to biological fitness and virulence of S. suis in the host.

  4. Risk and risk reduction involving arginine intake and meat consumption in colorectal tumorigenesis and survival

    OpenAIRE

    Zell, Jason A.; Ignatenko, Natalia A.; Yerushalmi, Hagit F; Ziogas, Argyrios; Besselsen, David G; Gerner, Eugene W.; Anton-Culver, Hoda

    2007-01-01

    Elevated polyamine and nitric oxide levels (both derived from arginine) promote tumorigenesis, whereas non-steroidal anti-inflammatory drugs (NSAIDs) inhibit colorectal cancer (CRC) incidence in experimental and epidemiologic studies. We investigated dietary arginine-induced intestinal tumorigenesis and NSAID-inhibitory effects in Apc(Min/+) mice differentially expressing nitric oxide synthase-2 (Nos2). We also studied effects of estimated arginine exposures through meat consumption on tumor ...

  5. Remission of diabetes mellitus in cats cannot be predicted by the arginine stimulation test

    OpenAIRE

    Tschuor, F

    2011-01-01

    Background: Responsiveness of β-cells to arginine persists the longest during diabetes progression, making the intravenous arginine stimulation test (IVAST) a useful tool to assess residual insulin and glucagon secretion. Hypothesis: Diabetic cats with and without remission will have different arginine-induced insulin or glucagon response. Animals: 17 cats with diabetes, 7 healthy cats. Methods: Response to IVAST was assessed by calculating insulin and glucagon area under the c...

  6. Hyponatraemia in the first week of life in preterm infants. Part I. Arginine vasopressin secretion.

    OpenAIRE

    Rees, L; Brook, C G; Shaw, J C; Forsling, M L

    1984-01-01

    Continuous sequential urinary arginine vasopressin measurements in 14 preterm, ventilated infants suggest that both osmoreceptor and volume receptor systems are able to stimulate the prolonged secretion of arginine vasopressin from 26 weeks' gestation. The kidney is able to respond to arginine vasopressin stimulation from the first day of life and from 26 weeks' gestation. A maximum urine osmolality not exceeding 550 mOsm/kg was reached which varied with hydration of the infant. Excretion of ...

  7. Protective Effect of Arginine on Oxidative Stress in Transgenic Sickle Mouse Models

    OpenAIRE

    Dasgupta, Trisha; Hebbel, Robert P.; Kaul, Dhananjay K.

    2006-01-01

    Sickle cell disease (SCD) is characterized by reperfusion injury and chronic oxidative stress. Oxidative stress and hemolysis in SCD result in inactivation of nitric oxide (NO) and depleted arginine levels. We hypothesized that augmenting NO production by arginine supplementation will reduce oxidative stress in SCD. To this end, we measured the effect of arginine (5% in mouse chow) on NO metabolites (NOx), lipid peroxidation (LPO) and selected antioxidants in transgenic sickle mouse models. U...

  8. Arginine Consumption by the Intestinal Parasite Giardia intestinalis Reduces Proliferation of Intestinal Epithelial Cells

    OpenAIRE

    Stadelmann, Britta; Merino, Maria C.; Persson, Lo; Svard, Staffan G.

    2012-01-01

    In the field of infectious diseases the multifaceted amino acid arginine has reached special attention as substrate for the host´s production of the antimicrobial agent nitric oxide (NO). A variety of infectious organisms interfere with this part of the host immune response by reducing the availability of arginine. This prompted us to further investigate additional roles of arginine during pathogen infections. As a model we used the intestinal parasite Giardia intestinalis that actively consu...

  9. Oral Arginine Metabolism May Decrease the Risk for Dental Caries in Children

    OpenAIRE

    Nascimento, M. M.; Liu, Y.; Kalra, R.; Perry, S.; Adewumi, A.; Xu, X.; Primosch, R.E.; Burne, R A

    2013-01-01

    Arginine metabolism by oral bacteria via the arginine deiminase system (ADS) increases the local pH, which can neutralize the effects of acidification from sugar metabolism and reduce the cariogenicity of oral biofilms. To explore the relationship between oral arginine metabolism and dental caries experience in children, we measured ADS activity in oral samples from 100 children and correlated it with their caries status and type of dentition. Supragingival dental plaque was collected from to...

  10. Bioinformatic evaluation of L-arginine catabolic pathways in 24 cyanobacteria and transcriptional analysis of genes encoding enzymes of L-arginine catabolism in the cyanobacterium Synechocystis sp. PCC 6803

    Directory of Open Access Journals (Sweden)

    Pistorius Elfriede K

    2007-11-01

    Full Text Available Abstract Background So far very limited knowledge exists on L-arginine catabolism in cyanobacteria, although six major L-arginine-degrading pathways have been described for prokaryotes. Thus, we have performed a bioinformatic analysis of possible L-arginine-degrading pathways in cyanobacteria. Further, we chose Synechocystis sp. PCC 6803 for a more detailed bioinformatic analysis and for validation of the bioinformatic predictions on L-arginine catabolism with a transcript analysis. Results We have evaluated 24 cyanobacterial genomes of freshwater or marine strains for the presence of putative L-arginine-degrading enzymes. We identified an L-arginine decarboxylase pathway in all 24 strains. In addition, cyanobacteria have one or two further pathways representing either an arginase pathway or L-arginine deiminase pathway or an L-arginine oxidase/dehydrogenase pathway. An L-arginine amidinotransferase pathway as a major L-arginine-degrading pathway is not likely but can not be entirely excluded. A rather unusual finding was that the cyanobacterial L-arginine deiminases are substantially larger than the enzymes in non-photosynthetic bacteria and that they are membrane-bound. A more detailed bioinformatic analysis of Synechocystis sp. PCC 6803 revealed that three different L-arginine-degrading pathways may in principle be functional in this cyanobacterium. These are (i an L-arginine decarboxylase pathway, (ii an L-arginine deiminase pathway, and (iii an L-arginine oxidase/dehydrogenase pathway. A transcript analysis of cells grown either with nitrate or L-arginine as sole N-source and with an illumination of 50 μmol photons m-2 s-1 showed that the transcripts for the first enzyme(s of all three pathways were present, but that the transcript levels for the L-arginine deiminase and the L-arginine oxidase/dehydrogenase were substantially higher than that of the three isoenzymes of L-arginine decarboxylase. Conclusion The evaluation of 24

  11. Dietary Arginine Requirements for Growth Are Dependent on the Rate of Citrulline Production in Mice123

    Science.gov (United States)

    Marini, Juan C; Agarwal, Umang; Didelija, Inka C

    2015-01-01

    Background: In many species, including humans, arginine is considered a semiessential amino acid because under certain conditions endogenous synthesis cannot meet its demand. The requirements of arginine for growth in mice are ill defined and seem to vary depending on the genetic background of the mice. Objective: The objective of this study was to determine the metabolic and molecular basis for the requirement of arginine in 2 mouse strains. Methods: Institute of Cancer Research (ICR) and C57BL/6 (BL6) male mice were fed arginine-free or arginine-sufficient diets (Expt. 1) or 1 of 7 diets with increasing arginine concentration (from 0- to 8-g/kg diet, Expt. 2) between day 24 and 42 of life to determine the arginine requirements for growth. Citrulline production and “de novo” arginine synthesis were measured with use of stable isotopes, and arginine requirements were determined by breakpoint analysis and enzyme expression by reverse transcriptase-polymerase chain reaction. Results: In Expt. 1, ICR mice grew at the same rate regardless of the arginine concentration of the diet (mean ± SE: 0.66 ± 0.04 g/d, P = 0.80), but BL6 mice had a reduced growth rate when fed the arginine-free diet (0.25 ± 0.02 g/d, P < 0.001) compared to the 8-g arginine/kg diet (0.46 ± 0.03 g/d). ICR mice showed at least a 2-fold greater expression (P < 0.001) of ornithine transcarbamylase (OTC) than BL6 mice, which translated into a greater rate of citrulline (25%) and arginine synthesis (49%, P < 0.002). In Expt. 2, breakpoint analysis showed that the requirement for growth of BL6 mice was met with 2.32 ± 0.39 g arginine/kg diet; for ICR mice, however, no breakpoint was found. Conclusion: Our data indicate that a reduced expression of OTC in BL6 mice translates into a reduced production of citrulline and arginine compared with ICR mice, which results in a dietary arginine requirement for growth in BL6 mice, but not in ICR mice. PMID:25855119

  12. Temporal Expression of the Bacillus subtilis secA Gene, Encoding a Central Component of the Preprotein Translocase

    OpenAIRE

    Herbort, Markus; Klein, Michael; Manting, Erik H.; Driessen, Arnold J. M.; Freudl, Roland

    1999-01-01

    In Bacillus subtilis, the secretion of extracellular proteins strongly increases upon transition from exponential growth to the stationary growth phase. It is not known whether the amounts of some or all components of the protein translocation apparatus are concomitantly increased in relation to the increased export activity. In this study, we analyzed the transcriptional organization and temporal expression of the secA gene, encoding a central component of the B. subtilis preprotein transloc...

  13. Expression, purification, crystallization and preliminary crystallographic study of isolated modules of the mouse coactivator-associated arginine methyltransferase 1

    International Nuclear Information System (INIS)

    Isolated modules of mouse coactivator-associated arginine methyltransferase 1 encompassing the protein arginine N-methyltransferase catalytic domain have been overexpressed, purified and crystallized. X-ray diffraction data have been collected and have enabled determination of the structures by multiple isomorphous replacement using anomalous scattering. Coactivator-associated arginine methyltransferase 1 (CARM1) plays a crucial role in gene expression as a coactivator of several nuclear hormone receptors and also of non-nuclear receptor systems. Its recruitment by the transcriptional machinery induces protein methylation, leading to chromatin remodelling and gene activation. CARM128–507 and two structural states of CARM1140–480 were expressed, purified and crystallized. Crystals of CARM128–507 belong to space group P6222, with unit-cell parameters a = b = 136.0, c = 125.3 Å; they diffract to beyond 2.5 Å resolution using synchrotron radiation and contain one monomer in the asymmetric unit. The structure of CARM128–507 was solved by multiple isomorphous replacement and anomalous scattering methods. Crystals of apo CARM1140–480 belong to space group I222, with unit-cell parameters a = 74.6, b = 99.0, c = 207.4 Å; they diffract to beyond 2.7 Å resolution and contain two monomers in the asymmetric unit. Crystals of CARM1140–480 in complex with S-adenosyl-l-homocysteine belong to space P21212, with unit-cell parameters a = 74.6, b = 98.65, c = 206.08 Å; they diffract to beyond 2.6 Å resolution and contain four monomers in the asymmetric unit. The structures of apo and holo CARM1140–480 were solved by molecular-replacement techniques from the structure of CARM128–507

  14. To involvement the conformation of the adenine nucleotide translocase in opening the Tl(+)-induced permeability transition pore in Ca(2+)-loaded rat liver mitochondria.

    Science.gov (United States)

    Korotkov, Sergey M; Konovalova, Svetlana A; Brailovskaya, Irina V; Saris, Nils-Erik L

    2016-04-01

    The conformation of adenine nucleotide translocase (ANT) has a profound impact in opening the mitochondrial permeability transition pore (MPTP) in the inner membrane. Fixing the ANT in 'c' conformation by phenylarsine oxide (PAO), tert-butylhydroperoxide (tBHP), and carboxyatractyloside as well as the interaction of 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS) with mitochondrial thiols markedly attenuated the ability of ADP to inhibit the MPTP opening. We earlier found (Korotkov and Saris, 2011) that calcium load of rat liver mitochondria in medium containing TlNO3 and KNO3 stimulated the Tl(+)-induced MPTP opening in the inner mitochondrial membrane. The MPTP opening as well as followed increase in swelling, a drop in membrane potential (ΔΨmito), and a decrease in state 3, state 4, and 2,4-dinitrophenol-uncoupled respiration were visibly enhanced in the presence of PAO, tBHP, DIDS, and carboxyatractyloside. However, these effects were markedly inhibited by ADP and membrane-penetrant hydrophobic thiol reagent, N-ethylmaleimide (NEM) which fix the ANT in 'm' conformation. Cyclosporine A additionally potentiated these effects of ADP and NEM. Our data suggest that conformational changes of the ANT may be directly involved in the opening of the Tl(+)-induced MPTP in the inner membrane of Ca(2+)-loaded rat liver mitochondria. Using the Tl(+)-induced MPTP model is discussed in terms finding new transition pore inhibitors and inducers among different chemical and natural compounds. PMID:26835787

  15. Identification of novel inhibitors of phospho-MurNAc-pentapeptide translocase MraY from library screening: Isoquinoline alkaloid michellamine B and xanthene dye phloxine B.

    Science.gov (United States)

    Mihalyi, Agnes; Jamshidi, Shirin; Slikas, Justinas; Bugg, Timothy D H

    2014-09-01

    The National Cancer Institute (NCI) Diversity Set was screened for potential inhibitors of phospho-MurNAc-pentapeptide translocase MraY from Escherichia coli using a primary fluorescence enhancement assay, followed by a secondary radiochemical assay. One new MraY inhibitor was identified from this screen, a naphthylisoquinoline alkaloid michellamine B, which inhibited E. coli MraY (IC50 456μM) and Bacillus subtilis MraY (IC50 386μM), and which showed antimicrobial activity against B. subtilis (MIC 16μg/mL). Following an earlier report of halogenated fluoresceins identified from a combined MraY/MurG screen, three halogenated fluoresceins were tested as inhibitors of E. coli MraY and E. coli MurG, and phloxine B was identified as an inhibitor of E. coli MraY (IC50 32μM). Molecular docking of inhibitor structures against the structure of Aquifex aeolicus MraY indicates that phloxine B appears to bind to the Mg(2+) cofactor in the enzyme active site, while michellamine B binds to a hydrophobic groove formed between transmembrane helices 5 and 9. PMID:25127465

  16. Type 1 diabetes: can exercise impair the autoimmune event? The L-arginine/glutamine coupling hypothesis.

    Science.gov (United States)

    Krause, Maurício da Silva; de Bittencourt, Paulo Ivo Homem

    2008-06-01

    Prevention of type 1 diabetes mellitus (T1DM) requires early intervention in the autoimmune process directed against beta-cells of the pancreatic islets of Langerhans, which is believed to result from a disorder of immunoregulation. According to this concept, a T-helper lymphocyte of type 1 (Th1) subset of T-lymphocytes and their cytokine products, the type 1 cytokines [e.g. interleukin 2 (IL-2), interferon gamma (IFN-gamma) and tumour necrosis factor beta (TNF-beta)] prevail over immunoregulatory (anti-inflammatory) Th2 subset and its cytokine products, i.e. type 2 cytokines (e.g. IL-4, IL-6 and IL-10). This allows type 1 cytokines to initiate a cascade of immune/inflammatory processes in the islet (insulitis), culminating in beta-cell destruction. Activation of sympathetic-corticotropin-releasing hormone (CRH) axis by psychological stress induces specifically Th1 cell overactivity that determines enhanced glutamine utilization and consequent poor L-arginine supply for nitric oxide (NO)-assisted insulin secretion. This determines the shift of intraislet glutamate metabolism from the synthesis of glutathione (GSH) to that of L-arginine, leading to a redox imbalance that activates nuclear factor kappaB exacerbating inflammation and NO-mediated cytotoxicity. Physical exercise is capable of inducing changes in the pattern of cytokine production and release towards type 2 class and to normalize the glutamine supply to the circulation, which reduces the need for glutamate, whose metabolic fate may be restored in the direction of GSH synthesis and antioxidant defence. Also, the 70-kDa heat shock protein (hsp70), which is immunoregulatory, may modulate exercise-induced anti-inflammation. In this work, we envisage how exercise can intervene in the mechanisms involved in the autoimmune process against beta-cells and how novel therapeutic approaches may be inferred from these observations. PMID:18383559

  17. Thermoanalytical study of N{sub {alpha}}-benzoyl-L-argininate ethyl ester chloride

    Energy Technology Data Exchange (ETDEWEB)

    Fonseca, A.C. [Department of Chemical Engineering, University of Coimbra, Polo II, Pinhal de Marrocos, Coimbra 3030-790 (Portugal); Jarmelo, S. [Department of Chemical Engineering, University of Coimbra, Polo II, Pinhal de Marrocos, Coimbra 3030-790 (Portugal); Department of Chemistry, University of Coimbra, Coimbra 3004-535 (Portugal); Canotilho, J. [Faculty of Pharmacy, University of Coimbra, Polo das Ciencias da Saude, Azinhaga de Santa Comba, Coimbra 3000-548 (Portugal); Eusebio, M.E.S.; Fausto, R. [Department of Chemistry, University of Coimbra, Coimbra 3004-535 (Portugal); Gil, M.H.; Simoes, P.N. [Department of Chemical Engineering, University of Coimbra, Polo II, Pinhal de Marrocos, Coimbra 3030-790 (Portugal)

    2012-01-10

    Highlights: Black-Right-Pointing-Pointer Thermal analysis of crystalline N{sub {alpha}}-benzoyl-L-argininate ethyl ester chloride. Black-Right-Pointing-Pointer Characterization by STA, MDSC, PLTM, and HiRes-MTGA. Black-Right-Pointing-Pointer Non-isothermal kinetic analysis of the thermal decomposition. - Abstract: N{sub {alpha}}-benzoyl-L-argininate ethyl ester chloride (BAEEH{sup +}{center_dot}Cl{sup -}) has been used as a model drug in the development of polymeric delivery systems for peptides and proteins. In this study, crystalline BAEEH{sup +}{center_dot}Cl{sup -} was characterized by different thermoanalytical techniques. The main thermal events were identified by modulated differential scanning calorimetry (MDSC). Polarized light thermomicroscopy (PLTM) showed that no phase transitions but fusion occur before thermal decomposition. A crystalline phase could not be obtained from the melt in cooling/heating cycles between 150 Degree-Sign C and -150 Degree-Sign C, at 10 Degree-Sign C min{sup -1} scanning rate. The thermal stability and the non-isothermal kinetic analysis of the thermal decomposition were studied by conventional and high resolution modulated thermogravimetric analysis (HiRes-MTGA). Depending on the conditions of the study, the decomposition starts in the range from 197 Degree-Sign C (HiRes-MTGA at {phi} = 2 Degree-Sign C min{sup -1}) to 261 Degree-Sign C (TGA at 20 Degree-Sign C min{sup -1}). It was found that the first stage of the decomposition process can be described by the kinetic triplet E{sub a} = 142 kJ mol{sup -1}, A = 1.86 Multiplication-Sign 10{sup 11} s{sup -1}, and f({alpha}) = (1 - {alpha}){sup 2.78}.

  18. Thermoanalytical study of Nα-benzoyl-L-argininate ethyl ester chloride

    International Nuclear Information System (INIS)

    Highlights: ► Thermal analysis of crystalline Nα-benzoyl-L-argininate ethyl ester chloride. ► Characterization by STA, MDSC, PLTM, and HiRes-MTGA. ► Non-isothermal kinetic analysis of the thermal decomposition. - Abstract: Nα-benzoyl-L-argininate ethyl ester chloride (BAEEH+·Cl−) has been used as a model drug in the development of polymeric delivery systems for peptides and proteins. In this study, crystalline BAEEH+·Cl− was characterized by different thermoanalytical techniques. The main thermal events were identified by modulated differential scanning calorimetry (MDSC). Polarized light thermomicroscopy (PLTM) showed that no phase transitions but fusion occur before thermal decomposition. A crystalline phase could not be obtained from the melt in cooling/heating cycles between 150 °C and −150 °C, at 10 °C min−1 scanning rate. The thermal stability and the non-isothermal kinetic analysis of the thermal decomposition were studied by conventional and high resolution modulated thermogravimetric analysis (HiRes-MTGA). Depending on the conditions of the study, the decomposition starts in the range from 197 °C (HiRes-MTGA at φ = 2 °C min−1) to 261 °C (TGA at 20 °C min−1). It was found that the first stage of the decomposition process can be described by the kinetic triplet Ea = 142 kJ mol−1, A = 1.86 × 1011 s−1, and f(α) = (1 − α)2.78.

  19. Secretory production of an FAD cofactor-containing cytosolic enzyme (sorbitol–xylitol oxidase from Streptomyces coelicolor) using the twin-arginine translocation (Tat) pathway of Corynebacterium glutamicum

    Science.gov (United States)

    Scheele, Sandra; Oertel, Dan; Bongaerts, Johannes; Evers, Stefan; Hellmuth, Hendrik; Maurer, Karl-Heinz; Bott, Michael; Freudl, Roland

    2013-01-01

    Carbohydrate oxidases are biotechnologically interesting enzymes that require a tightly or covalently bound cofactor for activity. Using the industrial workhorse Corynebacterium glutamicum as the expression host, successful secretion of a normally cytosolic FAD cofactor-containing sorbitol–xylitol oxidase from Streptomyces coelicolor was achieved by using the twin-arginine translocation (Tat) protein export machinery for protein translocation across the cytoplasmic membrane. Our results demonstrate for the first time that, also for cofactor-containing proteins, a secretory production strategy is a feasible and promising alternative to conventional intracellular expression strategies. PMID:23163932

  20. Arginine vasopressin stimulates proliferation of adult rat cardiac fibroblasts via protein kinase C-extracellular signal-regulated kinase 1/2 pathway%蛋白激酶C-细胞外信号调节激酶1/2通路介导精氨酸升压素对成年大鼠心肌成纤维细胞的促增殖作用

    Institute of Scientific and Technical Information of China (English)

    何燕萍; 赵连友; 郑强荪; 刘少伟; 赵晓燕; 陆晓龙; 牛晓琳

    2008-01-01

    精氨酸升压素(arginine vasopressin, AVP)是高血压和心力衰竭时激活的神经体液和血流动力学因子,同时,它还具有直接的生长刺激作用.我们以往的研究显示AVP可诱导新生大鼠心肌成纤维细胞(cardiac fibroblasts, CFs)增殖.本研究旨在进一步观察AVP是否对成年大鼠CFs具有促增殖作用,并探计其机制.采用组织块法培养成年大鼠CFs,用[3H]-TdR掺入法和流式细胞仪方法观察AVP作用下CFs的DNA合成和细胞周期分布.根据特异性底物髓磷脂基质蛋白(myelin basic protein, MBP)的磷酸化水平测定细胞外信号调节激酶1/2 (extracellular signal-regulated kinase 1/2, ERK1/2)的活性.用Western blot检测ERK1/2的磷酸化和p27Kip1、细胞周期蛋白D1、 A、 E的表达.结果显示,AVP(0.1μmol/L)可促进成年大鼠CFs的DNA合成,该作用可被V1受体拮抗剂d(CH2)5[Tyr2(Me),Arg8]-vasopressin (0.1μmol/L)阻断,而不受V2受体拮抗剂desglycinamide [d(CH2)5, D-Ile2, Ile4, Arg8]-vasopressin (0.1μmol/L)的影响.AVP可激活ERK1/2,用蛋白激酶C(protein kinase C, PKC)激动剂佛波酯(phorbol 12-myristate 13-acetate, PMA, 30nmol/L, 5min)急性刺激可模拟该作用,而PMA持续慢性作用(2.5μmol/L,24h)耗竭PKC后则抑制AVP对ERK1/2的激活.AVP可抑制p27Kip1的蛋白表达,升高细胞周期蛋白D1、 A和E的表达,同时促进细胞周期由G0/G1期进入S期.ERK1/2抑制剂PD98059 (30μmol/L)阻断AVP对DNA合成、p27Kip1、细胞周期蛋白D1、A和E蛋白表达的作用,并抑制细胞周期进程.以上结果表明,AVP可促进成年大鼠CFs增殖,该作用由V1受体和PKC-ERK1/2通路介导.AVP可通过ERK1/2调控p27Kip1、细胞周期蛋白D1、A和E的表达,从而促进成年大鼠CFs的细胞周期进程.%Arginine vasopressin (AVP), a neurohormone and hemodynamic factor implicated in the pathophysiology of hypertension and congestive heart failure, can also act as a growth-stimulating factor. Our previous work

  1. The crystal structure of the complex of PII and acetylglutamate kinase reveals how PII controls the storage of nitrogen as arginine.

    Science.gov (United States)

    Llácer, José L; Contreras, Asunción; Forchhammer, Karl; Marco-Marín, Clara; Gil-Ortiz, Fernando; Maldonado, Rafael; Fita, Ignacio; Rubio, Vicente

    2007-11-01

    Photosynthetic organisms can store nitrogen by synthesizing arginine, and, therefore, feedback inhibition of arginine synthesis must be relieved in these organisms when nitrogen is abundant. This relief is accomplished by the binding of the PII signal transduction protein to acetylglutamate kinase (NAGK), the controlling enzyme of arginine synthesis. Here, we describe the crystal structure of the complex between NAGK and PII of Synechococcus elongatus, at 2.75-A resolution. We prove the physiological relevance of the observed interactions by site-directed mutagenesis and functional studies. The complex consists of two polar PII trimers sandwiching one ring-like hexameric NAGK (a trimer of dimers) with the threefold axes of these molecules aligned. The binding of PII favors a narrow ring conformation of the NAGK hexamer that is associated with arginine sites having low affinity for this inhibitor. Each PII subunit contacts one NAGK subunit only. The contacts map in the inner circumference of the NAGK ring and involve two surfaces of the PII subunit. One surface is on the PII body and interacts with the C-domain of the NAGK subunit, helping widen the arginine site found on the other side of this domain. The other surface is at the distal region of a protruding large loop (T-loop) that presents a novel compact shape. This loop is inserted in the interdomain crevice of the NAGK subunit, contacting mainly the N-domain, and playing key roles in anchoring PII on NAGK, in activating NAGK, and in complex formation regulation by MgATP, ADP, 2-oxoglutarate, and by phosphorylation of serine-49. PMID:17959776

  2. L-Canavanine potentiates the cytotoxicity of doxorubicin and cisplatin in arginine deprived human cancer cells

    Science.gov (United States)

    Wink, Michael

    2016-01-01

    The non-protein amino acid L-canavanine (L-CAV), an antimetabolite of L-arginine (L-ARG), can alter the 3D conformation of proteins when incorporated into a protein instead of L-ARG. L-CAV inhibits the proliferation of some tumour cells. The deprivation of L-ARG in the culture medium enhances the response of cells to L-CAV. This study aimed to investigate the interaction of L-CAV in combination with the chemotherapeutic drugs, doxorubicin (DOX) or cisplatin (CIS), in cancer cells, especially in the absence of L-ARG. A combination method based on the median-effect principle and mass-action law was used. The following cancer cells were employed: HeLa and Caco-2 cells, overexpressing argininosuccinate synthase (ASS), pancreatic cells (MIA PaCa-2 and BxPC-3) and hepatocellular carcinoma cells (Hep G2 and SK-HEP-1), with down-regulated ASS. When constant and non-constant ratios of L-CAV were combined with DOX and CIS, a synergistic potentiation of cytotoxicity was recorded. Cells expressing high levels of ASS were more sensitive to the treatment as compared to the cells with reduced ASS levels. Overall, this study may provide a new approach to targeting some cancer cells with L-CAV in combination with DNA-targeting drugs such as DOX and CIS, especially those cells which overexpress ASS, such as human cervical and colorectal carcinoma cells. PMID:26839743

  3. Dietary L-arginine supplementation reduces Methotrexate-induced intestinal mucosal injury in rat

    Directory of Open Access Journals (Sweden)

    Koppelmann Tal

    2012-04-01

    Full Text Available Abstract Background Arginine (ARG and nitric oxide maintain the mucosal integrity of the intestine in various intestinal disorders. In the present study, we evaluated the effects of oral ARG supplementation on intestinal structural changes, enterocyte proliferation and apoptosis following methotrexate (MTX-induced intestinal damage in a rat. Methods Male rats were divided into four experimental groups: Control rats, CONTR-ARG rats, were treated with oral ARG given in drinking water 72 hours before and 72 hours following vehicle injection, MTX rats were treated with a single dose of methotrexate, and MTX-ARG rats were treated with oral ARG following injection of MTX. Intestinal mucosal damage, mucosal structural changes, enterocyte proliferation and enterocyte apoptosis were determined 72 hours following MTX injection. RT-PCR was used to determine bax and bcl-2 mRNA expression. Results MTX-ARG rats demonstrated greater jejunal and ileal bowel weight, greater ileal mucosal weight, greater ileal mucosal DNA and protein levels, greater villus height in jejunum and ileum and crypt depth in ileum, compared to MTX animals. A significant decrease in enterocyte apoptosis in the ileum of MTX-ARG rats (vs MTX was accompanied by decreased bax mRNA and protein expression and increased bcl-2 protein levels. Conclusions Treatment with oral ARG prevents mucosal injury and improves intestinal recovery following MTX- injury in the rat.

  4. Stability and resilience of oral microcosms toward acidification and Candida outgrowth by arginine supplementation.

    Science.gov (United States)

    Koopman, Jessica E; Röling, Wilfred F M; Buijs, Mark J; Sissons, Christopher H; ten Cate, Jacob M; Keijser, Bart J F; Crielaard, Wim; Zaura, Egija

    2015-02-01

    Dysbiosis induced by low pH in the oral ecosystem can lead to caries, a prevalent bacterial disease in humans. The amino acid arginine is one of the pH-elevating agents in the oral cavity. To obtain insights into the effect of arginine on oral microbial ecology, a multi-plaque "artificial mouth" (MAM) biofilm model was inoculated with saliva from a healthy volunteer and microcosms were grown for 4 weeks with 1.6 % (w/v) arginine supplement (Arginine) or without (Control), samples were taken at several time-points. A cariogenic environment was mimicked by sucrose pulsing. The bacterial composition was determined by 16S rRNA gene amplicon sequencing, the presence and amount of Candida and arginine deiminase system genes arcA and sagP by qPCR. Additionally, ammonium and short-chain fatty acid concentrations were determined. The Arginine microcosms were dominated by Streptococcus, Veillonella, and Neisseria and remained stable in time, while the composition of the Control microcosms diverged significantly in time, partially due to the presence of Megasphaera. The percentage of Candida increased 100-fold in the Control microcosms compared to the Arginine microcosms. The pH-raising effect of arginine was confirmed by the pH and ammonium results. The abundances of sagP and arcA were highest in the Arginine microcosms, while the concentration of butyrate was higher in the Control microcosms. We demonstrate that supplementation with arginine serves a health-promoting function; it enhances microcosm resilience toward acidification and suppresses outgrowth of the opportunistic pathogen Candida. Arginine facilitates stability of oral microbial communities and prevents them from becoming cariogenic. PMID:25433583

  5. The subcellular compartmentalization of arginine metabolizing enzymes and their role in endothelial dysfunction

    Directory of Open Access Journals (Sweden)

    Feng eChen

    2013-07-01

    Full Text Available The endothelial production of nitric oxide (NO mediates endothelium-dependent vasorelaxation and restrains vascular inflammation, smooth muscle proliferation and platelet aggregation. Impaired production of NO is a hallmark of endothelial dysfunction and promotes the development of cardiovascular disease. In endothelial cells, NO is generated by endothelial nitric oxide synthase (eNOS through the conversion of its substrate, L-arginine to L-citrulline. Reduced access to L-arginine has been proposed as a major mechanism underlying reduced eNOS activity and NO production in cardiovascular disease. The arginases (Arg1 and Arg2 metabolize L-arginine to generate L-ornithine and urea and increased expression of arginase has been proposed as a mechanism of reduced eNOS activity secondary to the depletion of L-arginine. Indeed, supplemental L-arginine and suppression of arginase activity has been shown to improve endothelium-dependent relaxation and ameliorate cardiovascular disease. However, L-arginine concentrations in endothelial cells remain sufficiently high to support NO synthesis suggesting additional mechanisms. The compartmentalization of intracellular L-arginine into poorly interchangeable pools has been proposed to allow for the local depletion of L-arginine. Indeed the subcellular location of L-arginine metabolizing enzymes plays important functional roles. In endothelial cells, eNOS is found in discrete intracellular locations and the capacity to generate NO is heavily influenced by its localtion. Arg1 and Arg2 also reside in different subcellular environments and are thought to differentially influence endothelial function. The plasma membrane solute transporter, CAT-1 and the arginine recycling enzyme, ASL, co-localize with eNOS and facilitate NO release. This review highlights the importance of the subcellular location of eNOS and arginine transporting and metabolizing enzymes to NO release and cardiovascular disease.

  6. Oral L-arginine supplementation impacts several reproductive parameters during the postpartum period in mares.

    Science.gov (United States)

    Kelley, Dale E; Warren, Lori K; Mortensen, Christopher J

    2013-05-01

    L-arginine is an amino acid which can alter pituitary function and increase blood flow to the reproductive tract. The objective was to determine the effect of supplementing 100g of L-arginine on plasma arginine concentrations, follicular dynamics and ovarian and uterine artery blood flow during the estrus that occurs subsequent to foaling. In Experiment 1, mares were fed 100g L-arginine for 1 day during the last 3 weeks of pregnancy and plasma samples taken for every hour for the first 4h and every other hour until 12h.L-arginine supplementation elevated plasma arginine concentrations from 1 to 8h post feeding; arginine peaked at 6h (arginine: 515±33μmol/L; control: 80±33μmol/L). In Experiment 2, mares received either 100g L-arginine or control diets beginning 21 d before the expected foaling date and continued for 30 d postpartum. The reproductive tract was evaluated by transrectal Doppler ultrasonography from Day 1 postpartum through Day 30. There were no differences in ovarian follicular dynamics, ovarian or uterine resistance indices between groups. Vascular perfusion of the F1 follicular wall was greater in L-arginine supplemented mares (37.3±2.6%) than controls (25.4±2.7%; Pmares had a smaller uterine body and horns and accumulated less uterine fluid than controls (Pfollicular development, raises the possible use of L-arginine supplementation as a breeding management tool during the postpartum period to increase reproductive success. PMID:23523236

  7. Evidence for an Arginine Exporter Encoded by yggA (argO) That Is Regulated by the LysR-Type Transcriptional Regulator ArgP in Escherichia coli

    OpenAIRE

    Nandineni, Madhusudan R.; J.Gowrishankar

    2004-01-01

    The anonymous open reading frame yggA of Escherichia coli was identified in this study as a gene that is under the transcriptional control of argP (previously called iciA), which encodes a LysR-type transcriptional regulator protein. Strains with null mutations in either yggA or argP were supersensitive to the arginine analog canavanine, and yggA-lac expression in vivo exhibited argP+-dependent induction by arginine. Lysine supplementation phenocopied the argP null mutation in that it virtual...

  8. Adverse effects associated with arginine alpha-ketoglutarate containing supplements.

    Science.gov (United States)

    Prosser, J M; Majlesi, N; Chan, G M; Olsen, D; Hoffman, R S; Nelson, L S

    2009-05-01

    The athletic performance supplement industry is a multibillion-dollar business and one popular category claims to increase nitric oxide (NO) production. We report three patients presenting to the emergency department with adverse effects. A 33-year-old man presented with palpitations, dizziness, vomiting, and syncope, after the use of NO(2) platinum. His examination and electrocardiogram (ECG) were normal. The dizziness persisted, requiring admission overnight. A 21-year-old man with palpitations and near syncope had used a "nitric oxide" supplement. He was tachycardic to 115 bpm with otherwise normal examination. Laboratory values including methemoglobin, and ECG were unremarkable. He was treated with 1 L of saline with no change in heart rate. He was admitted for observation. A 24-year-old man presented after taking NO-Xplode with palpitations and a headache. His examination, laboratory values, and ECG were normal. He was discharged. The purported active ingredient in these products is arginine alpha-ketoglutarate (AAKG), which is claimed to increase NO production by supplying the precursor L-arginine. The symptoms could be due to vasodilation from increased levels of NO, though other etiologies cannot be excluded. AAKG containing supplements may be associated with adverse effects requiring hospital admission. PMID:19755457

  9. Cloning and expression of phosphoenolpyruvate carboxykinase from a cestode parasite and its solubilization from inclusion bodies using l-arginine.

    Science.gov (United States)

    Dutta, Asim K; Ramnath; Dkhar, Barilin; Tandon, Veena; Das, Bidyadhar

    2016-09-01

    Phosphoenolpyruvate carboxykinase is an essential regulatory enzyme of glycolysis in the cestode parasite, Raillietina echinobothrida, and is considered a potential target for anthelmintic action because of its differential activity from that of its avian host. However, due to the unavailability of its structure, the mechanism of regulation of PEPCK from R. echinobothrida (rePEPCK) and its interaction with possible modulators remain unclear. Hence, in this study, the rePEPCK gene was cloned into pGEX-4T-3 and overexpressed for its characterization. On being induced by IPTG, the recombinant rePEPCK was expressed as inclusion bodies (IBs); hence, various agents, like different inducer concentrations, temperature, time, host cell types, culture media, pH, and additives, were used to bring the protein to soluble form. Finally, a significant amount (∼46%) of rePEPCK was solubilized from IBs by adding 2M l-arginine. Near-UV circular dichroism spectra analysis indicated that l-arginine (2M) had no effect on the conformation of the protein. In this study, we have reported a yield of ∼73mg of purified rePEPCK per 1L of culture. The purified rePEPCK retained its biological activity, and Km of the enzyme for its substrate was determined and discussed. The availability of recombinant rePEPCK may help in biochemical- and biophysical-studies to explore its molecular mechanisms and regulations. PMID:26363119

  10. TP53 Codon 72 Polymorphism and P53 Protein Expression in Colorectal Cancer Specimens in Isfahan

    Directory of Open Access Journals (Sweden)

    Mehdi Nikbahkt Dastjerdi

    2011-02-01

    Full Text Available The TP53 tumor suppressor gene plays important roles in genomic stability. A common polymorphism at codon 72 of TP53 gene has been associated with increased risk for many human cancers. The p53 protein is expressed in colorectal cancer, but the reported prevalence of its expression varies widely. In the present study, the p53 protein expression in different genotypes of its codon 72 , was investigated. We undertook a case-control study on 250 controls and 250 paraffin block specimens of sporadic colorectal adenocarcinomas from the city of Isfahan. PCR amplification of TP53 codon 72 polymorphism: TP53 codon 72 genotypes were detected by PCR using specific primer pairs for amplifying the proline or the arginine Alleles. The PCR reaction was done separately for each of the two polymorphic variants. The amplified products were subjected to electrophoresis on 1% agarose gel in 1× TBE buffer and visualized on a transilluminator using ethidium bromide. Immunohistochemical Staining: We evaluated the expression patterns of p53 protein, as potential prognostic marker in colorectal cancer specimens by immunohistochemical staining. Statistical analyses: The χ2-test was used to assess the significance of any difference in the prevalence of TP53 codon 72 polymorphism between colorectal cancer patients and controls. The odds ratio and 95% CI (confidence intervals was used as a measure of the strength of the association. Statistical significance level was set to P≤0.05. In control samples, the genotype distribution for TP53 polymorphism showed 30.4%, 45.2% and 24.4% for the arginine/arginine, arginine/proline and proline/proline genotypes, respectively. Allelic frequencies corresponded to 0.663 for the arginine allele and 0.338 for the proline allele. In the cancer group 38.8% of the cases were arginine/arginine, 40.4% were arginine/proline and 20.8% were proline/proline. The corresponding frequencies were 0.590 for the arginine allele and 0.410 for the

  11. Synthesis, characterization and behaviour of trans-bis (argininate) copper (II) to gamma radiation

    International Nuclear Information System (INIS)

    The synthesis, the characterization and the behaviour to gamma radiation of trans-bis (argininate) copper (II) are presented. The synthesis is made from copper sulfate, sodium hydroxide and hydrochloride of L (+) arginine, in aqueous medium, and the characterization by infrared spectroscopy, visible and ultraviolet spectroscopy and elementary analysis. (C.G.C.)

  12. Acute hypothalamic administration of L-arginine increases feed intake in rats

    OpenAIRE

    Carlos Ricardo Maneck Malfatti; Luiz Augusto da Silva; Ricardo Aparecido Pereira; Renan Garcia Michel; André Luiz Snak; Fabio Seidel dos Santos

    2015-01-01

    Objective: This study investigated the chronic (oral) and acute (hypothalamic infusion) effects of L-arginine supplementation on feed intake, body composition, and behavioral changes in rats. Methods: Twenty rats were divided into two groups treated orally for 60 days; one group received L-arginine (1 g/kg body weight) and one group received saline (1 mL/NaCl ...

  13. Facilitation of peptide fibre formation by arginine-phosphate/carboxylate interactions

    Indian Academy of Sciences (India)

    K Krishna Prasad; Sandeep Verma

    2008-01-01

    This study describes peptide fibre formation in a hexapeptide, derived from the V3 loop of HIV-1, mediated by the interactions between arginine residues and phosphate/carboxylate anions. This charge neutralization approach was further confirmed when the deletion of arginine residue from the hexapeptide sequence resulted in fibre formation, which was studied by a combination of microscopic techniques.

  14. Antibacterial action of a novel functionalized chitosan-arginine against Gram-negative bacteria.

    Science.gov (United States)

    Tang, Hong; Zhang, Peng; Kieft, Thomas L; Ryan, Shannon J; Baker, Shenda M; Wiesmann, William P; Rogelj, Snezna

    2010-07-01

    The antimicrobial activity of chitosan and chitosan derivatives has been well established. However, although several mechanisms have been proposed, the exact mode of action is still unclear. Here we report on the investigation of antibacterial activity and the antibacterial mode of action of a novel water-soluble chitosan derivative, arginine-functionalized chitosan, on the Gram-negative bacteria Pseudomonas fluorescens and Escherichia coli. Two different arginine-functionalized chitosans (6% arginine-substituted and 30% arginine-substituted) each strongly inhibited P. fluorescens and E. coli growth. Time-dependent killing efficacy experiments showed that 5000 mg l(-1) of 6%- and 30%-substituted chitosan-arginine killed 2.7 logs and 4.5 logs of P. fluorescens, and 4.8 logs and 4.6 logs of E. coli in 4h, respectively. At low concentrations, the 6%-substituted chitosan-arginine was more effective in inhibiting cell growth even though the 30%-substituted chitosan-arginine appeared to be more effective in permeabilizing the cell membranes of both P. fluorescens and E. coli. Studies using fluorescent probes, 1-N-phenyl-naphthylamine (NPN), nile red (NR) and propidium iodide (PI), and field emission scanning electron microscopy (FESEM) suggest that chitosan-arginine's antibacterial activity is, at least in part, due to its interaction with the cell membrane, in which it increases membrane permeability. PMID:20060936

  15. Endothelial arginine resynthesis contributes to the maintenance of vasomotor function in male diabetic mice

    DEFF Research Database (Denmark)

    Chennupati, Ramesh; Meens, Merlijn J P M T; Marion, Vincent;

    2014-01-01

    : Absence of endothelial citrulline recycling to arginine did not affect blood pressure and systemic arterial vasomotor responses in healthy mice. EDNO-mediated vasodilatation was significantly more impaired in diabetic Ass-KOTie2 than in control mice demonstrating that endothelial arginine recycling...

  16. Arginine Catabolism by Sourdough Lactic Acid Bacteria: Purification and Characterization of the Arginine Deiminase Pathway Enzymes from Lactobacillus sanfranciscensis CB1

    OpenAIRE

    De Angelis, Maria; Mariotti, Liberato; Rossi, Jone; Servili, Maurizio; Fox, Patrick F.; Rollán, Graciela; Gobbetti, Marco

    2002-01-01

    The cytoplasmic extracts of 70 strains of the most frequently isolated sourdough lactic acid bacteria were screened initially for arginine deiminase (ADI), ornithine transcarbamoylase (OTC), and carbamate kinase (CK) activities, which comprise the ADI (or arginine dihydrolase) pathway. Only obligately heterofermentative strains such as Lactobacillus sanfranciscensis CB1; Lactobacillus brevis AM1, AM8, and 10A; Lactobacillus hilgardii 51B; and Lactobacillus fructivorans DD3 and DA106 showed al...

  17. Eukaryotic LYR Proteins Interact with Mitochondrial Protein Complexes

    OpenAIRE

    Heike Angerer

    2015-01-01

    In eukaryotic cells, mitochondria host ancient essential bioenergetic and biosynthetic pathways. LYR (leucine/tyrosine/arginine) motif proteins (LYRMs) of the Complex1_LYR-like superfamily interact with protein complexes of bacterial origin. Many LYR proteins function as extra subunits (LYRM3 and LYRM6) or novel assembly factors (LYRM7, LYRM8, ACN9 and FMC1) of the oxidative phosphorylation (OXPHOS) core complexes. Structural insights into complex I accessory subunits LYRM6 and LYRM3 have bee...

  18. Supplementation with l-arginine stabilizes plasma arginine and nitric oxide metabolites, suppresses elevated liver enzymes and peroxidation in sickle cell anaemia.

    Science.gov (United States)

    Jaja, S I; Ogungbemi, S O; Kehinde, M O; Anigbogu, C N

    2016-06-01

    The effect of l-arginine on liver function in SCD has received little or no attention. The effect of a chronic, oral, low-dose supplementation with l-arginine (1gm/day for 6 weeks) on some liver enzymes, lipid peroxidation and nitric oxide metabolites was studied in 20 normal (non-sickle cell anaemia; NSCA) subjects and 20 sickle cell anaemia (SCA) subjects. Ten milliliters of blood was withdrawn from an ante-cubital vein for the estimation of plasma arginine concentration ([R]), alanine aminotransaminase (ALT), aspartate aminotransaminase (AST) and alkaline phosphatase (ALP), plasma total bilirubin concentration [TB], malondialdehyde concentration [MDA] and nitric oxide metabolites concentration [NOx]. Before supplementation, ALT, AST, ALP (pNOx] were higher in NSCA subjects (pNOX] in SCA than in NSCA subjects (plow-dose supplementation with l-arginine improved liver function, oxidative stress, plasma arginine concentration and nitric oxide metabolites levels in NSCA and SCA subjects. Responses in SCA subjects to l-arginine were more sensitive than in NSCA subjects. PMID:27156372

  19. Potentiality of application of the conductometric L-arginine biosensors for the real sample analysis

    Directory of Open Access Journals (Sweden)

    Jaffrezic-Renault N.

    2012-12-01

    Full Text Available Aim. To determine an influence of serum components on the L-arginine biosensor sensitivity and to formulate practical recommendations for its reliable analysis. Methods. The L-arginine biosensor comprised arginase and urease co-immobilized by cross-linking. Results. The biosensor specificity was investigated based on a series of representative studies (namely, through urea determination in the serum; inhibitory effect studies of mercury ions; high temperature treatment of sensors; studying the biosensor sensitivity to the serum treated by enzymes, and selectivity studies. It was found that the response of the biosensor to the serum injections was determined by high sensitivity of the L-arginine biosensor toward not only to L-arginine but also toward two other basic amino acids (L-lysine and L-histidine. Conclusions. A detailed procedure of optimization of the conductometric biosensor for L-arginine determination in blood serum has been proposed.

  20. Sequestration and metabolism of host cell arginine by the intraerythrocytic malaria parasite Plasmodium falciparum.

    Science.gov (United States)

    Cobbold, Simon A; Llinás, Manuel; Kirk, Kiaran

    2016-06-01

    Human erythrocytes have an active nitric oxide synthase, which converts arginine into citrulline and nitric oxide (NO). NO serves several important functions, including the maintenance of normal erythrocyte deformability, thereby ensuring efficient passage of the red blood cell through narrow microcapillaries. Here, we show that following invasion by the malaria parasite Plasmodium falciparum the arginine pool in the host erythrocyte compartment is sequestered and metabolized by the parasite. Arginine from the extracellular medium enters the infected cell via endogenous host cell transporters and is taken up by the intracellular parasite by a high-affinity cationic amino acid transporter at the parasite surface. Within the parasite arginine is metabolized into citrulline and ornithine. The uptake and metabolism of arginine by the parasite deprive the erythrocyte of the substrate required for NO production and may contribute to the decreased deformability of infected erythrocytes. PMID:26633083

  1. Differential effects of cranial radiation on growth hormone response to arginine and insulin infusion

    International Nuclear Information System (INIS)

    The growth hormone responses to arginine infusion and to insulin-induced hypoglycemia were studied in 13 patients with neoplastic disease after treatment with radiation and chemotherapy. Patients who received intensive cranial radiation (greater than 2,400 rads) had no response to either arginine or insulin; those who received moderate cranial radiation (greater than or equal to 2,400 rads) had GH response to arginine but not to insulin; patients receiving no cranial radiation responded to both arginine and insulin. These data support the hypothesis that GH secretion in response to arginine infusion has a different mechanism in contrast to the response to insulin-induced hypoglycemia and that the latter is more vulnerable to cranial radiation

  2. Life on arginine for Mycoplasma hominis: clues from its minimal genome and comparison with other human urogenital mycoplasmas.

    Directory of Open Access Journals (Sweden)

    Sabine Pereyre

    2009-10-01

    Full Text Available Mycoplasma hominis is an opportunistic human mycoplasma. Two other pathogenic human species, M. genitalium and Ureaplasma parvum, reside within the same natural niche as M. hominis: the urogenital tract. These three species have overlapping, but distinct, pathogenic roles. They have minimal genomes and, thus, reduced metabolic capabilities characterized by distinct energy-generating pathways. Analysis of the M. hominis PG21 genome sequence revealed that it is the second smallest genome among self-replicating free living organisms (665,445 bp, 537 coding sequences (CDSs. Five clusters of genes were predicted to have undergone horizontal gene transfer (HGT between M. hominis and the phylogenetically distant U. parvum species. We reconstructed M. hominis metabolic pathways from the predicted genes, with particular emphasis on energy-generating pathways. The Embden-Meyerhoff-Parnas pathway was incomplete, with a single enzyme absent. We identified the three proteins constituting the arginine dihydrolase pathway. This pathway was found essential to promote growth in vivo. The predicted presence of dimethylarginine dimethylaminohydrolase suggested that arginine catabolism is more complex than initially described. This enzyme may have been acquired by HGT from non-mollicute bacteria. Comparison of the three minimal mollicute genomes showed that 247 CDSs were common to all three genomes, whereas 220 CDSs were specific to M. hominis, 172 CDSs were specific to M. genitalium, and 280 CDSs were specific to U. parvum. Within these species-specific genes, two major sets of genes could be identified: one including genes involved in various energy-generating pathways, depending on the energy source used (glucose, urea, or arginine and another involved in cytadherence and virulence. Therefore, a minimal mycoplasma cell, not including cytadherence and virulence-related genes, could be envisaged containing a core genome (247 genes, plus a set of genes required for

  3. Dimethyl fumarate protects pancreatic islet cells and non-endocrine tissue in L-arginine-induced chronic pancreatitis.

    Directory of Open Access Journals (Sweden)

    Lourdes Robles

    Full Text Available Chronic pancreatitis (CP is a progressive disorder resulting in the destruction and fibrosis of the pancreatic parenchyma which ultimately leads to impairment of the endocrine and exocrine functions. Dimethyl Fumarate (DMF was recently approved by FDA for treatment of patients with multiple sclerosis. DMF's unique anti-oxidant and anti-inflammatory properties make it an interesting drug to test on other inflammatory conditions. This study was undertaken to determine the effects of DMF on islet cells and non-endocrine tissue in a rodent model of L-Arginine-induced CP.Male Wistar rats fed daily DMF (25 mg/kg or vehicle by oral gavage were given 5 IP injections of L-Arginine (250 mg/100 g × 2, 1 hr apart. Rats were assessed with weights and intra-peritoneal glucose tolerance tests (IPGTT, 2 g/kg. Islets were isolated and assessed for islet mass and viability with flow cytometry. Non-endocrine tissue was assessed for histology, myeloperoxidase (MPO, and lipid peroxidation level (MDA. In vitro assessments included determination of heme oxygenase (HO-1 protein expression by Western blot.Weight gain was significantly reduced in untreated CP group at 6 weeks. IPGTT revealed significant impairment in untreated CP group and its restoration with DMF therapy (P <0.05. Untreated CP rats had pancreatic atrophy, severe acinar architectural damage, edema, and fatty infiltration as well as elevated MDA and MPO levels, which were significantly improved by DMF treatment. After islet isolation, the volume of non-endocrine tissue was significantly smaller in untreated CP group. Although islet counts were similar in the two groups, islet viability was significantly reduced in untreated CP group and improved with DMF treatment. In vitro incubation of human pancreatic tissue with DMF significantly increased HO-1 expression.Administration of DMF attenuated L-Arginine-induced CP and islet function in rats. DMF treatment could be a possible strategy to improve clinical

  4. Effect of oral take L-arginine on [Ca2+]i of human platelets

    International Nuclear Information System (INIS)

    To study the effect of oral take L-arginine on [Ca2+]i of human platelet, ten male volunteers were selected to take L-arginine for 4 days (20g/three times/a day, the 4th day took about 7g). They are neither smokers nor drinkers and no drugs were taken by them during 2 weeks before study. Tests were performed before and after taking L-arginine. Before and after taking L-arginine, platelet intracellular cGMP were 1.91 ± 0.20 vs 2.14 ± 0.34pmol/109 platelets, P 2+]i (A/B value: 0.45 ± 0.14 vs 0.32 ± 0.09) were inhibited. The results has showed that oral take L-arginine could inhibit platelet activation. This inhibition should be ascribed to such two points: (1) The amount of NO released by endothelial cells has increased. A basal release of NO has been found to regulate the blood vessel intensity and inhibit platelet adhesion. After taking L-arginine, the endothelial cells released more NO and the plasma NO3- level also increased. (2) The platelet has synthesized more NO. There is an L-arginine/NO pathway in platelet and platelet could synthesize NO from L-arginine to inhibit its activation. Taking L-arginine has enhanced the negative effect of this pathway by producing more NO. The study will be helpful to further study on the possibility of oral take L-arginine to avoid thrombotic and hemorrhagic diseases

  5. Biochemical Characterization of An Arginine-Specific Alkaline Trypsin from Bacillus licheniformis

    Directory of Open Access Journals (Sweden)

    Jin-Song Gong

    2015-12-01

    Full Text Available In the present study, we isolated a trypsin-producing strain DMN6 from the leather waste and identified it as Bacillus licheniformis through a two-step screening strategy. The trypsin activity was increased up to 140 from 20 U/mL through culture optimization. The enzyme was purified to electrophoretic homogeneity with a molecular mass of 44 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the specific activity of purified enzyme is 350 U/mg with Nα-Benzoyl-l-arginine ethylester as the substrate. The optimum temperature and pH for the trypsin are 65 °C and pH 9.0, respectively. Also, the enzyme can be significantly activated by Ba2+. This enzyme is relatively stable in alkaline environment and displays excellent activity at low temperatures. It could retain over 95% of enzyme activity after 180 min of incubation at 45 °C. The distinguished activity under low temperature and prominent stability enhance its catalytic potential. In the current work, the open reading frame was obtained with a length of 1371 nucleotides that encoded a protein of 456 amino acids. These data would warrant the B. licheniformis trypsin as a promising candidate for catalytic application in collagen preparation and leather bating through further protein engineering.

  6. Evidence that L-Arginine inhibits glycation of human serum albumin (HSA) in vitro

    International Nuclear Information System (INIS)

    Previous work by Brownlee has shown that glycation of bovine serum albumin can be reduced in the presence of aminoguanidine (AG). Presumably, the guanidinium group on AG interferes with further rearrangement of amadori products to advanced glycosylated end products (AGE). Since L-arginine (ARG) also contains a guanidinium group, its ability to inhibit the formation of AGE products was investigated. HSA was incubated at 37 degrees C in the presence or absence of glucose; with glucose and fructose; or with sugars in the presence or absence of ARG or AG. A tracer amount of U-14C-glucose was added to each tube containing sugars. Protein bound glucose was separated from unreacted glucose by gel filtration. Radioactivity, total protein, fluorescence, and glucose concentration were measured. Preliminary data show enhanced binding of 14C-glucose to HSA with fructose at all time points. A 30-40% decrease in 14C-glucose incorporation was observed when ARG or AG as present. ARG and AG were equally effective in inhibiting incorporation of 14C-glucose. FPLC analysis is in progress to determine the type and degree of HSA crosslinking during the 2 week incubation period

  7. Effects of "Bioactive" amino acids leucine, glutamate, arginine and tryptophan on feed intake and mRNA expression of relative neuropeptides in broiler chicks

    Directory of Open Access Journals (Sweden)

    Wang Songbo

    2012-08-01

    Full Text Available Abstract Feed intake control is vital to ensuring optimal nutrition and achieving full potential for growth and development in poultry. The aim of the present study was to investigate the effects of L-leucine, L-glutamate, L-tryptophan and L-arginine on feed intake and the mRNA expression levels of hypothalamic Neuropeptide involved in feed intake regulation in broiler chicks. Leucine, glutamate, tryptophan or arginine was intra-cerebroventricularly (ICV administrated to 4d-old broiler chicks respectively and the feed intake were recorded at various time points. Quantitative PCR was performed to determine the hypothalamic mRNA expression levels of Neuropeptide Y (NPY, agouti related protein (AgRP, pro-opiomelanocortin (POMC, melanocortin receptor 4 (MC4R and corticotrophin releasing factor (CRF. Our results showed that ICV administration of L-leucine (0.15 or 1.5  μmol significantly (P P 

  8. Altered Nitrogen Balance and Decreased Urea Excretion in Male Rats Fed Cafeteria Diet Are Related to Arginine Availability

    Directory of Open Access Journals (Sweden)

    David Sabater

    2014-01-01

    rats, but low arginine levels point to a block in the urea cycle between ornithine and arginine, thereby preventing the elimination of excess nitrogen as urea. The ultimate consequence of this paradoxical block in the urea cycle seems to be the limitation of arginine production and/or availability.

  9. Loading of PAX3 to Mitotic Chromosomes Is Mediated by Arginine Methylation and Associated with Waardenburg Syndrome.

    Science.gov (United States)

    Wu, Tsu-Fang; Yao, Ya-Li; Lai, I-Lu; Lai, Chien-Chen; Lin, Pei-Lun; Yang, Wen-Ming

    2015-08-14

    PAX3 is a transcription factor critical to gene regulation in mammalian development. Mutations in PAX3 are associated with Waardenburg syndrome (WS), but the mechanism of how mutant PAX3 proteins cause WS remains unclear. Here, we found that PAX3 loads on mitotic chromosomes using its homeodomain. PAX3 WS mutants with mutations in homeodomain lose the ability to bind mitotic chromosomes. Moreover, loading of PAX3 on mitotic chromosomes requires arginine methylation, which is regulated by methyltransferase PRMT5 and demethylase JMJD6. Mutant PAX3 proteins that lose mitotic chromosome localization block cell proliferation and normal development of zebrafish. These results reveal the molecular mechanism of PAX3s loading on mitotic chromosomes and the importance of this localization pattern in normal development. Our findings suggest that PAX3 WS mutants interfere with the normal functions of PAX3 in a dominant negative manner, which is important to the understanding of the pathogenesis of Waardenburg syndrome. PMID:26149688

  10. Loading of PAX3 to Mitotic Chromosomes Is Mediated by Arginine Methylation and Associated with Waardenburg Syndrome*

    Science.gov (United States)

    Wu, Tsu-Fang; Yao, Ya-Li; Lai, I-Lu; Lai, Chien-Chen; Lin, Pei-Lun; Yang, Wen-Ming

    2015-01-01

    PAX3 is a transcription factor critical to gene regulation in mammalian development. Mutations in PAX3 are associated with Waardenburg syndrome (WS), but the mechanism of how mutant PAX3 proteins cause WS remains unclear. Here, we found that PAX3 loads on mitotic chromosomes using its homeodomain. PAX3 WS mutants with mutations in homeodomain lose the ability to bind mitotic chromosomes. Moreover, loading of PAX3 on mitotic chromosomes requires arginine methylation, which is regulated by methyltransferase PRMT5 and demethylase JMJD6. Mutant PAX3 proteins that lose mitotic chromosome localization block cell proliferation and normal development of zebrafish. These results reveal the molecular mechanism of PAX3s loading on mitotic chromosomes and the importance of this localization pattern in normal development. Our findings suggest that PAX3 WS mutants interfere with the normal functions of PAX3 in a dominant negative manner, which is important to the understanding of the pathogenesis of Waardenburg syndrome. PMID:26149688

  11. Biosynthesis of homoarginine (hArg) and asymmetric dimethylarginine (ADMA) from acutely and chronically administered free L-arginine in humans.

    Science.gov (United States)

    Kayacelebi, Arslan Arinc; Langen, Jennifer; Weigt-Usinger, Katharina; Chobanyan-Jürgens, Kristine; Mariotti, François; Schneider, Jessica Y; Rothmann, Sabine; Frölich, Jürgen C; Atzler, Dorothee; Choe, Chi-Un; Schwedhelm, Edzard; Huneau, Jean François; Lücke, Thomas; Tsikas, Dimitrios

    2015-09-01

    Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide (NO) synthesis, whereas L-arginine (Arg) and L-homoarginine (hArg) serve as substrates for NO synthesis. ADMA and other methylated arginines are generally believed to exclusively derive from guanidine (N (G))-methylated arginine residues in proteins by protein arginine methyltransferases (PRMTs) that use S-adenosylmethionine (SAM) as the methyl donor. L-Lysine is known for decades as a precursor for hArg, but only recent studies indicate that arginine:glycine amidinotransferase (AGAT) is responsible for the synthesis of hArg. AGAT catalyzes the formation of guanidinoacetate (GAA) that is methylated to creatine by guanidinoacetate methyltransferase (GAMT) which also uses SAM. The aim of the present study was to learn more about the mechanisms of ADMA and hArg formation in humans. Especially, we hypothesized that ADMA is produced by N (G)-methylation of free Arg in addition to the known PRMTs-involving mechanism. In knockout mouse models of AGAT- and GAMT-deficiency, we investigated the contribution of these enzymes to hArg synthesis. Arg infusion (0.5 g/kg, 30 min) in children (n = 11) and ingestion of high-fat protein meals by overweight men (n = 10) were used to study acute effects on ADMA and hArg synthesis. Daily Arg ingestion (10 g) or placebo for 3 or 6 months by patients suffering from peripheral arterial occlusive disease (PAOD, n = 20) or coronary artery disease (CAD, n = 30) was used to study chronic effects of Arg on ADMA synthesis. Mass spectrometric methods were used to measure all biochemical parameters in plasma and urine samples. In mice, AGAT but not GAMT was found to contribute to plasma hArg, while ADMA synthesis was independent of AGAT and GAMT. Arg infusion acutely increased plasma Arg, hArg and ADMA concentrations, but decreased the plasma hArg/ADMA ratio. High-fat protein meals acutely increased plasma Arg, hArg, ADMA concentrations, as well as the plasma h

  12. Microwave heating of arginine yields highly fluorescent nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Philippidis, Aggelos [Foundation for Research and Technology-Hellas, Institute of Electronic Structure and Laser (Greece); Stefanakis, Dimitrios [University of Crete, Department of Chemistry (Greece); Anglos, Demetrios, E-mail: anglos@iesl.forth.gr [Foundation for Research and Technology-Hellas, Institute of Electronic Structure and Laser (Greece); Ghanotakis, Demetrios, E-mail: ghanotakis@chemistry.uoc.gr [University of Crete, Department of Chemistry (Greece)

    2013-01-15

    Brightly fluorescent nanoparticles were produced via a single-step, single-precursor procedure based on microwave heating of an aqueous solution of the amino acid arginine. Key structural and optical properties of the resulting Arg nanoparticles, Arg-dots, are reported and discussed with emphasis on the pH dependence of their fluorescence emission. The surface of the Arg-dots was functionalised through coupling to folic acid, opening up ways for connecting fluorescent nanoparticles to cancer cells. The generality and versatility of the microwave heating procedure was further demonstrated by the synthesis of different types of carbon nanoparticles, such as CE-dots, that were produced by use of citric acid and ethanolamine as precursors and compared to the Arg-dots.

  13. Microwave heating of arginine yields highly fluorescent nanoparticles

    International Nuclear Information System (INIS)

    Brightly fluorescent nanoparticles were produced via a single-step, single-precursor procedure based on microwave heating of an aqueous solution of the amino acid arginine. Key structural and optical properties of the resulting Arg nanoparticles, Arg-dots, are reported and discussed with emphasis on the pH dependence of their fluorescence emission. The surface of the Arg-dots was functionalised through coupling to folic acid, opening up ways for connecting fluorescent nanoparticles to cancer cells. The generality and versatility of the microwave heating procedure was further demonstrated by the synthesis of different types of carbon nanoparticles, such as CE-dots, that were produced by use of citric acid and ethanolamine as precursors and compared to the Arg-dots.

  14. Polymerization on the rocks: beta-amino acids and arginine

    Science.gov (United States)

    Liu, R.; Orgel, L. E.; Bada, J. L. (Principal Investigator)

    1998-01-01

    We have studied the accumulation of long oligomers of beta-amino acids on the surface of minerals using the 'polymerization on the rocks' protocol. We find that long oligopeptides of beta-glutamic acid which cannot be formed in homogeneous aqueous solution are accumulated efficiently on the surface of hydroxylapatite using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDAC) as condensing agent. The EDAC-induced oligomerization of aspartic acid on hydroxylapatite proceeds even more efficiently. Hydroxylapatite can also facilitate the ligation of the tripeptide (glu)3. The 'polymerization on the rocks' scenario is not restricted to negatively-charged amino acids. Oligoarginines are accumulated on the surface of illite using carbonyldiimidizole (CDI) as condensing agent. We find that FeS2 catalyzes the CDI-induced oligomerization of arginine, although it does not adsorb oligoarginines. These results are relevant to the formation of polypeptides on the primitive earth.

  15. The functional relationship between the Cdc50p-Drs2p putative aminophospholipid translocase and the Arf GAP Gcs1p in vesicle formation in the retrieval pathway from yeast early endosomes to the TGN.

    OpenAIRE

    Sakane, Hiroshi; Yamamoto, Takaharu; Tanaka, Kazuma

    2006-01-01

    Drs2p, the catalytic subunit of the Cdc50p-Drs2p putative aminophospholipid translocase, has been implicated in conjunction with the Arf1 signaling pathway in the formation of clathrin-coated vesicles (CCVs) from the TGN. Herein, we searched for Arf regulator genes whose mutations were synthetically lethal with cdc50Δ, and identified the Arf GAP gene GCS1. Most of the examined transport pathways in the Cdc50p-depleted gcs1Δ mutant were nearly normal, including endocytic transport to vacuoles,...

  16. Arginine becomes an essential amino acid after massive resection of rat small intestine.

    Science.gov (United States)

    Wakabayashi, Y; Yamada, E; Yoshida, T; Takahashi, H

    1994-12-23

    We compared effects of feeding arginine- and/or proline- deficient diets (-Arg, -Pro, and -Arg, Pro) with those of a complete diet (Complete) in rats whose small intestine had been massively resected. After 4 weeks, the rats fed -Arg and -Arg, Pro lost weight (a mean of 28 and 32 g, respectively), whereas those fed Complete and -Pro gained 80 and 58 g, respectively. The average nitrogen balance was about 117,100, -20 and -14 mg/day for Complete, -Pro, -Arg, and -Arg, Pro diets, respectively. The concentration of arginine in skeletal muscle was about 310, 330, 91, and 65 nmol/g for Complete, -Pro, -Arg, and -Arg, Pro, respectively; while plasma arginine concentration averaged 95, 107, 56, and 46 microM, respectively. The weight loss, the negative nitrogen balance, and the markedly reduced arginine concentration in the muscle observed in rats fed -Arg and -Arg, Pro clearly indicate that arginine becomes a strictly essential amino acid in the rats with massive resection of the small intestine. However, sufficient proline can be synthesized from arginine in tissues such as the liver and kidney in the absence of the small intestine. Plasma glutamine, citrulline in the muscle and plasma, urinary excretion of orotic acid and nitrate (to assess nitric oxide formation from arginine) were also measured, and the changes in these metabolites are discussed. PMID:7798273

  17. One-pot green synthesis of biocompatible arginine-stabilized magnetic nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Wang Zhongjun; Zhu Hui; Wang Xiaolei; Yang Fan; Yang Xiurong, E-mail: xryang@ciac.jl.c [State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun, Jilin, 130022 (China)

    2009-11-18

    A green one-step approach has been developed for the synthesis of amino-functionalized magnetite nanoparticles. The synthesis was accomplished by simply mixing FeCl{sub 2} with arginine under ambient conditions. It was found that the Fe{sup 2+}/arginine molar ratio, reaction duration and temperature greatly influence the size, morphology and composition of magnetic nanoparticles. The arginine-stabilized magnetic nanoparticles were characterized by transmission electron microscopy, x-ray diffraction, x-ray photoelectron spectroscopy, thermogravimetric analysis, and Fourier transform infrared spectroscopy techniques. The results show that the prepared nanoparticles are spherically shaped with a nearly uniform size distribution and pure magnetite phase. The presence of arginine on the magnetic nanoparticle surface has been confirmed and the amount of surface arginine varies with the Fe{sup 2+}/arginine molar ratio. The surface amine densities are calculated to be 5.60 and 7.84 {mu}mol mg{sup -1} for magnetic nanoparticles prepared at 1:1 and 1:2 Fe{sup 2+}/arginine molar ratio, respectively. The as-synthesized nanoparticles show superparamagnetic behavior at room temperature and good solubility in water. In addition, using a similar synthesis procedure, we have been able to synthesize superparamagnetic manganese and cobalt ferrite nanoparticles.

  18. One-pot green synthesis of biocompatible arginine-stabilized magnetic nanoparticles

    International Nuclear Information System (INIS)

    A green one-step approach has been developed for the synthesis of amino-functionalized magnetite nanoparticles. The synthesis was accomplished by simply mixing FeCl2 with arginine under ambient conditions. It was found that the Fe2+/arginine molar ratio, reaction duration and temperature greatly influence the size, morphology and composition of magnetic nanoparticles. The arginine-stabilized magnetic nanoparticles were characterized by transmission electron microscopy, x-ray diffraction, x-ray photoelectron spectroscopy, thermogravimetric analysis, and Fourier transform infrared spectroscopy techniques. The results show that the prepared nanoparticles are spherically shaped with a nearly uniform size distribution and pure magnetite phase. The presence of arginine on the magnetic nanoparticle surface has been confirmed and the amount of surface arginine varies with the Fe2+/arginine molar ratio. The surface amine densities are calculated to be 5.60 and 7.84 μmol mg-1 for magnetic nanoparticles prepared at 1:1 and 1:2 Fe2+/arginine molar ratio, respectively. The as-synthesized nanoparticles show superparamagnetic behavior at room temperature and good solubility in water. In addition, using a similar synthesis procedure, we have been able to synthesize superparamagnetic manganese and cobalt ferrite nanoparticles.

  19. Functional variation in the arginine vasopressin 2 receptor as a modifier of human plasma von Willebrand factor levels

    DEFF Research Database (Denmark)

    Nossent, Anne Yaël; Robben, J H; Deen, P M T; Vos, H L; Rosendaal, F R; Doggen, C J M; Hansen, J L; Sheikh, S P; Bertina, R M; Eikenboom, J C J

    2010-01-01

    -green fluorescent protein (GFP) cells, which is in accordance with increased levels of VWF propeptide associated with the 12E variant. The dissociation constant (K(D)) was 4.5 nm [95% confidence interval (CI) 3.6-5.4] for 12E-V2R-GFP and 16.5 nm (95% CI 10.1-22.9) for 12G-V2R-GFP. AVP-induced cAMP generation was......SUMMARY OBJECTIVES: Stimulation of arginine vasopressin 2 receptor (V2R) with arginine vasopressin (AVP) results in a rise in von Willebrand factor (VWF) and factor VIII plasma levels. We hypothesized that gain-of-function variations in the V2R gene (AVPR2) would lead to higher plasma levels of VWF...... levels. The functionality of the G12E variant was studied in stably transfected MDCKII cells, expressing constructs of either 12G-V2R or 12E-V2R. Both V2R variants were fully glycosylated and expressed on the basolateral membrane. The binding affinity of V2R for AVP was increased three-fold in 12E-V2R...

  20. Tripartite ATP-independent Periplasmic (TRAP) Transporters Use an Arginine-mediated Selectivity Filter for High Affinity Substrate Binding.

    Science.gov (United States)

    Fischer, Marcus; Hopkins, Adam P; Severi, Emmanuele; Hawkhead, Judith; Bawdon, Daniel; Watts, Andrew G; Hubbard, Roderick E; Thomas, Gavin H

    2015-11-01

    Tripartite ATP-independent periplasmic (TRAP) transporters are secondary transporters that have evolved an obligate dependence on a substrate-binding protein (SBP) to confer unidirectional transport. Different members of the DctP family of TRAP SBPs have binding sites that recognize a diverse range of organic acid ligands but appear to only share a common electrostatic interaction between a conserved arginine and a carboxylate group in the ligand. We investigated the significance of this interaction using the sialic acid-specific SBP, SiaP, from the Haemophilus influenzae virulence-related SiaPQM TRAP transporter. Using in vitro, in vivo, and structural methods applied to SiaP, we demonstrate that the coordination of the acidic ligand moiety of sialic acid by the conserved arginine (Arg-147) is essential for the function of the transporter as a high affinity scavenging system. However, at high substrate concentrations, the transporter can function in the absence of Arg-147 suggesting that this bi-molecular interaction is not involved in further stages of the transport cycle. As well as being required for high affinity binding, we also demonstrate that the Arg-147 is a strong selectivity filter for carboxylate-containing substrates in TRAP transporters by engineering the SBP to recognize a non-carboxylate-containing substrate, sialylamide, through water-mediated interactions. Together, these data provide biochemical and structural support that TRAP transporters function predominantly as high affinity transporters for carboxylate-containing substrates. PMID:26342690

  1. Asymmetric arginine dimethylation of RelA provides a repressive mark to modulate TNFα/NF-κB response.

    Science.gov (United States)

    Reintjes, Anja; Fuchs, Julian E; Kremser, Leopold; Lindner, Herbert H; Liedl, Klaus R; Huber, Lukas A; Valovka, Taras

    2016-04-19

    Nuclear factor kappa B (NF-κB) is an inducible transcription factor that plays critical roles in immune and stress responses and is often implicated in pathologies, including chronic inflammation and cancer. Although much has been learned about NF-κB-activating pathways, the specific repression of NF-κB is far less well understood. Here we identified the type I protein arginine methyltransferase 1 (PRMT1) as a restrictive factor controlling TNFα-induced activation of NF-κB. PRMT1 forms a cellular complex with NF-κB through direct interaction with the Rel homology domain of RelA. We demonstrate that PRMT1 methylates RelA at evolutionary conserved R30, located in the DNA-binding L1 loop, which is a critical residue required for DNA binding. Asymmetric R30 dimethylation inhibits the binding of RelA to DNA and represses NF-κB target genes in response to TNFα. Molecular dynamics simulations of the DNA-bound RelA:p50 predicted structural changes in RelA caused by R30 methylation or a mutation that interferes with the stability of the DNA-NF-κB complex. Our findings provide evidence for the asymmetric arginine dimethylation of RelA and unveil a unique mechanism controlling TNFα/NF-κB signaling. PMID:27051065

  2. The effect of L-arginine on guinea-pig and rabbit airway smooth muscle function in vitro

    OpenAIRE

    Perez A.C.; Paul W.; Harrison S.; Page C.P.; Spina D.

    1998-01-01

    We have investigated the effects of L-arginine, D-arginine and L-lysine on airway smooth muscle responsiveness to spasmogens in vitro. Both L-arginine and D-arginine (100 mM) significantly reduced the contractile potency and maximal contractile response to histamine but not to methacholine or potassium chloride in guinea-pig epithelium-denuded isolated trachea. Similarly, the contractile response to histamine was significantly reduced by L-arginine (100 mM) in rabbit epithelium-denuded isolat...

  3. Hepatic adaptation compensates inactivation of intestinal arginine biosynthesis in suckling mice.

    Directory of Open Access Journals (Sweden)

    Vincent Marion

    Full Text Available Suckling mammals, including mice, differ from adults in the abundant expression of enzymes that synthesize arginine from citrulline in their enterocytes. To investigate the importance of the small-intestinal arginine synthesis for whole-body arginine production in suckling mice, we floxed exon 13 of the argininosuccinate synthetase (Ass gene, which codes for a key enzyme in arginine biosynthesis, and specifically and completely ablated Ass in enterocytes by crossing Ass (fl and Villin-Cre mice. Unexpectedly, Ass (fl/fl /VilCre (tg/- mice showed no developmental impairments. Amino-acid fluxes across the intestine, liver, and kidneys were calculated after determining the blood flow in the portal vein, and hepatic and renal arteries (86%, 14%, and 33%, respectively, of the transhepatic blood flow in 14-day-old mice. Relative to control mice, citrulline production in the splanchnic region of Ass (fl/fl /VilCre (tg/- mice doubled, while arginine production was abolished. Furthermore, the net production of arginine and most other amino acids in the liver of suckling control mice declined to naught or even changed to consumption in Ass (fl/fl /VilCre (tg/- mice, and had, thus, become remarkably similar to that of post-weaning wild-type mice, which no longer express arginine-biosynthesizing enzymes in their small intestine. The adaptive changes in liver function were accompanied by an increased expression of genes involved in arginine metabolism (Asl, Got1, Gpt2, Glud1, Arg1, and Arg2 and transport (Slc25a13, Slc25a15, and Slc3a2, whereas no such changes were found in the intestine. Our findings suggest that the genetic premature deletion of arginine synthesis in enterocytes causes a premature induction of the post-weaning pattern of amino-acid metabolism in the liver.

  4. Mechanism of Allosteric Inhibition of N-Acetyl-L-glutamate Synthase by L-Arginine*

    OpenAIRE

    Min, Li; Jin, Zhongmin; Caldovic, Ljubica; Morizono, Hiroki; Allewell, Norma M.; Tuchman, Mendel; Shi, Dashuang

    2009-01-01

    N-Acetylglutamate synthase (NAGS) catalyzes the first committed step in l-arginine biosynthesis in plants and micro-organisms and is subject to feedback inhibition by l-arginine. This study compares the crystal structures of NAGS from Neisseria gonorrhoeae (ngNAGS) in the inactive T-state with l-arginine bound and in the active R-state complexed with CoA and l-glutamate. Under all of the conditions examined, the enzyme consists of two stacked ...

  5. High Throughput Screen for Escherichia coli Twin Arginine Translocation (Tat) Inhibitors

    Science.gov (United States)

    Bageshwar, Umesh K.; VerPlank, Lynn; Baker, Dwight; Dong, Wen; Hamsanathan, Shruthi; Whitaker, Neal; Sacchettini, James C.; Musser, Siegfried M.

    2016-01-01

    The twin arginine translocation (Tat) pathway transports fully-folded and assembled proteins in bacteria, archaea and plant thylakoids. The Tat pathway contributes to the virulence of numerous bacterial pathogens that cause disease in humans, cattle and poultry. Thus, the Tat pathway has the potential to be a novel therapeutic target. Deciphering the Tat protein transport mechanism has been challenging since the active translocon only assembles transiently in the presence of substrate and a proton motive force. To identify inhibitors of Tat transport that could be used as biochemical tools and possibly as drug development leads, we developed a high throughput screen (HTS) to assay the effects of compounds in chemical libraries against protein export by the Escherichia coli Tat pathway. The primary screen is a live cell assay based on a fluorescent Tat substrate that becomes degraded in the cytoplasm when Tat transport is inhibited. Consequently, low fluorescence in the presence of a putative Tat inhibitor was scored as a hit. Two diverse chemical libraries were screened, yielding average Z'-factors of 0.74 and 0.44, and hit rates of ~0.5% and 0.04%, respectively. Hits were evaluated by a series of secondary screens. Electric field gradient (Δψ) measurements were particularly important since the bacterial Tat transport requires a Δψ. Seven low IC50 hits were eliminated by Δψ assays, suggesting ionophore activity. As Δψ collapse is generally toxic to animal cells and efficient membrane permeability is generally favored during the selection of library compounds, these results suggest that secondary screening of hits against electrochemical effects should be done early during hit validation. Though none of the short-listed compounds inhibited Tat transport directly, the screening and follow-up assays developed provide a roadmap to pursue Tat transport inhibitors. PMID:26901445

  6. Arginine Decarboxylase expression, polyamines biosynthesis and reactive oxygen species during organogenic nodule formation in hop.

    Science.gov (United States)

    Fortes, Ana M; Costa, Joana; Santos, Filipa; Seguí-Simarro, José M; Palme, Klaus; Altabella, Teresa; Tiburcio, Antonio F; Pais, Maria S

    2011-02-01

    Hop (Humulus lupulus L.) is an economically important plant species used in beer production and as a health-promoting medicine. Hop internodes develop upon stress treatments organogenic nodules which can be used for genetic transformation and micropropagation. Polyamines are involved in plant development and stress responses. Arginine decarboxylase (ADC; EC 4·1.1·19) is a key enzyme involved in the biosynthesis of putrescine in plants. Here we show that ADC protein was increasingly expressed at early stages of hop internode culture (12h). Protein continued accumulating until organogenic nodule formation after 28 days, decreasing thereafter. The same profile was observed for ADC transcript suggesting transcriptional regulation of ADC gene expression during morphogenesis. The highest transcript and protein levels observed after 28 days of culture were accompanied by a peak in putrescine levels. Reactive oxygen species accumulate in nodular tissues probably due to stress inherent to in vitro conditions and enhanced polyamine catabolism. Conjugated polyamines increased during plantlet regeneration from nodules suggesting their involvement in plantlet formation and/or in the control of free polyamine levels. Immunogold labeling revealed that ADC is located in plastids, nucleus and cytoplasm of nodular cells. In vacuolated cells, ADC immunolabelling in plastids doubled the signal of proplastids in meristematic cells. Location of ADC in different subcellular compartments may indicate its role in metabolic pathways taking place in these compartments. Altogether these data suggest that polyamines play an important role in organogenic nodule formation and represent a progress towards understanding the role played by these growth regulators in plant morphogenesis. PMID:21415599

  7. Mutation of Residue Arginine18 of Cytochrome b559 α-Subunit and its Effects on Photosystem Ⅱ Activities in Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    It has been known that arginine is used as the basic amino acid in the ?subunit of cytochrome b559 (Cyt b559) except histidine. However, previous studies have focused on the function of histidine in the activities of photosystem (PS) Ⅱ and there are no reports regarding the structural and/or functional roles of arginine in PSII complexes. In the present study,two arginine18 (R18) mutants of Chlamydomonas reinhardtii were constructed using site-directed mutagenesis, in which R18 was replaced by glutamic acid (E) and glycine (G). The results show that the oxygen evolution of the PSII complex in the R18G and R18E mutants was approximately 60% of wild-type (WT) levels and that, after irradiation at high light intensity, oxygen evolution for the PSII of mutants was reduced to zero compared with 40% in WT cells. The efficiency of light capture by PSII (Fv/Fm) of R18G and R18E mutants was approximately 42%-46% that of WT cells. Furthermore, levels of the ?subunit of Cyt b559 and PsbO proteins were reduced in thylakoid membranes compared with WT. Overall, these data suggest that R18 plays a significant role in helping Cyt b559 maintain the structure of the PSII complex and its activity,although it is not directly bound to the heme group.

  8. Transgenic approach to express the channelrhodopsin 2 gene in arginine vasopressin neurons of rats.

    Science.gov (United States)

    Ishii, Masahiro; Hashimoto, Hirofumi; Ohkubo, Jun-Ichi; Ohbuchi, Toyoaki; Saito, Takeshi; Maruyama, Takashi; Yoshimura, Mitsuhiro; Yamamoto, Yukiyo; Kusuhara, Koichi; Ueta, Yoichi

    2016-09-01

    Optogenetics provides a powerful tool to regulate neuronal activity by light-sensitive ion channels such as channelrhodopsin 2 (ChR2). Arginine vasopressin (AVP; also known as the anti-diuretic hormone) is a multifunctional hormone which is synthesized in the magnocellular neurosecretory cells (MNCs) of the hypothalamus. Here, we have generated a transgenic rat that expresses an AVP-ChR2-enhanced green fluorescent protein (eGFP) fusion gene in the MNCs of the hypothalamus. The eGFP fluorescence that indicates the expression of ChR2-eGFP was observed in the supraoptic nucleus (SON) and in the magnocellular division of the paraventricular nucleus (PVN) that is known to contain AVP-secreting neurons. The eGFP fluorescence intensities in those nuclei and posterior pituitary were markedly increased after chronic salt loading (2% NaCl in drinking water for 5days). ChR2-eGFP was localized mainly in the membrane of AVP-positive MNCs. Whole-cell patch-clamp recordings were performed from single MNCs isolated from the SON of the transgenic rats, and blue light evoked repetitive action potentials. Our work provides for the first time an optogenetic approach to selectively activate AVP neurons in the rat. PMID:27493075

  9. Continuous spectrophotometric assays for three regulatory enzymes of the arginine biosynthetic pathway.

    Science.gov (United States)

    Takahara, Kentaro; Akashi, Kinya; Yokota, Akiho

    2007-09-15

    N-Acetylglutamate synthase (AGS), N-acetylglutamate kinase (AGK), and glutamate N-acetyltransferase (GAT) are the key enzymes in the synthesis of arginine that serves as an important precursor for the synthesis of protein, polyamines, urea, and nitric oxide. Current assays available for these three enzymes are laborious and time-consuming and do not allow continuous monitoring of enzyme activities. Here we established continuous enzyme assays for AGS, AGK, and GAT based on the coupling of AGS and GAT reactions to AGK followed by coupling of the AGK reaction to N-acetylglutamate 5-phosphate reductase (AGPR). The rate of AGPR-dependent oxidation of reduced nicotinamide adenine dinucleotide phosphate was monitored continuously as a change in absorbance at 340 nm using spectrophotometry. These methods were applied to kinetic analyses for Escherichia coli AGK, E. coli AGS, and Saccharomyces cerevisiae GAT, and the kinetic parameters obtained in the coupling assays showed nearly the same values as those obtained previously using discontinuous assays. The specificity of these coupled assays was confirmed by the lack of enzyme activity from extracts of E. coli AGS-, E. coli AGK-, and S. cerevisiae GAT-deletion mutants. Moreover, the coupled assay enabled us to measure AGS activity from mammalian liver mitochondrial extracts, known to be an important regulatory enzyme for the urea cycle. These coupled enzyme assays are rapid, highly sensitive, and reproducible. PMID:17651682

  10. Substitution of arginine for histidine-47 in the coenzyme binding site of yeast alcohol dehydrogenase I

    International Nuclear Information System (INIS)

    Molecular modeling of alcohol dehydrogenases suggests that His-47 in the yeast enzyme (His-44 in the protein sequence, corresponding to Arg-47 in the horse liver enzyme) binds the pyrophosphate of the NAD coenzyme. His-47 in the Saccharomyces cerevisiae isoenzyme I was substituted with an arginine by a directed mutation. Steady-state kinetic results at pH 7.3 and 30 degree C of the mutant and wild-type enzymes were consistent with an ordered Bi-Bi mechanism. The substitution decreased dissociation constants by 4-fold for NAD+ and 2-fold for NADH while turnover numbers were decreased by 4-fold for ethanol oxidation and 6-fold for acetaldehyde reduction. The magnitudes of these effects are smaller than those found for the same mutation in the human liver β enzyme, suggesting that other amino acid residues in the active site modulate the effects of the substitution. The pH dependencies of dissociation constants and other kinetic constants were similar in the two yeast enzymes. Thus, it appears that His-47 is not solely responsible for a pK value near 7 that controls activity and coenzyme binding rates in the wild-type enzyme. The small substrate deuterium isotope effect above pH 7 and the single exponential phase of NADH production during the transient oxidation of ethanol by the Arg-47 enzyme suggest that the mutation makes an isomerization of the enzyme-NAD+ complex limiting for turnover with ethanol

  11. Preliminary crystallographic studies of yeast mitochondrial peripheral membrane protein Tim44p

    International Nuclear Information System (INIS)

    Tim44p is an essential mitochondrial peripheral membrane protein. To investigate the mechanism by which Tim44p functions in the TIM23 translocon to deliver the mitochondrial protein precursors, the yeast Tim44p has been crystallized. Protein translocations across mitochondrial membranes play critical roles in mitochondrion biogenesis. Protein transport from the cell cytosol to the mitochondrial matrix is carried out by the translocase of the outer membrane (TOM) complex and the translocase of the inner membrane (TIM) complexes. Tim44p is an essential mitochondrial peripheral membrane protein and a major component of the TIM23 translocon. To investigate the mechanism by which Tim44p functions in the TIM23 translocon to deliver the mitochondrial protein precursors, the yeast Tim44p was crystallized. The crystals diffract to 3.2 Å using a synchrotron X-ray source and belong to space group P6322, with unit-cell parameters a = 124.25, c = 77.83 Å. There is one Tim44p molecule in one asymmetric unit, which corresponds to a solvent content of approximately 43%. Structure determination by MAD methods is under way

  12. Preliminary crystallographic studies of yeast mitochondrial peripheral membrane protein Tim44p

    Energy Technology Data Exchange (ETDEWEB)

    Josyula, Ratnakar [Department of Cell Biology, Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States); Jin, Zhongmin [SER-CAT, APS, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439 (United States); McCombs, Deborah; DeLucas, Lawrence [Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States); Sha, Bingdong, E-mail: bdsha@uab.edu [Department of Cell Biology, Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States)

    2006-02-01

    Tim44p is an essential mitochondrial peripheral membrane protein. To investigate the mechanism by which Tim44p functions in the TIM23 translocon to deliver the mitochondrial protein precursors, the yeast Tim44p has been crystallized. Protein translocations across mitochondrial membranes play critical roles in mitochondrion biogenesis. Protein transport from the cell cytosol to the mitochondrial matrix is carried out by the translocase of the outer membrane (TOM) complex and the translocase of the inner membrane (TIM) complexes. Tim44p is an essential mitochondrial peripheral membrane protein and a major component of the TIM23 translocon. To investigate the mechanism by which Tim44p functions in the TIM23 translocon to deliver the mitochondrial protein precursors, the yeast Tim44p was crystallized. The crystals diffract to 3.2 Å using a synchrotron X-ray source and belong to space group P6{sub 3}22, with unit-cell parameters a = 124.25, c = 77.83 Å. There is one Tim44p molecule in one asymmetric unit, which corresponds to a solvent content of approximately 43%. Structure determination by MAD methods is under way.

  13. Functional characterization and expression analysis of rice δ1-pyrroline-5-carboxylate dehydrogenase provide new insight into the regulation of proline and arginine catabolism

    Science.gov (United States)

    Forlani, Giuseppe; Bertazzini, Michele; Zarattini, Marco; Funck, Dietmar

    2015-01-01

    While intracellular proline accumulation in response to various stress conditions has been investigated in great detail, the biochemistry and physiological relevance of proline degradation in plants is much less understood. Moreover, the second and last step in proline catabolism, the oxidation of δ1-pyrroline-5-carboxylic acid (P5C) to glutamate, is shared with arginine catabolism. Little information is available to date concerning the regulatory mechanisms coordinating these two pathways. Expression of the gene coding for P5C dehydrogenase was analyzed in rice by real-time PCR either following the exogenous supply of amino acids of the glutamate family, or under hyperosmotic stress conditions. The rice enzyme was heterologously expressed in E. coli, and the affinity-purified protein was thoroughly characterized with respect to structural and functional properties. A tetrameric oligomerization state was observed in size exclusion chromatography, which suggests a structure of the plant enzyme different from that shown for the bacterial P5C dehydrogenases structurally characterized to date. Kinetic analysis accounted for a preferential use of NAD+ as the electron acceptor. Cations were found to modulate enzyme activity, whereas anion effects were negligible. Several metal ions were inhibitory in the micromolar range. Interestingly, arginine also inhibited the enzyme at higher concentrations, with a mechanism of uncompetitive type with respect to P5C. This implies that millimolar levels of arginine would increase the affinity of P5C dehydrogenase toward its specific substrate. Results are discussed in view of the involvement of the enzyme in either proline or arginine catabolism. PMID:26300893

  14. Functional characterization and expression analysis of rice δ(1)-pyrroline-5-carboxylate dehydrogenase provide new insight into the regulation of proline and arginine catabolism.

    Science.gov (United States)

    Forlani, Giuseppe; Bertazzini, Michele; Zarattini, Marco; Funck, Dietmar

    2015-01-01

    While intracellular proline accumulation in response to various stress conditions has been investigated in great detail, the biochemistry and physiological relevance of proline degradation in plants is much less understood. Moreover, the second and last step in proline catabolism, the oxidation of δ(1)-pyrroline-5-carboxylic acid (P5C) to glutamate, is shared with arginine catabolism. Little information is available to date concerning the regulatory mechanisms coordinating these two pathways. Expression of the gene coding for P5C dehydrogenase was analyzed in rice by real-time PCR either following the exogenous supply of amino acids of the glutamate family, or under hyperosmotic stress conditions. The rice enzyme was heterologously expressed in E. coli, and the affinity-purified protein was thoroughly characterized with respect to structural and functional properties. A tetrameric oligomerization state was observed in size exclusion chromatography, which suggests a structure of the plant enzyme different from that shown for the bacterial P5C dehydrogenases structurally characterized to date. Kinetic analysis accounted for a preferential use of NAD(+) as the electron acceptor. Cations were found to modulate enzyme activity, whereas anion effects were negligible. Several metal ions were inhibitory in the micromolar range. Interestingly, arginine also inhibited the enzyme at higher concentrations, with a mechanism of uncompetitive type with respect to P5C. This implies that millimolar levels of arginine would increase the affinity of P5C dehydrogenase toward its specific substrate. Results are discussed in view of the involvement of the enzyme in either proline or arginine catabolism. PMID:26300893

  15. L-canavanine and protein synthesis in the tobacco hornworm Manduca sexta

    Energy Technology Data Exchange (ETDEWEB)

    Rosenthal, G.A.; Dahlman, D.L.

    1986-01-01

    L-Canavanine, a nonprotein amino acid of certain leguminous plants, manifests potent insecticidal properties in a canavanine-sensitive insect such as the tobacco hornworm Manduca sexta (L.) (Sphingidae). This arginine analog is activated and aminoacylated by arginyl-tRNA synthetase and incorporated into nascent polypeptide chains to create structurally aberrant, canavanine-containing proteins. Analysis of incorporation of (TH)leucine into protein in M. sexta larvae that had been injected with canavanine revealed that this arginine analog stimulates protein synthesis. During the first 3 hr after injection of canavanine, canavanine-mediated net stimulation of protein formation was readily discerned. Thereafter, the stimulation of protein synthesis appeared to be offset by the preferential of degradation of anomalous proteins. Double-label protein-turnover experiments with larvae injected with ( UC)canavanine- and (TH)arginine-containing hemolymph proteins showed that canavanine-containing proteins were degraded preferentially.

  16. L-canavanine and protein synthesis in the tobacco hornworm Manduca sexta

    International Nuclear Information System (INIS)

    L-Canavanine, a nonprotein amino acid of certain leguminous plants, manifests potent insecticidal properties in a canavanine-sensitive insect such as the tobacco hornworm Manduca sexta (L.) (Sphingidae). This arginine analog is activated and aminoacylated by arginyl-tRNA synthetase and incorporated into nascent polypeptide chains to create structurally aberrant, canavanine-containing proteins. Analysis of incorporation of [3H]leucine into protein in M. sexta larvae that had been injected with canavanine revealed that this arginine analog stimulates protein synthesis. During the first 3 hr after injection of canavanine, canavanine-mediated net stimulation of protein formation was readily discerned. Thereafter, the stimulation of protein synthesis appeared to be offset by the preferential of degradation of anomalous proteins. Double-label protein-turnover experiments with larvae injected with [14C]canavanine- and [3H]arginine-containing hemolymph proteins showed that canavanine-containing proteins were degraded preferentially

  17. Synthesis, characterization and properties of L-arginine-passivated silver nanocolloids

    Science.gov (United States)

    Sunatkari, A. L.; Talwatkar, S. S.; Tamgadge, Y. S.; Muley, G. G.

    2016-05-01

    We investigate the effect of L-arginine-surface passivation on localised surface plasmon resonance (LSPR), size and stability of colloidal Silver Nanoparticles (AgNPs) synthesized by chemical reduction method. The surface Plasmon resonance absorption peak of AgNPs shows blue shift with the increase in L-arginine concentration. Transmission electron microscopy (TEM) analysis confirmed that the average size of AgNPs reduces from 10 nm to 6 nm as the concentration of L-Arginine increased from 1 to 5 mM. The X-ray diffraction study (XRD) confirmed the formation face-centred cubic (fcc) structured AgNPs. FT-IR studies revealed strong bonding between L-arginine functional groups and AgNPs.

  18. Streptococcus pneumoniae arginine synthesis genes promote growth and virulence in pneumococcal meningitis

    NARCIS (Netherlands)

    J.R. Piet; M. Geldhoff; B.D.C. van Schaik; M.C. Brouwer; M. Valls Seron; M.E. Jakobs; K. Schipper; Y. Pannekoek; A.H. Zwinderman; T. van der Poll; A.H.C. van Kampen; F. Baas; A van der Ende; D. van de Beek

    2014-01-01

    Streptococcus pneumoniae (pneumococcus) is a major human pathogen causing pneumonia, sepsis and bacterial meningitis. Using a clinical phenotype based approach with bacterial whole-genome sequencing we identified pneumococcal arginine biosynthesis genes to be associated with outcome in patients with

  19. Thermal, FT–IR and SHG efficiency studies of L-arginine doped KDP crystals

    Indian Academy of Sciences (India)

    K D Parikh; D J Dave; B B Parekh; M J Joshi

    2007-04-01

    Potassium dihydrogen phosphate (KDP) is a well known nonlinear optical (NLO) material with different applications. Since most of the amino acids exhibit NLO property, it is of interest to dope them in KDP. In the present study, amino acid L-arginine was doped in KDP. The doping of L-arginine was confirmed by FT–IR and paper chromatography. Thermogravimetry suggested that as the amount of doping increases the thermal stability decreases as well as the value of thermodynamic and kinetic parameters decreases. The second harmonic generation (SHG) efficiency of L-arginine doped KDP crystals was found to be increasing with doping concentration of L-arginine. The results are discussed here.

  20. Resveratrol inhibits Trypanosoma cruzi arginine kinase and exerts a trypanocidal activity.

    Science.gov (United States)

    Valera Vera, Edward A; Sayé, Melisa; Reigada, Chantal; Damasceno, Flávia S; Silber, Ariel M; Miranda, Mariana R; Pereira, Claudio A

    2016-06-01

    Arginine kinase catalyzes the reversible transphosphorylation between ADP and phosphoarginine which plays a critical role in the maintenance of cellular energy homeostasis. Arginine kinase from the protozoan parasite Trypanosoma cruzi, the etiologic agent of Chagas disease, meets the requirements to be considered as a potential therapeutic target for rational drug design including being absent in its mammalian hosts. In this study a group of polyphenolic compounds was evaluated as potential inhibitors of arginine kinase using molecular docking techniques. Among the analyzed compounds with the lowest free binding energy to the arginine kinase active site (market price; and (3) has as a well-defined target enzyme which is absent in the mammalian host, it is a promising compound as a trypanocidal drug for Chagas disease. PMID:26976067

  1. Inhibition of corrosion of carbon steel in well water by arginine-Zn2+ system

    Directory of Open Access Journals (Sweden)

    ANTHONY SAMY SAHAYA RAJA

    2012-06-01

    Full Text Available The environmental friendly inhibitor system arginine-Zn2+, has been investigated by weight-loss method. A synergistic effect exists between arginine and Zn2+ system. The formulation consisting of 250 ppm of arginine and 5 ppm of Zn2+ offers good inhibition efficiency of 98 %. Polarization study reveals that this formulation functions as an anodic inhibitor. AC impedance spectra reveal that a protective film is formed on the metal sur­face. The FTIR spectral study leads to the conclusion that the Fe2+- DL-arginine complex, formed on anodic sites of the metal surface, controls the anodic reaction. Zn(OH2 formed on the cathodic sites of the metal surface controls the cathodic reaction. The surface morphology and the roughness of the metal surface were analyzed with Atomic Force Microscope. A suitable mechanism of corrosion inhibition is proposed based on the results obtained from weight loss study and surface analysis technique.

  2. Metabolomic analysis of plasma and liver from surplus arginine fed Atlantic salmon

    Science.gov (United States)

    Andersen, Synne M.; Assaad, Houssein I.; Lin, Gang; Wang, Junjun; Aksnes, Anders; Wu, Guoyao; Espe, Marit

    2016-01-01

    The aim of this study was to determine the metabolic effect of surplus arginine (36.1 g/kg dry matter) compared to a control diet with required arginine (21.1 g/kg dry matter) in adult Atlantic salmon (Salmo salar L.). Although the feeding trial had no significant effect on growth, there were significant differences in the metabolite profile in both plasma and liver in experimental group as compared to the control group. There was increased concentrations of biliverdin, PGF-2 alpha, oxidized glutathione, selenocysteine, two monoacylglycerols and a tripeptide in the liver as well as decreased concentrations of valine and a vitamin D3 metabolite in plasma of arginine supplemented fish. These results indicate that while surplus arginine does not affect growth or body weight, it induces metabolic changes in Atlantic salmon. PMID:25553364

  3. Arginine does not exacerbate markers of inflammation in cocultures of human enterocytes and leukocytes

    DEFF Research Database (Denmark)

    Parlesak, Alexandr; Negrier, I.; Neveux, N.;

    2007-01-01

    Enteral arginine supplementation in the critically ill has become a matter of controversy. In this study, we investigated effects of the addition of 0.4 and 1.2 mmol/L arginine in a coculture model on markers of inflammation, enterocyte layer integrity, and amino acid transport. In this model, a...... transepithelial flux of 22 amino acids, their catabolism, and the integrity of the enterocyte layer assessed as permeability of fluorescein dextran (M(r) 4400). Bacterial stimulation of intestinal epithelial cells enhanced the basolateral concentration of nitric oxide and all cytokines measured. Supplementation...... the catabolism of serine, asparagine, and lysine, and reduced glutamine catabolism. Addition of arginine increased ornithine formation and moderately reduced transepithelial transport of methionine and other amino acids. Hence, arginine supplementation does not interfere with inflammation...

  4. Arginine metabolism by macrophages promotes cardiac and muscle fibrosis in mdx muscular dystrophy.

    Directory of Open Access Journals (Sweden)

    Michelle Wehling-Henricks

    Full Text Available BACKGROUND: Duchenne muscular dystrophy (DMD is the most common, lethal disease of childhood. One of 3500 new-born males suffers from this universally-lethal disease. Other than the use of corticosteroids, little is available to affect the relentless progress of the disease, leading many families to use dietary supplements in hopes of reducing the progression or severity of muscle wasting. Arginine is commonly used as a dietary supplement and its use has been reported to have beneficial effects following short-term administration to mdx mice, a genetic model of DMD. However, the long-term effects of arginine supplementation are unknown. This lack of knowledge about the long-term effects of increased arginine metabolism is important because elevated arginine metabolism can increase tissue fibrosis, and increased fibrosis of skeletal muscles and the heart is an important and potentially life-threatening feature of DMD. METHODOLOGY: We use both genetic and nutritional manipulations to test whether changes in arginase metabolism promote fibrosis and increase pathology in mdx mice. Our findings show that fibrotic lesions in mdx muscle are enriched with arginase-2-expressing macrophages and that muscle macrophages stimulated with cytokines that activate the M2 phenotype show elevated arginase activity and expression. We generated a line of arginase-2-null mutant mdx mice and found that the mutation reduced fibrosis in muscles of 18-month-old mdx mice, and reduced kyphosis that is attributable to muscle fibrosis. We also observed that dietary supplementation with arginine for 17-months increased mdx muscle fibrosis. In contrast, arginine-2 mutation did not reduce cardiac fibrosis or affect cardiac function assessed by echocardiography, although 17-months of dietary supplementation with arginine increased cardiac fibrosis. Long-term arginine treatments did not decrease matrix metalloproteinase-2 or -9 or increase the expression of utrophin, which have

  5. Tailored second line therapy in asthmatic children with the arginine-16 genotype

    OpenAIRE

    Lipworth, Brian J; Basu, Kaninika; Donald, Helen P; Tavendale, Roger; Macgregor, Donald F; Ogston, Simon A; Palmer, Colin N. A.; Mukhopadhyay, Somnath

    2012-01-01

    The arginine-16 beta-2 receptor genotype confers increased susceptibility to exacerbations in asthmatic children taking regular long acting beta-2 agonists. We therefore evaluated using montelukast as an alternative to salmeterol as tailored second line asthma controller therapy in children expressing this susceptible genotype. 62 persistent asthmatic children with the homozygous arginine-16 genotype were randomized to receive salmeterol 50ug bid or montelukast 5/10mg od as add on to inhaled ...

  6. Arginine and pyrimidine biosynthetic defects in Neisseria gonorrhoeae strains isolated from patients.

    OpenAIRE

    Shinners, E N; Catlin, B W

    1982-01-01

    Neisseria gonorrhoeae strains with nutritional requirements that include arginine (Arg-), uracil (Ura-), and hypoxanthine have attracted attention because of their tendency to cause disseminated infections, as a basis for genetic studies of arginine and pyrimidine biosynthesis, we examined the activities of four enzymes of these pathways in cell-free extracts of both prototrophic and Arg- Ura- strains. Activities of glutamate acetyltransferase, aspartate transcarbamylase, and orotate phosphor...

  7. Chiral pharmacokinetics and inversion of NG-nitro-arginine in the rat

    Institute of Scientific and Technical Information of China (English)

    Yan-feiXIN; RuiTONG; YangFANG; Xiang-junZHOU; Yong-xiangWANG

    2004-01-01

    AIM: To explore pharmacokinetics of NG-nitro-D-arginine (D-NNA) and NG-nitro-L-arginine (L-NNA) in conscious rats.METHODS: The plasma concentration of D-NNA and L-NNA were determined by chiral ligand exchange method with capillary electrochromatography (CEC). Pharmacokinetic parameters were estimated using non-compartment model and were fitted using a computer program DAS. Chiral inversion rate of D-NNA to L-

  8. Arginine intake and risk of coronary heart disease mortality in elderly men

    OpenAIRE

    Oomen, C.M.; Erk, van, M.J.; Feskens, E.J.M.; Kok, F.J.; Kromhout, D.

    2000-01-01

    From experimental studies, the hypothesis is derived that the amino acid arginine, the precursor of NO, could restore the impaired endothelial function and increased platelet activation observed in atherosclerosis. We investigated whether dietary intake of arginine is associated with reduced coronary heart disease risk in elderly persons. The study population consisted of 806 men aged 64 to 84 years at baseline who participated in the Zutphen Elderly Study, a population-based cohort followed ...

  9. L-Arginine but not L-glutamine likely increases exogenous carbohydrate oxidation during endurance exercise.

    Science.gov (United States)

    Rowlands, David S; Clarke, Jim; Green, Jackson G; Shi, Xiaocai

    2012-07-01

    The addition of L-arginine or L-glutamine to glucose-electrolyte solutions can increase intestinal water, glucose, and sodium absorption in rats and humans. We evaluated the utility of L-arginine and L-glutamine in energy-rehydration beverages through assessment of exogenous glucose oxidation and perceptions of exertion and gastrointestinal distress during endurance exercise. Eight cyclists rode 150 min at 50% of peak power on four occasions while ingesting solutions at a rate of 150 mL 15 min(-1) that contained (13)C-enriched glucose (266 mmol L(-1)) and sodium citrate ([Na(+)] 60 mmol L(-1)), and either: 4.25 mmol L(-1) L-arginine or 45 mmol L(-1) L-glutamine, and as controls glucose only or no glucose. Relative to glucose only, L-arginine invoked a likely 12% increase in exogenous glucose oxidation (90% confidence limits: ± 8%); however, the effect of L-glutamine was possibly trivial (4.5 ± 7.3%). L-Arginine also led to very likely small reductions in endogenous fat oxidation rate relative to glucose (12 ± 4%) and L-glutamine (14 ± 4%), and relative to no glucose, likely reductions in exercise oxygen consumption (2.6 ± 1.5%) and plasma lactate concentration (0.20 ± 0.16 mmol L(-1)). Effects on endogenous and total carbohydrate oxidation were inconsequential. Compared with glucose only, L-arginine and L-glutamine caused likely small-moderate effect size increases in perceptions of stomach fullness, abdominal cramp, exertion, and muscle tiredness during exercise. Addition of L-arginine to a glucose and electrolyte solution increases the oxidation of exogenous glucose and decreases the oxygen cost of exercise, although the mechanisms responsible and impact on endurance performance require further investigation. However, L-arginine also increases subjective feelings of gastrointestinal distress, which may attenuate its other benefits. PMID:22048324

  10. PRE-EXERCISE ARGININE SUPPLEMENTATION INCREASES TIME TO EXHAUSTION IN ELITE MALE WRESTLERS

    OpenAIRE

    2014-01-01

    Dietary supplements containing arginine are among the most popular ergogenics intended to enhance strength, power and muscle recovery associated with both anaerobic and aerobic exercise. The aim of the present study was to evaluate the possible effect of pre-exercise acute intake of arginine on performance and exercise metabolism during incremental exhaustive exercise in elite male wrestlers. Nine volunteer elite male wrestlers (24.7±3.8 years) participated in this study. The test-retest prot...

  11. Nonspecific blockade of vascular free radical signals by methylated arginine analogues

    Directory of Open Access Journals (Sweden)

    Pedro M.A.

    1998-01-01

    Full Text Available Methylated arginine analogues are often used as probes of the effect of nitric oxide; however, their specificity is unclear and seems to be frequently overestimated. This study analyzed the effects of NG-methyl-L-arginine (L-NMMA on the endothelium-dependent release of vascular superoxide radicals triggered by increased flow. Plasma ascorbyl radical signals measured by direct electron paramagnetic resonance spectroscopy in 25 rabbits increased by 3.8 ± 0.7 nmol/l vs baseline (28.7 ± 1.4 nmol/l, P<0.001 in response to papaverine-induced flow increases of 121 ± 12%. In contrast, after similar papaverine-induced flow increases simultaneously with L-NMMA infusions, ascorbyl levels were not significantly changed compared to baseline. Similar results were obtained in isolated rabbit aortas perfused ex vivo with the spin trap a-phenyl-N-tert-butylnitrone (N = 22. However, in both preparations, this complete blockade was not reversed by co-infusion of excess L-arginine and was also obtained by N-methyl-D-arginine, thus indicating that it is not related to nitric oxide synthase. L-arginine alone was ineffective, as previously demonstrated for NG-methyl-L-arginine ester (L-NAME. In vitro, neither L-arginine nor its analogues scavenged superoxide radicals. This nonspecific activity of methylated arginine analogues underscores the need for careful controls in order to assess nitric oxide effects, particularly those related to interactions with active oxygen species.

  12. Abnormal mitochondrial L-arginine transport contributes to the pathogenesis of heart failure and rexoygenation injury.

    Directory of Open Access Journals (Sweden)

    David Williams

    Full Text Available BACKGROUND: Impaired mitochondrial function is fundamental feature of heart failure (HF and myocardial ischemia. In addition to the effects of heightened oxidative stress, altered nitric oxide (NO metabolism, generated by a mitochondrial NO synthase, has also been proposed to impact upon mitochondrial function. However, the mechanism responsible for arginine transport into mitochondria and the effect of HF on such a process is unknown. We therefore aimed to characterize mitochondrial L-arginine transport and to investigate the hypothesis that impaired mitochondrial L-arginine transport plays a key role in the pathogenesis of heart failure and myocardial injury. METHODS AND RESULTS: In mitochondria isolated from failing hearts (sheep rapid pacing model and mouse Mst1 transgenic model we demonstrated a marked reduction in L-arginine uptake (p<0.05 and p<0.01 respectively and expression of the principal L-arginine transporter, CAT-1 (p<0.001, p<0.01 compared to controls. This was accompanied by significantly lower NO production and higher 3-nitrotyrosine levels (both p<0.05. The role of mitochondrial L-arginine transport in modulating cardiac stress responses was examined in cardiomyocytes with mitochondrial specific overexpression of CAT-1 (mtCAT1 exposed to hypoxia-reoxygenation stress. mtCAT1 cardiomyocytes had significantly improved mitochondrial membrane potential, respiration and ATP turnover together with significantly decreased reactive oxygen species production and cell death following mitochondrial stress. CONCLUSION: These data provide new insights into the role of L-arginine transport in mitochondrial biology and cardiovascular disease. Augmentation of mitochondrial L-arginine availability may be a novel therapeutic strategy for myocardial disorders involving mitochondrial stress such as heart failure and reperfusion injury.

  13. Protein methylation is required to maintain optimal HIV-1 infectivity

    OpenAIRE

    Piller Sabine C; Warrilow David; Apolloni Ann; Bodetti Tracey J; Hitchen Eleanor M; Willemsen Nicole M; Harrich David

    2006-01-01

    Abstract Background: Protein methylation is recognized as a major protein modification pathway regulating diverse cellular events such as protein trafficking, transcription, and signal transduction. More recently, protein arginine methyltransferase activity has been shown to regulate HIV-1 transcription via Tat. In this study, adenosine periodate (AdOx) was used to globally inhibit protein methyltransferase activity so that the effect of protein methylation on HIV-1 infectivity could be asses...

  14. Cysteine and arginine-rich peptides as molecular carriers.

    Science.gov (United States)

    Shirazi, Amir Nasrolahi; El-Sayed, Naglaa Salem; Mandal, Dindayal; Tiwari, Rakesh K; Tavakoli, Kathy; Etesham, Matthew; Parang, Keykavous

    2016-01-15

    A number of linear and cyclic peptides containing alternative arginine and cysteine residues, namely linear (CR)3, linear (CR)4, linear (CR)5, cyclic [CR]4, and cyclic [CR]5, were synthesized. The peptides were evaluated for their ability to deliver two molecular cargos, fluorescence-labeled cell-impermeable negatively charged phosphopeptide (F'-GpYEEI) and fluorescence-labeled lamivudine (F'-3TC), intracellularly in human leukemia cancer (CCRF-CEM) cells. We investigated the role of cyclization and the number of amino acids in improving the transporting ability of the peptides. The flow cytometry studies suggested that the synthesized peptides were able to work efficiently as transporters for both cargos. Among all compounds, cyclic [CR]4 was found to be the most efficient peptide in transporting the cargo into cells. For instance, the cellular uptake of F'-3TC (5μM) and F'-GpYEEI (5μM) was enhanced by 16- and 20-fold, respectively, in the presence of cyclic [CR]4 compared to that of the parent compound alone. The mechanism of F'-GpYEEI uptake by cells was found to be energy-independent. The results showed that the number of amino acids and their cyclic nature can impact the efficiency of the peptide in transporting the molecular cargos. PMID:26631317

  15. Urinary and metabolic clearances of arginine vasopressin in normal subjects

    International Nuclear Information System (INIS)

    Synthetic arginine vasopressin (AVP) was infused into 11 hydrated normal subjects at five different infusion rates ranging from 10 to 350 μU kg-1 min-1. Each infusion rate was continued for 1 h, and urinary determinations were made on the 30- to 60-min specimens during which time there was no further rise in plasma AVP. Urinary AVP concentrations (μU/ml) and excretion rates (μU/min) increased linearly with increasing infusion rates, and the concentration of AVP in urine increased 120 times more rapid than plasma. Urinary and metabolic clearances of AVP also increased linearly with the maximum urinary clearance being 60.6% of the creatinine clearance. The total metabolic clearance of AVP (including urinary clearance) was 17.8 times that of the urinary clearance of AVP alone. These data clarify the relationships between plasma and urinary AVP in normal hydrated subjects during AVP infusion under steady-state conditions and emphasize the potential advantage of measuring urinary AVP as a monitor of endogenous AVP secretion. AVP was measured by radioimmunoassay

  16. tlpA gene expression is required for arginine and bicarbonate chemotaxis in Helicobacter pylori

    Directory of Open Access Journals (Sweden)

    Oscar A Cerda

    2011-01-01

    Full Text Available About half of the human population is infected with Helicobacter pylori, a bacterium causing gastritis, peptic ulcer and progression to gastric cancer. Chemotaxis and flagellar motility are required for colonization and persistence of H. pylori in the gastric mucus layer. It is not completely clear which chemical gradients are used by H. pylori to maintain its position. TlpA, a chemotaxis receptor for arginine/ bicarbonate, has been identified. This study aimed to find out whether tlpA gene expression is required for the chemotactic response to arginine/bicarbonate. Wild-type motile H. pylori ATCC 700392 and H. pylori ATCC 43504, a strain having an interrupted tlpA gene, were used. Also, a tlpA-knockout mutant of H. pylori 700392 (H. pylori 700-tlpA::cat was produced by homologous recombination. Expression of tlpA was assessed by a Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR assay. Chemotaxis was measured as a Relative Chemotaxis Response (RCR by a modified capillary assay. H. pylori 700392 presented chemotaxis to arginine and sodium bicarbonate. H. pylori 700-tlpA::cat showed neither tlpA gene expression nor chemotaxis towards arginine and bicarbonate. Besides confirming that TlpA is a chemotactic receptor for arginine/bicarbonate in H. pylori, this study showed that tlpA gene expression is required for arginine/bicarbonate chemotaxis.

  17. AcEST: DK958066 [AcEST

    Lifescience Database Archive (English)

    Full Text Available translocase prote... 106 1e-21 tr|B4WJ18|B4WJ18_9SYNE Twin arginine-targeting protein trans...dependent protein translocase TatC ... 102 2e-20 tr|B4VMB2|B4VMB2_9CYAN Twin arginine-targeting protein trans... OS=Physcomitrella paten... 147 5e-34 tr|Q9AVE6|Q9AVE6_ORYSJ cDNA clone:J013104L23, full insert sequen...TTLKVSGYCGLLLG 97 >tr|Q9AVE6|Q9AVE6_ORYSJ cDNA clone:J013104L23, full insert sequence OS=Oryza sativa subsp....mology search results ■■ - Swiss-Prot (release 56.9) Link to BlastX Result : Swiss-Prot sp_hit_id P54086 Defin

  18. N-terminal arginines modulate plasma-membrane localization of Kv7.1/KCNE1 channel complexes.

    Directory of Open Access Journals (Sweden)

    Zenawit Girmatsion

    Full Text Available BACKGROUND AND OBJECTIVE: The slow delayed rectifier current (I(Ks is important for cardiac action potential termination. The underlying channel is composed of Kv7.1 α-subunits and KCNE1 β-subunits. While most evidence suggests a role of KCNE1 transmembrane domain and C-terminus for the interaction, the N-terminal KCNE1 polymorphism 38G is associated with reduced I(Ks and atrial fibrillation (a human arrhythmia. Structure-function relationship of the KCNE1 N-terminus for I(Ks modulation is poorly understood and was subject of this study. METHODS: We studied N-terminal KCNE1 constructs disrupting structurally important positively charged amino-acids (arginines at positions 32, 33, 36 as well as KCNE1 constructs that modify position 38 including an N-terminal truncation mutation. Experimental procedures included molecular cloning, patch-clamp recording, protein biochemistry, real-time-PCR and confocal microscopy. RESULTS: All KCNE1 constructs physically interacted with Kv7.1. I(Ks resulting from co-expression of Kv7.1 with non-atrial fibrillation '38S' was greater than with any other construct. Ionic currents resulting from co-transfection of a KCNE1 mutant with arginine substitutions ('38G-3xA' were comparable to currents evoked from cells transfected with an N-terminally truncated KCNE1-construct ('Δ1-38'. Western-blots from plasma-membrane preparations and confocal images consistently showed a greater amount of Kv7.1 protein at the plasma-membrane in cells co-transfected with the non-atrial fibrillation KCNE1-38S than with any other construct. CONCLUSIONS: The results of our study indicate that N-terminal arginines in positions 32, 33, 36 of KCNE1 are important for reconstitution of I(Ks. Furthermore, our results hint towards a role of these N-terminal amino-acids in membrane representation of the delayed rectifier channel complex.

  19. Surprising arginine biosynthesis: a reappraisal of the enzymology and evolution of the pathway in microorganisms.

    Science.gov (United States)

    Xu, Ying; Labedan, Bernard; Glansdorff, Nicolas

    2007-03-01

    Major aspects of the pathway of de novo arginine biosynthesis via acetylated intermediates in microorganisms must be revised in light of recent enzymatic and genomic investigations. The enzyme N-acetylglutamate synthase (NAGS), which used to be considered responsible for the first committed step of the pathway, is present in a limited number of bacterial phyla only and is absent from Archaea. In many Bacteria, shorter proteins related to the Gcn5-related N-acetyltransferase family appear to acetylate l-glutamate; some are clearly similar to the C-terminal, acetyl-coenzyme A (CoA) binding domain of classical NAGS, while others are more distantly related. Short NAGSs can be single gene products, as in Mycobacterium spp. and Thermus spp., or fused to the enzyme catalyzing the last step of the pathway (argininosuccinase), as in members of the Alteromonas-Vibrio group. How these proteins bind glutamate remains to be determined. In some Bacteria, a bifunctional ornithine acetyltransferase (i.e., using both acetylornithine and acetyl-CoA as donors of the acetyl group) accounts for glutamate acetylation. In many Archaea, the enzyme responsible for glutamate acetylation remains elusive, but possible connections with a novel lysine biosynthetic pathway arose recently from genomic investigations. In some Proteobacteria (notably Xanthomonadaceae) and Bacteroidetes, the carbamoylation step of the pathway appears to involve N-acetylornithine or N-succinylornithine rather than ornithine. The product N-acetylcitrulline is deacetylated by an enzyme that is also involved in the provision of ornithine from acetylornithine; this is an important metabolic function, as ornithine itself can become essential as a source of other metabolites. This review insists on the biochemical and evolutionary implications of these findings. PMID:17347518

  20. Protective effect of Mimosa pudica L. in an L-arginine model of acute necrotising pancreatitis in rats.

    Science.gov (United States)

    Kaur, Jagdeep; Sidhu, Shabir; Chopra, Kanwaljit; Khan, M U

    2016-07-01

    Mimosa pudica is used in traditional medicine for treating various disorders such as inflammatory conditions, diarrhoea, insomnia, alopecia, urogenital infections and wounds. The present study investigated the effect of M. pudica extract (MPE) on L-arginine-induced acute necrotising pancreatitis in rats. The ethanolic extract of M. pudica leaves was studied for the presence of quercetin and gallic acid using high-performance liquid chromatography. Four groups were employed-normal control rats, L-arginine control rats (two intraperitoneal [i.p.] injections of 2 g/kg at an interval of 1 h), MPE-treated rats (400 mg/kg orally) and melatonin-treated rats (positive control 10 mg/kg i.p.), which were further divided into subgroups according to time points (24 h, 3 days and 14 days). Serum amylase, lipase, tumour necrosis factor-α (TNF-α), pancreatic amylase, nucleic acid content, protein, transforming growth factor-β1 (TGF-β1), thiobarbituric reactive substances, glutathione, nitrite/nitrate, collagen content and histopathological examination were carried out. MPE significantly improved acute necrotising pancreatitis by modulating diagnostic markers of pancreatitis such as serum lipase and pancreatic amylase, inflammation (TNF-α), and oxidative and nitrosative stress. Moreover, MPE administration induced regenerative changes in the pancreas evidenced by increased levels of pancreatic proteins, nucleic acid content and histopathology report. In addition, MPE improved TGF-β1 and collagen levels thereby preventing fibrosis. The current investigation indicates the novel role of MPE in reducing the severity of acute necrotising pancreatitis by plausible mechanisms such as anti-inflammatory and anti-fibrotic activity and by promoting repair and regeneration of the pancreas. PMID:27164910

  1. Gastropod arginine kinases from Cellana grata and Aplysia kurodai. Isolation and cDNA-derived amino acid sequences.

    Science.gov (United States)

    Suzuki, T; Inoue, N; Higashi, T; Mizobuchi, R; Sugimura, N; Yokouchi, K; Furukohri, T

    2000-12-01

    Arginine kinase (AK) was isolated from the radular muscle of the gastropod molluscs Cellana grata (subclass Prosobranchia) and Aplysia kurodai (subclass Opisthobranchia), respectively, by ammonium sulfate fractionation, Sephadex G-75 gel filtration and DEAE-ion exchange chromatography. The denatured relative molecular mass values were estimated to be 40 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isolated enzyme from Aplysia gave a Km value of 0.6 mM for arginine and a Vmax value of 13 micromole Pi min(-1) mg protein(-1) for the forward reaction. These values are comparable to other molluscan AKs. The cDNAs encoding Cellana and Aplysia AKs were amplified by polymerase chain reaction, and the nucleotide sequences of 1,608 and 1,239 bp, respectively, were determined. The open reading frame for Cellana AK is 1044 nucleotides in length and encodes a protein with 347 amino acid residues, and that for A. kurodai is 1077 nucleotides and 354 residues. The cDNA-derived amino acid sequences were validated by chemical sequencing of internal lysyl endopeptidase peptides. The amino acid sequences of Cellana and Aplysia AKs showed the highest percent identity (66-73%) with those of the abalone Nordotis and turbanshell Battilus belonging to the same class Gastropoda. These AK sequences still have a strong homology (63-71%) with that of the chiton Liolophura (class Polyplacophora), which is believed to be one of the most primitive molluscs. On the other hand, these AK sequences are less homologous (55-57%) with that of the clam Pseudocardium (class Bivalvia), suggesting that the biological position of the class Polyplacophora should be reconsidered. PMID:11281267

  2. 25 years on and no end in sight: a perspective on the role of RecG protein.

    OpenAIRE

    Lloyd, RG; Rudolph, CJ

    2016-01-01

    The RecG protein of Escherichia coli is a double-stranded DNA translocase that unwinds a variety of branched substrates in vitro. Although initially associated with homologous recombination and DNA repair, studies of cells lacking RecG over the past 25 years have led to the suggestion that the protein might be multi-functional and associated with a number of additional cellular processes, including initiation of origin-independent DNA replication, the rescue of stalled or damaged replication ...

  3. Copper-induced tight junction mRNA expression changes, apoptosis and antioxidant responses via NF-κB, TOR and Nrf2 signaling molecules in the gills of fish: Preventive role of arginine

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Biao [Animal Nutrition Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); Feng, Lin; Jiang, Wei-Dan [Animal Nutrition Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); Fish Nutrition and Safety Production University Key Laboratory of Sichuan Province, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); Key Laboratory for Animal Disease-Resistance Nutrition of China Ministry of Education, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); Wu, Pei [Animal Nutrition Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); Kuang, Sheng-Yao [Animal Nutrition Institute, Sichuan Academy of Animal Science, Chengdu, 610066, Sichuan (China); Jiang, Jun [Animal Nutrition Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); Fish Nutrition and Safety Production University Key Laboratory of Sichuan Province, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); Key Laboratory for Animal Disease-Resistance Nutrition of China Ministry of Education, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); Tang, Ling; Tang, Wu-Neng [Animal Nutrition Institute, Sichuan Academy of Animal Science, Chengdu, 610066, Sichuan (China); Zhang, Yong-An [Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072 (China); Liu, Yang, E-mail: kyckgk@hotmail.com [Animal Nutrition Institute, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); Fish Nutrition and Safety Production University Key Laboratory of Sichuan Province, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); Key Laboratory for Animal Disease-Resistance Nutrition of China Ministry of Education, Sichuan Agricultural University, Chengdu 611130, Sichuan (China); and others

    2015-01-15

    Highlights: • Cu exposure induced oxidative stress via disruption of antioxidant system. • Cu exposure disrupted TJ mRNA expression through regulation of cytokines in fish. • Cu induced gill apoptosis partly via intrinsic pathway but not extrinsic pathway. • Cu exposure can regulate Nrf2, NF-κB and TOR signaling molecules in fish. • Arginine can effectively prevent Cu-induced fish gill damage. - Abstract: This study explored the possible preventive effects of dietary arginine on copper (Cu)-induced tight junction mRNA expression changes, apoptosis and antioxidant responses in the gills of young grass carp (Ctenopharyngodon idella). The results indicated that exposure to 0.7 mg/L (11.01 μmol/L) Cu for 96 h induced the production of reactive oxygen species (ROS), thereby increasing protein oxidation, lipid peroxidation and DNA damage in the gills of fish. However, these oxidative effects were prevented by arginine supplementation. Arginine also prevented the toxic effects of Cu on the activities of copper/zinc superoxide dismutase (SOD1), glutathione-S-transferase (GST), glutathione peroxidase (GPx), glutathione reductase (GR) and the glutathione (GSH) content (P < 0.05). However, Cu induced an adaptive increase in the activity of catalase (CAT), and arginine supplementation further increased CAT activity (P < 0.05). Moreover, Cu induced increases in the relative mRNA expressions of SOD1, CAT, GPx, GST, caspase-3, caspase-9, NF-E2-related factor 2 (Nrf2), Kelch-like-ECH-associated protein 1a (Keap1a), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-8 (IL-8), transforming growth factor-β (TGF-β) and nuclear transcription factor-κB p65 (NF-κB p65) in the gills of grass carp (P < 0.05). In contrast, the relative mRNA expression levels of occludin, zonula occludens-1 (ZO-1), claudin b, claudin 3, claudin 12, target of rapamycin (TOR) and inhibitor factor κBα (IκBα) in the gills were decreased by Cu (P < 0.05). However, pre

  4. Copper-induced tight junction mRNA expression changes, apoptosis and antioxidant responses via NF-κB, TOR and Nrf2 signaling molecules in the gills of fish: Preventive role of arginine

    International Nuclear Information System (INIS)

    Highlights: • Cu exposure induced oxidative stress via disruption of antioxidant system. • Cu exposure disrupted TJ mRNA expression through regulation of cytokines in fish. • Cu induced gill apoptosis partly via intrinsic pathway but not extrinsic pathway. • Cu exposure can regulate Nrf2, NF-κB and TOR signaling molecules in fish. • Arginine can effectively prevent Cu-induced fish gill damage. - Abstract: This study explored the possible preventive effects of dietary arginine on copper (Cu)-induced tight junction mRNA expression changes, apoptosis and antioxidant responses in the gills of young grass carp (Ctenopharyngodon idella). The results indicated that exposure to 0.7 mg/L (11.01 μmol/L) Cu for 96 h induced the production of reactive oxygen species (ROS), thereby increasing protein oxidation, lipid peroxidation and DNA damage in the gills of fish. However, these oxidative effects were prevented by arginine supplementation. Arginine also prevented the toxic effects of Cu on the activities of copper/zinc superoxide dismutase (SOD1), glutathione-S-transferase (GST), glutathione peroxidase (GPx), glutathione reductase (GR) and the glutathione (GSH) content (P < 0.05). However, Cu induced an adaptive increase in the activity of catalase (CAT), and arginine supplementation further increased CAT activity (P < 0.05). Moreover, Cu induced increases in the relative mRNA expressions of SOD1, CAT, GPx, GST, caspase-3, caspase-9, NF-E2-related factor 2 (Nrf2), Kelch-like-ECH-associated protein 1a (Keap1a), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-8 (IL-8), transforming growth factor-β (TGF-β) and nuclear transcription factor-κB p65 (NF-κB p65) in the gills of grass carp (P < 0.05). In contrast, the relative mRNA expression levels of occludin, zonula occludens-1 (ZO-1), claudin b, claudin 3, claudin 12, target of rapamycin (TOR) and inhibitor factor κBα (IκBα) in the gills were decreased by Cu (P < 0.05). However, pre

  5. The role of nitric oxide in L-arginine-stimulated growth hormone secretion%一氧化氮在L-精氨酸促生长激素分泌中的作用

    Institute of Scientific and Technical Information of China (English)

    蒋明玉; 蔡德培

    2011-01-01

    Amino acids have been pointed out as potent stimulators of growth hormone (GH) secretion. Among them, L-arginine is considered one of the most effective GH releasers. L-arginine increases GH release in pituitary when given orallyor intravenously. L-arginine-stimulated GH secretion can be mediated by several signaling systems, but nitric oxide (NO) dependent soluble guanylyl cyclase (sGC) -cyclic guanosine monophosphate (cGMP)-protein kinase G (PKG) pathway plays a key role in the process of L-arginine-stimulated GH secretion.%氨基酸是促进垂体生长激素(GH)分泌的潜在刺激因子.在所有氨基酸中,左旋精氨酸(L-精氨酸)促GH分泌作用最为明显.口服或静脉滴注精氨酸均可刺激垂体GH的分泌.L-精氨酸促GH的分泌可能涉及多种机制,但一氧化氮(NO)依赖的可溶性鸟苷酸环化酶-环鸟苷酸-蛋白激酶G(sGC-cGMP-PKG)信号转导通路在此过程中起着重要作用.

  6. A novel mutation affecting the arginine-137 residue of AVPR2 in dizygous twins leads to nephrogenic diabetes insipidus and attenuated urine exosome aquaporin-2

    DEFF Research Database (Denmark)

    Hinrichs, Gitte R; Hansen, Louise H; Nielsen, Maria R;

    2016-01-01

    Mutations in the vasopressin V2 receptor gene AVPR2 may cause X-linked nephrogenic diabetes insipidus by defective apical insertion of aquaporin-2 in the renal collecting duct principal cell. Substitution mutations with exchange of arginine at codon 137 can cause nephrogenic syndrome of inappropr...... administration. While a similar urine exosome release rate was shown between probands and controls by western blotting for the marker ALIX, there was a selective decrease in exosome aquaporin-2 versus aquaporin-1 protein in probands compared to controls....

  7. Twin-arginine translocation system (tat) mutants of Salmonella are attenuated due to envelope defects, not respiratory defects.

    Science.gov (United States)

    Craig, Maureen; Sadik, Adam Y; Golubeva, Yekaterina A; Tidhar, Avital; Slauch, James M

    2013-09-01

    The twin-arginine translocation system (Tat) transports folded proteins across the cytoplasmic membrane and is critical to virulence in Salmonella and other pathogens. Experimental and bioinformatic data indicate that 30 proteins are exported via Tat in Salmonella Typhimurium. However, there are no data linking specific Tat substrates with virulence. We inactivated every Tat-exported protein and determined the virulence phenotype of mutant strains. Although a tat mutant is highly attenuated, no single Tat-exported substrate accounts for this virulence phenotype. Rather, the attenuation is due primarily to envelope defects caused by failure to translocate three Tat substrates, the N-acetylmuramoyl-l-alanine amidases, AmiA and AmiC, and the cell division protein, SufI. Strikingly, neither the amiA amiC nor the sufI mutations alone conferred any virulence defect. Although AmiC and SufI have previously been localized to the divisome, the synthetic phenotypes observed are the first to suggest functional overlap. Many Tat substrates are involved in anaerobic respiration, but we show that a mutant completely deficient in anaerobic respiration retains full virulence in both the oral and systemic phases of infection. Similarly, an obligately aerobic mutant is fully virulent. These results suggest that in the classic mouse model of infection, S. Typhimurium is replicating only in aerobic environments. PMID:23822642

  8. Entropic stabilization of proteins and its proteomic consequences.

    Directory of Open Access Journals (Sweden)

    2005-09-01

    Full Text Available Evolutionary traces of thermophilic adaptation are manifest, on the whole-genome level, in compositional biases toward certain types of amino acids. However, it is sometimes difficult to discern their causes without a clear understanding of underlying physical mechanisms of thermal stabilization of proteins. For example, it is well-known that hyperthermophiles feature a greater proportion of charged residues, but, surprisingly, the excess of positively charged residues is almost entirely due to lysines but not arginines in the majority of hyperthermophilic genomes. All-atom simulations show that lysines have a much greater number of accessible rotamers than arginines of similar degree of burial in folded states of proteins. This finding suggests that lysines would preferentially entropically stabilize the native state. Indeed, we show in computational experiments that arginine-to-lysine amino acid substitutions result in noticeable stabilization of proteins. We then hypothesize that if evolution uses this physical mechanism as a complement to electrostatic stabilization in its strategies of thermophilic adaptation, then hyperthermostable organisms would have much greater content of lysines in their proteomes than comparably sized and similarly charged arginines. Consistent with that, high-throughput comparative analysis of complete proteomes shows extremely strong bias toward arginine-to-lysine replacement in hyperthermophilic organisms and overall much greater content of lysines than arginines in hyperthermophiles. This finding cannot be explained by genomic GC compositional biases or by the universal trend of amino acid gain and loss in protein evolution. We discovered here a novel entropic mechanism of protein thermostability due to residual dynamics of rotamer isomerization in native state and demonstrated its immediate proteomic implications. Our study provides an example of how analysis of a fundamental physical mechanism of

  9. Intracellular L-arginine concentration does not determine NO production in endothelial cells: Implications on the “L-arginine paradox”

    International Nuclear Information System (INIS)

    Highlights: ► Our findings provide a possible solution to the “L-arginine paradox”. ► Extracellular L-arginine concentration is the major determinant of NO production. ► Cellular L-arginine action is limited by cellular ARG transport, not the Km of NOS. ► We explain how L-arginine supplementation can work to increase endothelial function. -- Abstract: We examined the relative contributory roles of extracellular vs. intracellular L-arginine (ARG) toward cellular activation of endothelial nitric oxide synthase (eNOS) in human endothelial cells. EA.hy926 human endothelial cells were incubated with different concentrations of 15N4-ARG, ARG, or L-arginine ethyl ester (ARG-EE) for 2 h. To modulate ARG transport, siRNA for ARG transporter (CAT-1) vs. sham siRNA were transfected into cells. ARG transport activity was assessed by cellular fluxes of ARG, 15N4-ARG, dimethylarginines, and L-citrulline by an LC–MS/MS assay. eNOS activity was determined by nitrite/nitrate accumulation, either via a fluorometric assay or by15N-nitrite or estimated 15N3-citrulline concentrations when 15N4-ARG was used to challenge the cells. We found that ARG-EE incubation increased cellular ARG concentration but no increase in nitrite/nitrate was observed, while ARG incubation increased both cellular ARG concentration and nitrite accumulation. Cellular nitrite/nitrate production did not correlate with cellular total ARG concentration. Reduced 15N4-ARG cellular uptake in CAT-1 siRNA transfected cells vs. control was accompanied by reduced eNOS activity, as determined by 15N-nitrite, total nitrite and 15N3-citrulline formation. Our data suggest that extracellular ARG, not intracellular ARG, is the major determinant of NO production in endothelial cells. It is likely that once transported inside the cell, ARG can no longer gain access to the membrane-bound eNOS. These observations indicate that the “L-arginine paradox” should not consider intracellular ARG concentration as a

  10. Effects of Arginine Vasopressin on musical short-term memory

    Directory of Open Access Journals (Sweden)

    Roni Y. Granot

    2013-10-01

    Full Text Available Previous genetic studies showed an association between variations in the gene coding for the 1a receptor of the neuro-hormone arginine vasopressin (AVP and musical working memory (WM. The current study set out to test the influence of intranasal administration (INA of AVP on musical as compared to verbal WM using a double blind crossover (AVP – placebo design. Two groups of 25 males were exposed to 20 IU of AVP in one session, and 20 IU of saline water (placebo in a second session, one week apart. In each session subjects completed the tonal subtest from Gordon's Musical Aptitude Profile, the interval subtest from the Montreal Battery for Evaluation of Amusias (MBEA, and the forward and backward digit span tests. Scores in the digit span tests were not influenced by AVP. In contrast, in the music tests there was an AVP effect. In the MBEA test, scores for the group receiving placebo in the first session (PV were higher than for the group receiving vasopressin in the first session (VP (p < .05 with no main Session effect nor Group * Session interaction. In the Gordon test there was a main Session effect (p < .05 with scores higher in the second as compared to the first session, a marginal main Group effect (p = .093 and a marginal Group X Session interaction (p = 0.88. In addition we found that the group that received AVP in the first session scored higher on scales indicative of happiness, and alertness on the Positive and Negative Affect Scale, (PANAS. Only in this group and only in the music test these scores were significantly correlated with memory scores. Together the results reflect a complex interaction between AVP, musical memory, arousal, and contextual effects such as session, and base levels of memory. The results are interpreted in light of music's universal use as a means to modulate arousal on the one hand, and AVP's influence on mood, arousal, and social interactions on the other.

  11. Genes, enzymes and regulation of arginine biosynthesis in plants.

    Science.gov (United States)

    Slocum, Robert D

    2005-08-01

    Arabidopsis genes encoding enzymes for each of the eight steps in L-arginine (Arg) synthesis were identified, based upon sequence homologies with orthologs from other organisms. Except for N-acetylglutamate synthase (NAGS; EC 2.3.1.1), which is encoded by two genes, all remaining enzymes are encoded by single genes. Targeting predictions for these enzymes, based upon their deduced sequences, and subcellular fractionation studies, suggest that most enzymes of Arg synthesis reside within the plastid. Synthesis of the L-ornthine (Orn) intermediate in this pathway from L-glutamate occurs as a series of acetylated intermediates, as in most other organisms. An N-acetylornithine:glutamate acetyltransferase (NAOGAcT; EC 2.3.1.35) facilitates recycling of the acetyl moiety during Orn formation (cyclic pathway). A putative N-acetylornithine deacetylase (NAOD; EC 3.5.1.16), which participates in the "linear" pathway for Orn synthesis in some organisms, was also identified. Previous biochemical studies have indicated that allosteric regulation of the first and, especially, the second steps in Orn synthesis (NAGS; N-acetylglutamate kinase (NAGK), EC 2.7.2.8) by the Arg end-product are the major sites of metabolic control of the pathway in organisms using the cyclic pathway. Gene expression profiling for pathway enzymes further suggests that NAGS, NAGK, NAOGAcT and NAOD are coordinately regulated in response to changes in Arg demand during plant growth and development. Synthesis of Arg from Orn is further coordinated with pyrimidine nucleotide synthesis, at the level of allocation of the common carbamoyl-P intermediate. PMID:16122935

  12. Effects of arginine vasopressin on musical working memory.

    Science.gov (United States)

    Granot, Roni Y; Uzefovsky, Florina; Bogopolsky, Helena; Ebstein, Richard P

    2013-01-01

    Previous genetic studies showed an association between variations in the gene coding for the 1a receptor of the neuro-hormone arginine vasopressin (AVP) and musical working memory (WM). The current study set out to test the influence of intranasal administration (INA) of AVP on musical as compared to verbal WM using a double blind crossover (AVP-placebo) design. Two groups of 25 males were exposed to 20 IU of AVP in one session, and 20 IU of saline water (placebo) in a second session, 1 week apart. In each session subjects completed the tonal subtest from Gordon's "Musical Aptitude Profile," the interval subtest from the "Montreal Battery for Evaluation of Amusias (MBEA)," and the forward and backward digit span tests. Scores in the digit span tests were not influenced by AVP. In contrast, in the music tests there was an AVP effect. In the MBEA test, scores for the group receiving placebo in the first session (PV) were higher than for the group receiving vasopressin in the first session (VP) (p effect nor Group × Session interaction. In the Gordon test there was a main Session effect (p effect (p = 0.093) and a marginal Group × Session interaction (p = 0.88). In addition we found that the group that received AVP in the first session scored higher on scales indicative of happiness, and alertness on the positive and negative affect scale, (PANAS). Only in this group and only in the music test these scores were significantly correlated with memory scores. Together the results reflect a complex interaction between AVP, musical memory, arousal, and contextual effects such as session, and base levels of memory. The results are interpreted in light of music's universal use as a means to modulate arousal on the one hand, and AVP's influence on mood, arousal, and social interactions on the other. PMID:24151474

  13. Arginine deiminase resistance in melanoma cells is associated with metabolic reprogramming, glucose dependence, and glutamine addiction.

    Science.gov (United States)

    Long, Yan; Tsai, Wen-Bin; Wangpaichitr, Medhi; Tsukamoto, Takashi; Savaraj, Niramol; Feun, Lynn G; Kuo, Macus Tien

    2013-11-01

    Many malignant human tumors, including melanomas, are auxotrophic for arginine due to reduced expression of argininosuccinate synthetase-1 (ASS1), the rate-limiting enzyme for arginine biosynthesis. Pegylated arginine deiminase (ADI-PEG20), which degrades extracellular arginine, resulting in arginine deprivation, has shown favorable results in clinical trials for treating arginine-auxotrophic tumors. Drug resistance is the major obstacle for effective ADI-PEG20 usage. To elucidate mechanisms of resistance, we established several ADI-PEG20-resistant (ADI(R)) variants from A2058 and SK-Mel-2 melanoma cells. Compared with the parental lines, these ADI(R) variants showed the following characteristics: (i) all ADI(R) cell lines showed elevated ASS1 expression, resulting from the constitutive binding of the transcription factor c-Myc on the ASS1 promoter, suggesting that elevated ASS1 is the major mechanism of resistance; (ii) the ADI(R) cell lines exhibited enhanced AKT signaling and were preferentially sensitive to PI3K/AKT inhibitors, but reduced mTOR signaling, and were preferentially resistant to mTOR inhibitor; (iii) these variants showed enhanced expression of glucose transporter-1 and lactate dehydrogenase-A, reduced expression of pyruvate dehydrogenase, and elevated sensitivity to the glycolytic inhibitors 2-deoxy-glucose and 3-bromopyruvate, consistent with the enhanced glycolytic pathway (the Warburg effect); (iv) the resistant cells showed higher glutamine dehydrogenase and glutaminase expression and were preferentially vulnerable to glutamine inhibitors. We showed that c-Myc, not elevated ASS1 expression, is involved in upregulation of many of these enzymes because knockdown of c-Myc reduced their expression, whereas overexpressed ASS1 by transfection reduced their expression. This study identified multiple targets for overcoming ADI-PEG resistance in cancer chemotherapy using recombinant arginine-degrading enzymes. PMID:23979920

  14. Expression pattern of a nuclear encoded mitochondrial arginine-ornithine translocator gene from Arabidopsis

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    Schneider Anja

    2003-01-01

    Full Text Available Abstract Background Arginine and citrulline serve as nitrogen storage forms, but are also involved in biosynthetic and catabolic pathways. Metabolism of arginine, citrulline and ornithine is distributed between mitochondria and cytosol. For the shuttle of intermediates between cytosol and mitochondria transporters present on the inner mitochondrial membrane are required. Yeast contains a mitochondrial translocator for ornithine and arginine, Ort1p/Arg11p. Ort1p/Arg11p is a member of the mitochondrial carrier family (MCF essential for ornithine export from mitochondria. The yeast arg11 mutant, which is deficient in Ort1p/Arg11p grows poorly on media lacking arginine. Results High-level expression of a nuclear encoded Arabidopsis thaliana homolog (AtmBAC2 of Ort1p/Arg11p was able to suppress the growth deficiency of arg11. RT-PCR analysis demonstrated expression of AtmBAC2 in all tissues with highest levels in flowers. Promoter-GUS fusions showed preferential expression in flowers, i.e. pollen, in the vasculature of siliques and in aborted seeds. Variable expression was observed in leaf vasculature. Induction of the promoter was not observed during the first two weeks in seedlings grown on media containing NH4NO3, arginine or ornithine as sole nitrogen sources. Conclusion AtmBAC2 was isolated as a mitochondrial transporter for arginine in Arabidopsis. The absence of expression in developing seeds and in cotyledons of seedlings indicates that other transporters are responsible for storage and mobilization of arginine in seeds.

  15. Deprivation of L-Arginine Induces Oxidative Stress Mediated Apoptosis in Leishmania donovani Promastigotes: Contribution of the Polyamine Pathway

    Science.gov (United States)

    Mandal, Abhishek; Das, Sushmita; Roy, Saptarshi; Ghosh, Ayan Kumar; Sardar, Abul Hasan; Verma, Sudha; Saini, Savita; Singh, Ruby; Abhishek, Kumar; Kumar, Ajay; Mandal, Chitra; Das, Pradeep

    2016-01-01

    The growth and survival of intracellular parasites depends on the availability of extracellular nutrients. Deprivation of nutrients viz glucose or amino acid alters redox balance in mammalian cells as well as some lower organisms. To further understand the relationship, the mechanistic role of L-arginine in regulation of redox mediated survival of Leishmania donovani promastigotes was investigated. L-arginine deprivation from the culture medium was found to inhibit cell growth, reduce proliferation and increase L-arginine uptake. Relative expression of enzymes, involved in L-arginine metabolism, which leads to polyamine and trypanothione biosynthesis, were downregulated causing decreased production of polyamines in L-arginine deprived parasites and cell death. The resultant increase in reactive oxygen species (ROS), due to L-arginine deprivation, correlated with increased NADP+/NADPH ratio, decreased superoxide dismutase (SOD) level, increased lipid peroxidation and reduced thiol content. A deficiency of L-arginine triggered phosphatidyl serine externalization, a change in mitochondrial membrane potential, release of intracellular calcium and cytochrome-c. This finally led to DNA damage in Leishmania promastigotes. In summary, the growth and survival of Leishmania depends on the availability of extracellular L-arginine. In its absence the parasite undergoes ROS mediated, caspase-independent apoptosis-like cell death. Therefore, L-arginine metabolism pathway could be a probable target for controlling the growth of Leishmania parasites and disease pathogenesis. PMID:26808657

  16. 斑节对虾腺苷酸转移酶(PmANT)基因的cDNA克隆与表达分析%Molecular cloning and expression analysis of adenine nucleotide translocase (PmANT) in Penaeus monodon

    Institute of Scientific and Technical Information of China (English)

    孙文文; 周发林; 黄建华; 邱丽华; 杨其彬; 江世贵

    2013-01-01

    利用RACE技术获得了斑节对虾(Penaeus monodon)ANT基因(PmANT)的cDNA序列.该序列全长1 388 bp,开放阅读框(ORF)为930 bp,3’非编码区(UTR)为393 bp,5 '非编码区(UTR)为65 bp.ORF可编码309个氨基酸,分子量大约为33.622 ku.与所有ANT家族成员一样,PmANT蛋白具有3个重复同源的线粒体跨膜结构域,但不含信号肽和糖基化位点.相似性、同源性及系统进化树分析显示,斑节对虾的ANT基因与凡纳滨对虾的同源性和相似性最高,并与其聚为一支.采用荧光定量的方法研究了ANT基因在雌雄个体不同组织、卵巢不同发育阶段及未成熟和成熟精巢的差异表达情况.结果表明:PmANT的mRNA在各组织中都有表达,其中,在雄性个体的肌肉中表达量最高,其次为雌性肌肉,在精巢的表达量最低,且未成熟精巢低于成熟精巢.PmANT的mRNA在卵巢的表达量高于精巢,且在Ⅲ期卵巢表达量最高,Ⅳ期最低.为今后进一步研究该基因在斑节对虾性腺发育中的作用提供基础材料.%The adenine nucleotide translocase (ANT) is the most abundant mitochondrial inner membrane protein, which catalyzes the exchange of ADP and ATP between cytosol and mitochondria and participates in many models of mitochondrial apoptosis. In the present study, the full sequence of P. Monodon ANT gene was cloned and named PmANT. The full length cDNA of PmANT contained a 5' untranslated region (UTR) of 65 bp, a 3' UTR of 393 bp and an ORF of 930 bp encoding a polypeptide of 309 amino acids with an estimated molecular mass of 33. 622 ku. Like other animal ANTs, the structure of the PmANT protein consists of three homologous repeated domains. But there are no signal peptide and glycosylation sites in PmANT protein. Sequence alignment analysis showed that the PmANT with the ANT of Litopenaeus vannamei shared a similarity of 98. 7% and the homology of 97. 4%. Analysis of the tissue expression pattern of the PmANT showed that the PmANT m

  17. Effect of levo-dopa, arginine and exercise on pituitary growth hormone (GH) secretion

    International Nuclear Information System (INIS)

    Objective: To assess the stimulation effect on GH secretion with different agents (levo-dopa, arginine, exercise) in dwarf subjects. Methods: Growth hormone provocative tests were performed with levo-dopa (42 times), arginine (33) and standardized exercise (35) in 78 subjects classified as dwarfs. Serum GH levels were determined with RIA before the test and several times (at 30 min, intervals) afterwards with the peak value noted. The test results were divided into 3 categories: 1) peak value 10 ng/ml, test positive (no GH deficiency). Results: Peak values of serum GH after stimulation test with respective agents were: levo-dopa 14.09 ± 9.62 ng/ml, arginine 13.77 ± 6.83 ng/ml and exercise 12.68 ± 7.81 ng/ml with no significant differences among them. Positive rate after drug stimulation was significantly higher than that after exercise: levo-dopa 35.71% (15/42), arginine 36.36% (12/33) vs exercise 14.20% (5/35) P0.05). Conclusion: Diagnosis of GH deficiency (stimulation test negative) is best established after two negative provocative tests with different stimulant each time. Levo-dopa and arginine may be the drug of choice. (authors)

  18. L-arginine in combination with sildenafil potentiates the attenuation of hypoxic pulmonary hypertension in rats.

    Science.gov (United States)

    Al-Hiti, H; Chovanec, M; Melenovský, V; Vajnerová, O; Baňasová, A; Kautzner, J; Herget, J

    2013-01-01

    Chronic hypoxia induces an increased production of nitric oxide (NO) in pulmonary prealveolar arterioles. Bioavailability of the NO in the pulmonary vessels correlates with concentration of L-arginine as well as activity of phosphodiesterase-5 enzyme (PDE-5). We tested a hypothesis whether a combination of L-arginine and PDE-5 inhibitor sildenafil has an additive effect in reduction of the hypoxic pulmonary hypertension (HPH) in rats. Animals were exposed to chronic normobaric hypoxia for 3 weeks. In the AH group, rats were administered L-arginine during chronic hypoxic exposure. In the SH group, rats were administered sildenafil during chronic hypoxic exposure. In the SAH group, rats were treated by the combination of L-arginine as well as sildenafil during exposure to chronic hypoxia. Mean PAP, structural remodeling of peripheral pulmonary arterioles (%DL) and RV/LV+S ratio was significantly decreased in the SAH group compared to hypoxic controls even decreased compared to the AH and the SH groups in first two measured parameters. Plasmatic concentration of cGMP and NOx were significantly lower in the SAH group compared to hypoxic controls. We demonstrate that NO synthase substrate L-arginine and phosphodiesterase-5 inhibitor sildenafil administered in combination are more potent in attenuation of the HPH compared to a treatment by substances given alone. PMID:23869884

  19. The impact of arginine-modified chitosan-DNA nanoparticles on the function of macrophages

    International Nuclear Information System (INIS)

    It has been demonstrated that incorporation of arginine moieties into chitosan significantly elevates the transgenic efficacy of the chitosan. However, little is known about the impact of arginine-modified chitosan on the function of macrophages, which play a vitally important role in the inflammatory response of the body to foreign substances, especially particulate substances. This study was designed to investigate the impact of arginine-modified chitosan/DNA nanoparticles on the function of the murine macrophage through observation of phagocytic activity and production of pro-inflammatory cytokines (IL-1β, IL-6, IL-10, IL-12, and TNF-α). Results showed that both chitosan/DNA nanoparticles and arginine-modified chitosan/DNA nanoparticles, containing 20 μg/mL DNA, were internalized by almost all the macrophages in contact. This led to no significant changes, compared to the non-exposure group, in production of cytokines and phagocytic activity of the macrophages 24 h post co-incubation, whereas exposure to LPS induced obviously elevated cytokine production and phagocytic activity, suggesting that incorporation of arginine moieties into chitosan does not have a negative impact on the function of the macrophages.

  20. Arginine: A Potent Prey Attractant to Predatory Newts in Mountain Streams

    Science.gov (United States)

    Ferrer, R. P.; Zimmer, R. K.

    2005-05-01

    Chemoreception of aquatic organisms has been well-studied in the laboratory, but rarely in the field. The California newt, Taricha torosa, in natural stream habitats is an excellent animal for exploring behavioral responses to prey odors. Here, we selected 13 amino acids for field bioassays based on their concentrations in prey tissue extracts. Bioassays were calibrated for stimulus dilution by means of fluorescent dye releases and flow-through spectrofluorometry. Moreover, hydrodynamic properties of stream flows were determined using an electromagnetic current meter. Of all amino acids tested, only arginine, alanine and glycine were significantly attractive (relative to stream water controls). These three substances caused free-ranging newts to turn upstream and swim towards the odor sources. Additional experiments showed that arginine was the most effective attractant, evoking plume-tracking behavior at concentrations as low as 10 nM. In subsequent trials, nine arginine analogs were tested, but each compound failed to elicit a significant response. Even subtle changes to arginine, such as the addition of a single carbon to the side chain, destroyed all bioactivity. Within its natural habitat, the California newt thus exhibits keen sensitivity and narrow tuning to the free amino acid, arginine, a chemical signal of its prey.

  1. Biochemical and biological activity of arginine deiminase from Streptococcus pyogenes M22.

    Science.gov (United States)

    Starikova, Eleonora A; Sokolov, Alexey V; Vlasenko, Anna Yu; Burova, Larisa A; Freidlin, Irina S; Vasilyev, Vadim B

    2016-04-01

    Streptococcus pyogenes (group A Streptococcus; GAS) is an important gram-positive extracellular bacterial pathogen responsible for a number of suppurative infections. This micro-organism has developed complex virulence mechanisms to avoid the host's defenses. We have previously reported that SDSC from GAS type M22 causes endothelial-cell dysfunction, and inhibits cell adhesion, migration, metabolism, and proliferation in a dose-dependent manner, without affecting cell viability. This work aimed to isolate and characterize a component from GAS type M22 supernatant that suppresses the proliferation of endothelial cells (EA.hy926). In the process of isolating a protein possessing antiproliferative activity we identified arginine deiminase (AD). Further study showed that this enzyme is most active at pH 6.8. Calculating Km and Vmax gave the values of 0.67 mmol·L(-1) and 42 s(-1), respectively. A distinctive feature of AD purified from GAS type M22 is that its optimum activity and the maximal rate of the catalytic process is close to neutral pH by comparison with enzymes from other micro-organisms. AD from GAS type M22 suppressed the proliferative activity of endothelial cells in a dose-dependent mode. At the same time, in the presence of AD, the proportion of cells in G0/G1 phase increased. When l-Arg was added at increasing concentrations to the culture medium containing AD (3 μg·mL(-1)), the enzyme's capacity to inhibit cell proliferation became partially depressed. The proportion of cells in phases S/G2 increased concomitantly, although the cells did not fully recover their proliferation activity. This suggests that AD from GAS type M22 has potential for the suppression of excessive cell proliferation. PMID:26695833

  2. Mapping and characterization of antigenic epitopes of arginine kinase of Scylla paramamosain.

    Science.gov (United States)

    Yang, Yang; Cao, Min-Jie; Alcocer, Marcos; Liu, Qing-Mei; Fei, Dan-Xia; Mao, Hai-Yan; Liu, Guang-Ming

    2015-06-01

    Arginine kinase (AK) is a panallergen present in crustaceans, which can induce an immunoglobulin (Ig) E-mediated immune response in humans. The aim of this work was to map and characterize the antigenic epitopes of Scylla paramamosain AK. Specific-protein-A-enriched IgG raised in rabbits against purified S. paramamosain AK was used to screen a phage display random peptide library. Five AK mimotope clones were identified among 20 random clones after biopanning. Four conformational epitopes D3A4K43M1A5T49T44I7, L31K33V35T32E11E18F14S34D37, V177G172M173D176Q178T174L181K175L187, and R202L170Y203E190P205W204L187T206Y145 were identified with the program LocaPep, and mapped to S. paramamosain AK. The key amino acids of these conformational epitopes were D3, K33, T174, and W204, respectively. On the basis of biopanning, six IgE-specific peptides were mapped with synthetic overlapping peptides using the sera from crab-allergic patients, and four seropositive peptides (amino acids 113-127, 127-141, 141-155, and 204-218) were confirmed as linear epitopes in a degranulation assay in RBL-2H3 cells. Stability experiments showed that the structural integrity of AK is essential for its allergenicity, and the intramolecular disulfide bond at Cys201-Cys271 is essential for its structural stability. PMID:25728640

  3. Screening of Natural Antioxidants by using L-Arginine induced acute Pancreatitis Model

    Directory of Open Access Journals (Sweden)

    Sandeep Biradar

    2012-12-01

    Full Text Available Medicinal plants and their active constituents are traditionally used for herbal preparations and were proposed for their interesting antioxidant activities. Nearly all the medicinal plants are used for the therapeutic action and some of them are used in the investigation. Inflammation of pancreas of the exocrine part is called as acute pancreatitis. Inflammatory mediators and oxidative mediators are major factors for development of acute pancreatitis (AP. In the present study the protective effects of lawsone, myrcene, limonene, "-pinene and the underlying mechanisms in an experimental pancreatitis model. AP was induced in eleven groups of rats (n =6 by Larginine (2x2.5 g/kg, intraperitoneal, 1 h apart and 1 h later, they received a single oral dose of lawsone, myrcene, limonene, "- pinene, (100 and 200 mg/kg respectively, vehicle (3% Tween 80 and methylprednisolone (30 mg/kg. A saline (0.9% NaCl treated group served as a normal control. The efficacy was determined at 24 h by determination of serum levels of amylase, lipase and proinflammatory cytokines [tumor necrosis factor (TNF-a, C-reactive proteins and interleukin (IL], nitrate/nitrite levels, and the wet weight/body weight ratio. Lawsone, myrcene, limonene, "-pinene and methylprednisolone treatments significantly (P < 0.05 attenuated the L-arginine induced increases in pancreatic wet weight/body weight ratio, decreased the serum levels of amylase and lipase, TNF-a, IL-6 and significantly lowered pancreatic levels of TBARS, and nitrate/nitrite. The histoimmunological findings further proved the amelioration of pancreatic injury by lawsone, myrcene, limonene and "-pinene. Hence it proved anti-inflammatory and antioxidant agent property of lawsone, myrcene, limonene and "-pinene.

  4. The development of poly-L-arginine-coated liposomes for gene delivery

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    Opanasopit P

    2011-10-01

    Full Text Available Praneet Opanasopit1, Jintana Tragulpakseerojn1, Auayporn Apirakaramwong1, Tanasait Ngawhirunpat1, Theerasak Rojanarata1, Uracha Ruktanonchai21Faculty of Pharmacy, Silpakorn University, Nakhon Pathom, Thailand; 2National Nanotechnology Center, Thailand Science Park, Pathumthani, Thailand Abstract: In this study, liposomes coated with cationic polymers, poly-L-arginine (PLA, were assessed as a promising gene transfer system in human cervical carcinoma (HeLa cells and human hepatoma cell line (Huh7 cells. The liposomes were prepared using egg yolk phosphatidylcholine and sodium oleate in the molar ratio of 10:2 with an ultrasonic generator and then coated with PLA. The PLA-coated liposomes (PCLs formed complexes with plasmid DNA encoding green fluorescent protein. The complexes were characterized by agarose gel electrophoresis and investigated for their transfection efficiency in HeLa and Huh7 cells. The data were compared with PLA/DNA complexes and the positive control Lipofectamine 2000TM. The results showed that complete PCL/DNA complexes were formed at weight ratios of more than 0.05. Efficient gene transfer by PCLs was dependent on the cell type. The transfection efficiency of PCLs was about two times higher than that of PLA/DNA complexes in both HeLa cells and Huh7 cells. Cytotoxicity was determined by the 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyl tetrazolium bromide assay and showed that 80%-100% of both of the cells were viable after treating PCL/DNA complexes. The present results demonstrate that PCLs are a promising, nonviral gene carrier with low toxicity.Keywords: PLA-coated liposomes, PLA, gene delivery, transfection efficiency 

  5. Deletion of Genes Encoding Arginase Improves Use of "Heavy" Isotope-Labeled Arginine for Mass Spectrometry in Fission Yeast.

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    Weronika E Borek

    Full Text Available The use of "heavy" isotope-labeled arginine for stable isotope labeling by amino acids in cell culture (SILAC mass spectrometry in the fission yeast Schizosaccharomyces pombe is hindered by the fact that under normal conditions, arginine is extensively catabolized in vivo, resulting in the appearance of "heavy"-isotope label in several other amino acids, most notably proline, but also glutamate, glutamine and lysine. This "arginine conversion problem" significantly impairs quantification of mass spectra. Previously, we developed a method to prevent arginine conversion in fission yeast SILAC, based on deletion of genes involved in arginine catabolism. Here we show that although this method is indeed successful when (13C6-arginine (Arg-6 is used for labeling, it is less successful when (13C6(15N4-arginine (Arg-10, a theoretically preferable label, is used. In particular, we find that with this method, "heavy"-isotope label derived from Arg-10 is observed in amino acids other than arginine, indicating metabolic conversion of Arg-10. Arg-10 conversion, which severely complicates both MS and MS/MS analysis, is further confirmed by the presence of (13C5(15N2-arginine (Arg-7 in arginine-containing peptides from Arg-10-labeled cells. We describe how all of the problems associated with the use of Arg-10 can be overcome by a simple modification of our original method. We show that simultaneous deletion of the fission yeast arginase genes car1+ and aru1+ prevents virtually all of the arginine conversion that would otherwise result from the use of Arg-10. This solution should enable a wider use of heavy isotope-labeled amino acids in fission yeast SILAC.

  6. Mcp3 is a novel mitochondrial outer membrane protein that follows a unique IMP-dependent biogenesis pathway.

    Science.gov (United States)

    Sinzel, Monika; Tan, Tao; Wendling, Philipp; Kalbacher, Hubert; Özbalci, Cagakan; Chelius, Xenia; Westermann, Benedikt; Brügger, Britta; Rapaport, Doron; Dimmer, Kai Stefan

    2016-07-01

    Mitochondria are separated from the remainder of the eukaryotic cell by the mitochondrial outer membrane (MOM). The MOM plays an important role in different transport processes like lipid trafficking and protein import. In yeast, the ER-mitochondria encounter structure (ERMES) has a central, but poorly defined role in both activities. To understand the functions of the ERMES, we searched for suppressors of the deficiency of one of its components, Mdm10, and identified a novel mitochondrial protein that we named Mdm10 complementing protein 3 (Mcp3). Mcp3 partially rescues a variety of ERMES-related phenotypes. We further demonstrate that Mcp3 is an integral protein of the MOM that follows a unique import pathway. It is recognized initially by the import receptor Tom70 and then crosses the MOM via the translocase of the outer membrane. Mcp3 is next relayed to the TIM23 translocase at the inner membrane, gets processed by the inner membrane peptidase (IMP) and finally integrates into the MOM. Hence, Mcp3 follows a novel biogenesis route where a MOM protein is processed by a peptidase of the inner membrane. PMID:27226123

  7. GABA Production in Lactococcus lactis Is Enhanced by Arginine and Co-addition of Malate.

    Science.gov (United States)

    Laroute, Valérie; Yasaro, Chonthicha; Narin, Waranya; Mazzoli, Roberto; Pessione, Enrica; Cocaign-Bousquet, Muriel; Loubière, Pascal

    2016-01-01

    Lactococcus lactis NCDO 2118 was previously selected for its ability to decarboxylate glutamate to γ-aminobutyric acid (GABA), an interesting nutritional supplement able to improve mood and relaxation. Amino acid decarboxylation is generally considered as among the biochemical systems allowing lactic acid bacteria to counteracting acidic stress and obtaining metabolic energy. These strategies also include arginine deiminase pathway and malolactic fermentation but little is known about their possible interactions of with GABA production. In the present study, the effects of glutamate, arginine, and malate (i.e., the substrates of these acid-resistance pathways) on L. lactis NCDO 2118 growth and GABA production performances were analyzed. Both malate and arginine supplementation resulted in an efficient reduction of acidity and improvement of bacterial biomass compared to glutamate supplementation. Glutamate decarboxylation was limited to narrow environmental conditions (pH < 5.1) and physiological state (stationary phase). However, some conditions were able to improve GABA production or activate glutamate decarboxylation system even outside of this compass. Arginine clearly stimulated glutamate decarboxylation: the highest GABA production (8.6 mM) was observed in cultures supplemented with both arginine and glutamate. The simultaneous addition of arginine, malate, and glutamate enabled earlier GABA production (i.e., during exponential growth) at relatively high pH (6.5). As far as we know, no previous study has reported GABA production in such conditions. Although further studies are needed to understand the molecular basis of these phenomena, these results represent important keys suitable of application in GABA production processes. PMID:27458444

  8. Arginine-containing desensitizing toothpaste for the treatment of dentin hypersensitivity: a meta-analysis

    Science.gov (United States)

    Yang, Zheng-yan; Wang, Fei; Lu, Keke; Li, Yue-heng; Zhou, Zhi

    2016-01-01

    Objective To estimate the effect of arginine-containing desensitizing toothpaste on dentin hypersensitivity (DH). Methods Databases including China National Knowledge Infrastructure, VIP Database for Chinese Technical Periodicals, China Biology Medicine disc, Wangfang Data, PubMed, Web of Science, and Cochrane Trials Register were searched, and Google was used as a supplementary tool to search for information through February 2014. Randomized controlled trials (RCTs) of the treatment of DH with arginine-containing toothpaste were included. Relevant information was extracted, and a quality evaluation was performed. Meta-analyses were performed using RevMan 5.2 software. Results Eighteen RCTs with 1,423 patients were included. The results of the meta-analyses demonstrated that at days 0 and 3; weeks 2, 4, and 8; and more than 12 weeks, arginine-containing toothpaste led to significantly improved results on the tactile sensitivity test (standardized mean difference [SMD] =1.95, 95% confidence interval [CI] [1.14, 2.76]) and the air-blast test (SMD =−1.60, 95% CI [−2.14, −1.05]) at 4 weeks and the tactile sensitivity test (SMD =2.01, 95% CI [1.41, 2.61]) and the air-blast test (SMD =−1.41, 95% CI [−1.83, −0.98]) at 8 weeks compared to toothpastes containing other desensitizing components, thus indicating a superior therapeutic effect of arginine-containing desensitizing toothpaste. However, no significant differences between arginine-containing toothpaste and toothpastes containing other desensitizing components were observed in the air-blast test at days 0 and 3 and week 2 and in the tactile sensitivity and air-blast tests at more than 12 weeks. Conclusion The current evidence indicates that arginine-containing toothpaste is effective for DH. However, further high-quality, large-sample RCTs are needed. PMID:26793006

  9. Potential protective effect of arginine against 4-nitrophenol-induced ovarian damage in rats.

    Science.gov (United States)

    Xu, Wei-Feng; Li, Yan-Sen; Dai, Peng-Yuan; Li, Chun-Mei

    2016-01-01

    4-nitrophenol (PNP) is generally regarded as a diesel exhaust particle (DEP). Arginine plays an important role as a new feed additive, possessing highly efficient antioxidant activities. Here we investigated the effects of dietary supplementation with arginine against ovarian damage induced by PNP in rats. A total of thirty-two female rats postnatal day 28 (PND 28) were randomly divided into four groups. Two groups were fed with basal diet or 13 g/kg arginine in diet for 4 weeks, respectively; the other two groups were given PNP (100 mg/kg b.w.) daily by subcutaneous injection for 2 weeks following pretreatment with either basal diet or arginine diet for 2 weeks. The values of body weight gain (BWG), average daily gain (ADG) and percentage weight gain (PWG) upon PNP treatment were significantly reduced than those in other groups. The relative liver weight in the PNP group was significantly decreased compared with the control group. Treatment with PNP significant reduced the number of corpora lutea, although serum 17β-estradiol (E2) and progesterone (P4) concentrations were unchanged. The morphology of the ovaries in PNP-treated rats displayed necrosis, follicular deformation and granulosa cells irregular arrangement. Moreover, exposure to PNP enhanced production of malondialdehyde (MDA) and hydrogen peroxide (H2O2), and decreased the activities of total superoxide dismutase (T-SOD) and catalase (CAT), and the co-administration of arginine can attenuate the oxidative stress caused by PNP. These results suggest that arginine may have a protective effect against ovarian damage induced by PNP owing to its antioxidant capacity effect. PMID:27193729

  10. Linear short histidine and cysteine modified arginine peptides constitute a potential class of DNA delivery agents.

    Science.gov (United States)

    Mann, Anita; Shukla, Vasundhara; Khanduri, Richa; Dabral, Spoorti; Singh, Harpal; Ganguli, Munia

    2014-03-01

    The success of gene therapy relies on the development of safe and efficient multifunctional carriers of nucleic acids that can overcome extra- and intracellular barriers, protect the nucleic acid and mediate its release at the desired site allowing gene expression. Peptides bear unique properties that are indispensable for any carrier, e.g., they can mediate DNA condensation, cellular targeting, membrane translocation, endosomal escape and nuclear localization. In an effort to design a multifunctional peptide, we have modified an arginine homopeptide R16 by replacement of seven arginines with histidines and addition of one cysteine at each end respectively to impart endosomal escape property while maintaining the DNA condensation and release balance. Addition of histidines imparts endosomal escape property to arginine homopeptide, but their arrangement with respect to arginines is more critical in controlling DNA condensation, release and transfection efficiency. Intriguingly, R5H7R4 peptide where charge/arginine is distributed in blocks is preferred for strong condensation while more efficient transfection is seen in the variants R9H7 and H4R9H3, which exhibit weak condensation and strong release. Addition of cysteine to each of these peptides further fine-tuned the condensation-release balance without application of any oxidative procedure unlike other similar systems reported in the literature. This resulted in a large increase in the transfection efficiency in all of the histidine modified peptides irrespective of the arginine and histidine positions. This series of multifunctional peptides shows comparable transfection efficiency to commercially available transfection reagent Lipofectamine 2000 at low charge ratios, with simple preparative procedure and exhibits much less toxicity. PMID:24476132

  11. The Metabolic Conversion of Arginine in the Rumen Wall and its Importance in Ruminant Nitrogen Metabolism

    International Nuclear Information System (INIS)

    The functions of arginase and urease of the rumen wall were investigated in vitro and in vivo. Surviving ruminal mucosae of cattle were incubated for four hours. 14C-arginine-HCl, uniformly labelled, was added to the serosal side at a concentration of 10 pmol/mi. About 25% of the added arginine was used during the incubation by the ruminal tissue. In comparison with controls an increased amount of 14C-omithine, urea, and ammonia were formed in the mucosa and appeared on both sides. The increase was due to arginase and urease functions. It was estimated that about 50% of the urea formed by arginine breakdown were present at the mucosa side, mainly in the form of ammonia. Of the omithine simultaneously formed, 85% remained on the serosa side. Remarkable individual variations of omithine and urea formation were found from animal to animal. The in-vivo experiments were performed using goats with catheters placed in the right ruminal artery and vein. We injected 90 μCi of 14C-arginine into the ruminal artery. When 80 g of soluble starch were added to the rumen the activity and concentration of ornithine increased in the ruminal venous blood showing an arterial-venous difference. The radioactivity of urea in blood taken from the ruminal vein and the carotid artery did not show any difference. When starch was omitted from the rumen a comparable difference of omithine concentration was not found. It is assumed that the enzymes arginase and urease of the rumen wall are involved in nitrogen recycling processes. Blood arginine may be hydrolysed in the rumen wall forming urea and ornithine. Urea formed by arginine breakdown may be split to CO2 and ammonia. The experiments produced evidence that the ammonia formed preferably enters the rumen content. The nitrogen transfer through the rumen wall may be affected by varying activities of arginase. (author)

  12. Effects of chronic oral L-arginine administration on the L-arginine/NO pathway in patients with peripheral arterial occlusive disease or coronary artery disease: L-Arginine prevents renal loss of nitrite, the major NO reservoir.

    Science.gov (United States)

    Schneider, Jessica Y; Rothmann, Sabine; Schröder, Frank; Langen, Jennifer; Lücke, Thomas; Mariotti, François; Huneau, Jean François; Frölich, Jürgen C; Tsikas, Dimitrios

    2015-09-01

    Despite saturation of nitric oxide (NO) synthase (NOS) by its substrate L-arginine (Arg), oral and intravenous supplementation of Arg may enhance NO synthesis, a phenomenon known as "The L-arginine paradox". Yet, Arg is not only a source of NO, but is also a source for guanidine-methylated (N (G)) arginine derivatives which are all inhibitors of NOS activity. Therefore, Arg supplementation may not always result in enhanced NO synthesis. Concomitant synthesis of N (G)-monomethyl arginine (MMA), N (G),N (G)-dimethylarginine (asymmetric dimethylarginine, ADMA) and N (G),N (G´)-dimethylarginine (symmetric dimethylarginine, SDMA) from supplemented Arg may outweigh and even outbalance the positive effects of Arg on NO. Another possible, yet little investigated effect of Arg supplementation may be alteration of renal function, notably the influence on the excretion of nitrite in the urine. Nitrite is the autoxidation product of NO and the major reservoir of NO in the circulation. Nitrite and Arg are reabsorbed in the proximal tubule of the nephron and this reabsorption is coupled, at least in part, to the renal carbonic anhydrase (CA) activity. In the present placebo-controlled studies, we investigated the effect of chronic oral Arg supplementation of 10 g/day for 3 or 6 months in patients suffering from peripheral arterial occlusive disease (PAOD) or coronary artery disease (CAD) on the urinary excretion of nitrite relative to nitrate. We determined the urinary nitrate-to-nitrite molar ratio (UNOxR), which is a measure of nitrite-dependent renal CA activity before and after oral intake of Arg or placebo by the patients. The UNOxR was also determined in 6 children who underwent the Arg test, i.e., intravenous infusion of Arg (0.5 g Arg/kg bodyweight) for 30 min. Arg was well tolerated by the patients of the three studies. Oral Arg supplementation increased Arg (plasma and urine) and ADMA (urine) concentrations. No appreciable changes were seen in NO (in PAOD and CAD) and

  13. Untersuchungen zur Bedeutung von Arginin für die induzierte NO-Synthese im ZNS

    OpenAIRE

    Fischmann, Boris

    2005-01-01

    Die induzierbare NO-Synthase (iNOS) wird bei verschiedenen neurodegenerativen Erkrankungen verstärkt exprimiert und steht im Verdacht, an der Pathogenese dieser Erkrankungen beteiligt zu sein. Da Arginin der iNOS als Substrat dient, steht die induzierte NO-Produktion in engem Zusammenhang mit dem Argininstoffwechsel. In der vorliegenden Arbeit wurden in Modellsystemen für Microgliazellen (N11-Zellen) und für Astrocyten (C6-BU-1-Zellen) Aspekte der induzierten Produktion von NO und des Arginin...

  14. Effect of L-arginine on neuromuscular transmission of the chick biventer cervicis muscle

    Directory of Open Access Journals (Sweden)

    B. Esfandiar

    2008-01-01

    Full Text Available biventer cervicis muscleD. Effect of L-arginine on neuromuscular transmission of the chick EsfandiarAbstractBackground and Purpose: NO is a short-lived gas molecule generated by degradation of L-arg to citrulline and by the activation of enzyme NOS Ca2+/calmodulin-dependent. There are multiple NOS isoforms that strongly are expressed in skeletal muscle, suggesting the crucial role of NO in regulating muscular metabolism and function. In this study, the effect of L-arginine was examined at the neuromuscular junction of the chick biventer cervicis muscle.Materials and Methods: Biventer cervicis muscle preparations from chick’s age of 3 weeks were set up in the organ bath. The organ bath had a vessel with volume of about 70 ml; it contained Tyrode solution aerated with oxygen and was kept at 37º C. NO levels was also measured in the chick biventer cervicis muscle homogenates, using spectrophotometer method for the direct detection of NO, nitrite and nitrate. Total nitrite (nitrite+nitrate was measured by a spectrophotometer at 540 nm after the conversion of nitrate to nitrite by copperized cadmium granules.Results: L-Arginine at 500 µg/ml, decreased twitch response to electrical stimulation, and produced rightward shift of the dose-response curve for acetylcholine or carbachol. L-arginine at 1000 µg/ml produced a strong shift to the right of the dose-response curve for acetylcholine or carbachol with a reduction in efficacy. The inhibitory effect of L-arginine on the twitch response was blocked by caffeine (200 µg/ml. NO levels were found to be significantly increased in concentrations 500 and 1000 µg/ml of L-arginine in comparison with the control group (p < 0.001.Conclusion: These findings indicate a possible role of increased NO levels in the suppressive action of L-arginie on the twitch response. In addition, the results indicate that the post-junctional antagonistic action of L-arginine is probably the result of impaired sarcoplasmic

  15. Performance and nitrogen balance of laying hens fed increasing levels of digestible lysine and arginine

    OpenAIRE

    Fabyola Barros de Carvalho; José Henrique Stringhini; Maíra Silva Matos; Roberto Moraes Jardim Filho; Marcos Barcellos Café; Nadja Susana Mogyca Leandro; Maria Auxiliadora Andrade

    2012-01-01

    The objective of this experiment was to evaluate the effect of two digestible lysine levels and four digestible arginine levels on laying hens from 24 to 48 weeks of age. Three hundred and twenty Lohmann LSL laying hens were allotted in a completely randomized design in a 2 × 4 factorial arrangement, with two levels of digestible lysine (700 and 900 mg/kg of diet) and four digestible arginine levels (700, 800, 900 and 1000 mg/kg of diet). Results indicated requirement of 884 and 830 mg of dig...

  16. Arginine vasopressin stimulates phosphoinositide turnover in an enriched rat Leydig cell preparation

    DEFF Research Database (Denmark)

    Nielsen, J.R.; Hansen, Harald S.; Jensen, B.

    1989-01-01

    An enriched rat Leydig cell preparation was preincubated with [C]arachidonic acid. Stimulation of the cells with arginine vasopressin (AVP) (1 µM) for 2 min caused a significant increase in labelled phosphatidic acid and a significant fall in radioactivity in phosphatidylinositol and phosphatidyl......An enriched rat Leydig cell preparation was preincubated with [C]arachidonic acid. Stimulation of the cells with arginine vasopressin (AVP) (1 µM) for 2 min caused a significant increase in labelled phosphatidic acid and a significant fall in radioactivity in phosphatidylinositol and...

  17. The Role of Arginin and Uric Acid on Portulaca Grandiflora Growth under Saline Conditions.

    OpenAIRE

    Mahmoud Yagi

    2014-01-01

    Seeds of Portulaca grandiflora were soaked in distilled water, NaCl or NaCl with uric acid or arginine or in one of the amino acids. Treatment of seeds of Portulaca grandiflora with arginine and uric acid under saline conditions increased the percent of seed germination from 55% to 80%. Incorporation of these amino acids in the nutrient medium also significantly enhanced the dry weights as well as the contents of chlorophyll and ascorbic acid in the seedlings. Levels of both total amino acids...

  18. Reengineering of a Corynebacterium glutamicum l-Arginine and l-Citrulline Producer▿

    OpenAIRE

    Ikeda, Masato; Mitsuhashi, Satoshi; Tanaka, Kenji; Hayashi, Mikiro

    2009-01-01

    Toward the creation of a robust and efficient producer of l-arginine and l-citrulline (arginine/citrulline), we have performed reengineering of a Corynebacterium glutamicum strain by using genetic information of three classical producers. Sequence analysis of their arg operons identified three point mutations (argR123, argG92up, and argG45) in one producer and one point mutation (argB26 or argB31) in each of the other two producers. Reconstitution of the former three mutations or of each argB...

  19. Dietary l-Arginine Supplementation Protects Weanling Pigs from Deoxynivalenol-Induced Toxicity

    OpenAIRE

    Li Wu; Peng Liao; Liuqin He; Zemeng Feng; Wenkai Ren; Jie Yin; Jielin Duan; Tiejun Li; Yulong Yin

    2015-01-01

    This study was conducted to determine the positive effects of dietary supplementation with l-arginine (Arg) on piglets fed a deoxynivalenol (DON)-contaminated diet. A total of eighteen, 28-day-old healthy weanling pigs were randomly assigned into one of three groups: uncontaminated basal diet (control group), 6 mg/kg DON-contaminated diet (DON group) and 6 mg/kg DON + 1% l-arginine (DON + ARG group). After 21 days of Arg supplementation, piglets in the DON and DON + ARG groups were challenged...

  20. Nonspecific blockade of vascular free radical signals by methylated arginine analogues

    OpenAIRE

    Pedro M.A.; Augusto O.; Barbeiro H.V.; Carvalho M.H.C.; da-Luz P.L.; Laurindo F.R.M.

    1998-01-01

    Methylated arginine analogues are often used as probes of the effect of nitric oxide; however, their specificity is unclear and seems to be frequently overestimated. This study analyzed the effects of NG-methyl-L-arginine (L-NMMA) on the endothelium-dependent release of vascular superoxide radicals triggered by increased flow. Plasma ascorbyl radical signals measured by direct electron paramagnetic resonance spectroscopy in 25 rabbits increased by 3.8 ± 0.7 nmol/l vs baseline (28.7 ± 1.4 nmol...

  1. Efficacy L-Arginine In Patients With Nonalcoholic Steatohepatitis Associated With Metabolic Syndrome

    Directory of Open Access Journals (Sweden)

    Oleksandr Fediv

    2015-01-01

    Full Text Available Abstract Background and Purpose Recent research in the field of hematology indicate that among the many pathogenic mechanisms of development and progression of nonalcoholic steatohepatitis NASH which occurs on the background of the metabolic syndrome an important role is played by endothelial dysfunction and violations of haemocoagulation. The aim of this research was to study the effectiveness of L-arginine as it corrects endothelial dysfunction and disorders of homeostasis haemocoagulation link in patients with NASH associated with the metabolic syndrome. Subjects and Methods 128 patients with nonalcoholic steatohepatitis associated with metabolic syndrome were examined. Some patients 63 persons received standard treatment according to national guidelines. To another group 65 patients on the background of basic therapy L-arginine hydrochloride followed by transition to oral form of L-arginine aspartate was administered. Blood levels of stable nitrogen monoxide metabolites nitrites nitrates endothelin-1 and plasma recalcification time prothrombin time thrombin time activated partial thromboplastin time fibrinogen plasma level activity of antithrombin III and coagulation factor XIII potential activity of plasminogen plasma fibrinolytic blood activity were studied. Results Originally significantly increased levels of endothelin-1 decreased after the therapy in all studied groups but more noticeable changes in the group with L-arginine appointment were observed p0.05. In the studied groups normalization of stable nitrogen monoxide metabolites after treatment was also noticed. Significant p0.05 increase in all haemocoagulation time characteristics and activities of antithrombin-III and factor XIII was found. The positive effect of L-arginine on blood fibrinolytic activity was noted. Discussion and Conclusion Combined therapy of nonalcoholic steatohepatitis associated with metabolic syndrome with a differentiated degreeal L-arginine assignment by

  2. The microbiome, intestinal function, and arginine metabolism of healthy Indian women are different from those of American and Jamaican women

    Science.gov (United States)

    Indian women have slower arginine flux during pregnancy compared with American and Jamaican women. Arginine is a semi-essential amino acid that becomes essential during periods of rapid lean tissue deposition. It is synthesized only from citrulline, a nondietary amino acid produced mainly in the gut...

  3. DNA strand break dependence on Tris and arginine scavenger concentrations under ultra-soft X-ray irradiation: the contribution of secondary arginine radicals.

    Science.gov (United States)

    Souici, Mounir; Khalil, Talat Tariq; Boulanouar, Omar; Belafrites, Abdelfettah; Mavon, Christophe; Fromm, Michel

    2016-05-01

    In this study, we used a bench-top cold-cathode ultra-soft X-ray (USX) generator to expose aqueous DNA plasmid solutions to low-LET radiation under various scavenging conditions. Single- and double-strand breaks were assessed using classic gel electrophoresis quantification of linear, circular and supercoiled plasmid DNA topologies. With their very low penetration range in water, USX can only interact with matter up to short distances, of the order of 50 μm. We validated a stirring procedure which makes it possible to expose 100 µL of aqueous samples (2 mm thick). The scavenging of OH radicals by Tris buffer was studied at ambient temperature under aerobic conditions and compared to data gathered in the literature. A very good agreement was found with the rare data dealing with DNA plasmid exposed to Al Kα photons at low temperature (T ≤ 277 K), which therefore validated the experimental procedure. The yields for DNA single-strand breaks determined during this study enabled the ratio of indirect to direct effects to be determined at 96.2 %, in good agreement with the value of 97.7 % stemming from a study based on γ-ray irradiation of frozen solutions of plasmid DNA. Then, arginine was used both to create a "biological-like" chemical environment around the DNA plasmids and as an OH radical scavenger, in vitro. Although arginine has a greater scavenging (protecting) power than Tris, surprisingly, it led to higher rates of strand breakage. Based on the specific binding modes of arginine to DNA, we suggest that the side effects observed are due to the presence of arginine near to, but also inside, the DNA double helix. PMID:26994994

  4. Biochemical enrichment and biophysical characterization of a taste receptor for L-arginine from the catfish, Ictalurus puntatus

    Directory of Open Access Journals (Sweden)

    Spielman Andrew I

    2004-07-01

    Full Text Available Abstract Background The channel catfish, Ictalurus punctatus, is invested with a high density of cutaneous taste receptors, particularly on the barbel appendages. Many of these receptors are sensitive to selected amino acids, one of these being a receptor for L-arginine (L-Arg. Previous neurophysiological and biophysical studies suggested that this taste receptor is coupled directly to a cation channel and behaves as a ligand-gated ion channel receptor (LGICR. Earlier studies demonstrated that two lectins, Ricinus communis agglutinin I (RCA-I and Phaseolus vulgaris Erythroagglutinin (PHA-E, inhibited the binding of L-Arg to its presumed receptor sites, and that PHA-E inhibited the L-Arg-stimulated ion conductance of barbel membranes reconstituted into lipid bilayers. Results Both PHA-E and RCA-I almost exclusively labeled an 82–84 kDa protein band of an SDS-PAGE of solubilized barbel taste epithelial membranes. Further, both rhodamine-conjugated RCA-I and polyclonal antibodies raised to the 82–84 kDa electroeluted peptides labeled the apical region of catfish taste buds. Because of the specificity shown by RCA-I, lectin affinity was chosen as the first of a three-step procedure designed to enrich the presumed LGICR for L-Arg. Purified and CHAPS-solubilized taste epithelial membrane proteins were subjected successively to (1, lectin (RCA-I affinity; (2, gel filtration (Sephacryl S-300HR; and (3, ion exchange chromatography. All fractions from each chromatography step were evaluated for L-Arg-induced ion channel activity by reconstituting each fraction into a lipid bilayer. Active fractions demonstrated L-Arg-induced channel activity that was inhibited by D-arginine (D-Arg with kinetics nearly identical to those reported earlier for L-Arg-stimulated ion channels of native barbel membranes reconstituted into lipid bilayers. After the final enrichment step, SDS-PAGE of the active ion channel protein fraction revealed a single band at 82–84 k

  5. Docking of the Periplasmic FecB Binding Protein to the FecCD Transmembrane Proteins in the Ferric Citrate Transport System of Escherichia coli▿

    OpenAIRE

    Braun, Volkmar; Herrmann, Christina

    2007-01-01

    Citrate-mediated iron transport across the cytoplasmic membrane is catalyzed by an ABC transporter that consists of the periplasmic binding protein FecB, the transmembrane proteins FecC and FecD, and the ATPase FecE. Salt bridges between glutamate residues of the binding protein and arginine residues of the transmembrane proteins are predicted to mediate the positioning of the substrate-loaded binding protein on the transmembrane protein, based on the crystal structures of the ABC transporter...

  6. A reagentless electrochemiluminescent immunosensor for apurinic/apyrimidinic endonuclease 1 detection based on the new Ru(bpy)32+/bi-arginine system

    International Nuclear Information System (INIS)

    Highlights: • A reagentless ECL biosensor based on the new Ru(bpy)32+/bi-arginine system. • The successful preparation of bi-Arg/Au@Fe3O4–rGO as enhancer. • Using the APE-1 as target by the sandwich-type immunoassay format. - Abstract: Apurinic/apyrimidinic endonuclease 1 (APE-1), a kind of multifunctional protein widely-distributed in the body, plays an essential role in the DNA base excision repair and serves as multiple possible roles in the response of human cancer to radiotherapy and chemotherapy. In this work, an ultrasensitive solid-state electrochemiluminescence (ECL) immunosensor is designed to determine APE-1 based on the new Ru(bpy)32+/bi-arginine system. The bi-arginine (bi-Arg) is decorated on the Au nanoparticles functionalized magnetic Fe3O4/reduced graphene oxide (bi-Arg/Au@Fe3O4–rGO) according to the self-assembling and covalent cross-linking interaction to obtain the functionalized nanocomposite of bi-Arg/Au@Fe3O4–rGO. Herein, the bi-Arg/Au@Fe3O4–rGO plays not only an amplification label to enhance the ECL signal of Ru(bpy)32+ due to the coreactant of bi-Arg but also an ideal nanocarrier to load numerous secondary antibody. Based on sandwich-type immunoassay format, this proposed method offers a linear range of 1.0 fg mL−1–5.0 pg mL−1 and an estimated detection limit of 0.3 fg mL−1 for the APE-1. Moreover, the reagentless ECL immunosensor also exhibits high sensitivity, excellent selectivity and good stability, which has greatly potential development and application in clinical diagnostics, immunology and biomedical research

  7. ICE1 of Poncirus trifoliata functions in cold tolerance by modulating polyamine levels through interacting with arginine decarboxylase.

    Science.gov (United States)

    Huang, Xiao-San; Zhang, Qinghua; Zhu, Dexin; Fu, Xingzheng; Wang, Min; Zhang, Qian; Moriguchi, Takaya; Liu, Ji-Hong

    2015-06-01

    ICE1 (Inducer of CBF Expression 1) encodes a MYC-like basic helix-loop-helix transcription factor that acts as a central regulator of cold response. In this study, we elucidated the function and underlying mechanisms of PtrICE1 from trifoliate orange [Poncirus trifoliata (L.) Raf.]. PtrICE1 was upregulated by cold, dehydration, and salt, with the greatest induction under cold conditions. PtrICE1 was localized in the nucleus and could bind to a MYC-recognizing sequence. Ectopic expression of PtrICE1 in tobacco and lemon conferred enhanced tolerance to cold stresses at either chilling or freezing temperatures. Yeast two-hybrid screening revealed that 21 proteins belonged to the PtrICE1 interactome, in which PtADC (arginine decarboxylase) was confirmed as a bona fide protein interacting with PtrICE1. Transcript levels of ADC genes in the transgenic lines were slightly elevated under normal growth condition but substantially increased under cold conditions, consistent with changes in free polyamine levels. By contrast, accumulation of the reactive oxygen species, H2O2 and O2 (-), was appreciably alleviated in the transgenic lines under cold stress. Higher activities of antioxidant enzymes, such as superoxide dismutase and catalase, were detected in the transgenic lines under cold conditions. Taken together, these results demonstrated that PtrICE1 plays a positive role in cold tolerance, which may be due to modulation of polyamine levels through interacting with the ADC gene. PMID:25873670

  8. Presenting a foreign antigen on live attenuated Edwardsiella tarda using twin-arginine translocation signal peptide as a multivalent vaccine.

    Science.gov (United States)

    Wang, Yamin; Yang, Weizheng; Wang, Qiyao; Qu, Jiangbo; Zhang, Yuanxing

    2013-12-01

    The twin-arginine translocation (Tat) system is a major pathway for transmembrane translocation of fully folded proteins. In this study, a multivalent vaccine to present foreign antigens on live attenuated vaccine Edwardsiella tarda WED using screened Tat signal peptide was constructed. Because the Tat system increases the yields of folded antigens in periplasmic space or extracellular milieu, it is expected to contribute to the production of conformational epitope-derived specific antibodies. E. tarda Tat signal peptides fused with the green fluorescent protein (GFP) was constructed under the control of an in vivo inducible dps promoter. The resulting plasmids were electroporated into WED and the subcellular localizations of GFP were analyzed with Western blotting. Eight signal peptides with optimized GFP translocation efficiency were further fused to a protective antigen glyceraldehyde-3-phosphate dehydrogenase (GapA) from a fish pathogen Aeromonas hydrophila. Signal peptides of DmsA, NapA, and SufI displayed high efficiency for GapA translocation. The relative percent survival (RPS) of turbot was measured with a co-infection of E. tarda and A. hydrophila, and the strain with DmsA signal peptide showed the maximal protection. This study demonstrated a new platform to construct multivalent vaccines using optimized Tat signal peptide in E. tarda. PMID:23994481

  9. Diurnal changes in polyamine content, arginine and ornithine decarboxylase, and diamine oxidase in tobacco leaves

    Czech Academy of Sciences Publication Activity Database

    Gemperlová, Lenka; Nováková, Marie; Vaňková, Radomíra; Eder, Josef; Cvikrová, Milena

    2006-01-01

    Roč. 57, č. 6 (2006), s. 1413-1421. ISSN 0022-0957 R&D Projects: GA ČR GA206/03/0369 Institutional research plan: CEZ:AV0Z50380511 Keywords : Arginine decarboxylase * diamine oxidase * ornithine decarboxylase Subject RIV: ED - Physiology Impact factor: 3.630, year: 2006

  10. Impaired endothelial function after aneurysmal subarachnoid haemorrhage correlates with arginine:asymmetric dimethylarginine ratio

    DEFF Research Database (Denmark)

    Bergström, A; Staalsø, J M; Romner, B; Olsen, N V

    2014-01-01

    and arginine:ADMA ratio (r=0.43, P<0.005), but not with nitrite/nitrate, VMCA, or S-100B. CONCLUSIONS: Peripheral flow-mediated vasodilation is attenuated in the first days after SAH indicating acute systemic endothelial dysfunction. Impairment of endothelial function after SAH correlates with...

  11. Glutamine supplementation, citrulline production, and de novo arginine synthesis: Is there a relation?

    Science.gov (United States)

    We would like to comment on the recent publications by Buijs et al. The authors hypothesized that a parenteral supplement of glutamine stimulates citrulline formation and enhances de novo arginine synthesis. To test this hypothesis, they conducted an experiment with stable isotopes in patients under...

  12. Arginine-vasopressin stimulates the formation of phosphatidic acid in rat Leydig cells

    DEFF Research Database (Denmark)

    Nielsen, J.R.; Hansen, Harald S.; Jensen, B.

    1987-01-01

    Arginine-vasopressin (AVP) stimulated the formation of labelled phosphatidic acid (PA) in [C]arachidonic acid-prelabelled rat Leydig cells. After addition of 10 M AVP [C]arachidonoylphosphatidic acid reached a maximum within 2 min. The increase was dose-dependent (10-10 M). No change in labelling...

  13. Levels of arginine-vasopressin in cerebrospinal fluid during passive avoidance behavior in rats

    NARCIS (Netherlands)

    Kloet, E.R. de; Laczi, F.; Gaffori, O.; Fekete, M.; Wied, D. de

    1984-01-01

    The concentration of immunoreactive arginine-vasopressin (IR-AVP) was measured in the cerebrospinal fluid (CSF) during acquisition and retention of passive avoidance behavior. IR-AVP level in CSF of male Wistar rats immediately after the learning trial was increased; the rate of which was related to

  14. Reduced preabsorptive insulin response in aged rats : differential effects of amphetamine and arginine-vasopressin

    NARCIS (Netherlands)

    Buwalda, B.; Strubbe, J.H.; Bohus, B.

    1991-01-01

    The experiments presented here have been designed to investigate whether the age-related attenuation of the vagal reactivity to emotional stressors and its modulation by amphetamine (Amph) or arginine-vasopressin (AVP) can be generalized for other physiological response patterns. We therefore studie

  15. Evidence for a metabolic shift of arginine metabolism in sickle cell disease

    NARCIS (Netherlands)

    Schnog, JJB; Jager, EH; van der Dijs, FPL; Duits, AJ; Moshage, H; Muskiet, FD; Muskiet, FAJ

    2004-01-01

    Over the last few years, a pivotal role has been ascribed to reduced nitric oxide (NO) availability as a contributing factor to the vaso-occlusive process of sickle cell disease. We investigated whether arginine metabolism in sickle cell patients is different from healthy controls. Blood samples wer

  16. Watermelon enhances arginine availability in an animal model of type-II diabetes

    Science.gov (United States)

    Watermelon fruit contain lycopene, a red pigment known for its ability to scavenge free hydroxyl radicals. L-Citrulline, an amino acid that acts as a vasodilator and is a precursor of L-arginine, is found in all cucurbits, but is most plentiful in watermelon. In a study with Zucker diabetic fatty ...

  17. Synergistic Protection of L-Arginine and Vitamin E On Lipid Peroxidation of Asthenospermic Patients

    Directory of Open Access Journals (Sweden)

    Sudha Srivastava

    2008-01-01

    Full Text Available Background: Lipid peroxidation is known to cause various impairments to sperm cells and mayplay a major role in the etiology of male infertility. Asthenospermia is the main factor of maleinfertility and has significantly higher level of peroxidation than in normozoospermic males.Materials and Methods: Using thiobarbituric acid (TBA assay procedure, we have determinedthe level of lipid peroxidation as indicated by malondialdehyde (MDA in the spermatozoa obtainedfrom asthenospermic male semen.Results: An inverse correlation of MDA concentration with sperm motility is observed. Treatmentof cells with L-arginine and vitamin E significantly decreases the MDA concentration and improvesthe sperm motility as compared to that in case of control samples. A combination of L-arginine andvitamin E shows synergistic effect on sperm motility and prevention of lipid peroxidation.Conclusion: L-arginine and vitamin E protect the cells against the loss of sperm motility by lipidperoxidation. Therefore, supplementation of both L-arginine and vitamin E may improve spermmotility and increase the possibility of fertilization in asthenospermic subjects.

  18. Guanidinium Cations Pair with Positively Charged Arginine Side Chains in Water

    Czech Academy of Sciences Publication Activity Database

    Kubíčková, A.; Křížek, T.; Coufal, P.; Wernersson, Erik; Heyda, Jan; Jungwirth, Pavel

    2011-01-01

    Roč. 2, č. 12 (2011), s. 1387-1389. ISSN 1948-7185 R&D Projects: GA ČR GA203/08/0114; GA MŠk LC512 Institutional research plan: CEZ:AV0Z40550506 Keywords : guanidinium * cations * arginine * electrophoresis * water Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 6.213, year: 2011

  19. Effect of arginase inhibition on pulmonary L-arginine metabolism in murine Pseudomonas pneumonia.

    Directory of Open Access Journals (Sweden)

    Anne Mehl

    Full Text Available RATIONALE: Infection of the lung with Pseudomonas aeruginosa results in upregulation of nitric oxide synthases (NOS and arginase expression, and both enzymes compete for L-arginine as substrate. Nitric oxide (NO production may be regulated by arginase as it controls L-arginine availability for NOS. We here studied the effect of systemic arginase inhibition on pulmonary L-arginine metabolism in Pseudomonas pneumonia in the mouse. METHODS: Mice (C57BL/6, 8-10 weeks old, female underwent direct tracheal instillation of Pseudomonas (PAO-1-coated agar beads and were treated by repeated intra-peritoneal injections of the arginase inhibitor 2(S-amino-6-boronohexanoic acid (ABH or PBS until lungs were harvested on day 3 of the infection. L-arginine metabolites were quantified using liquid chromatography-tandem mass spectrometry, NO metabolites nitrate and nitrite by Griess reagent and cytokines by ELISA. RESULTS: NO metabolite concentrations (48.5±2.9 vs. 10.9±2.3 µM, p<0.0001, as well as L-ornithine (29.6±1.7 vs 2.3±0.4 µM, p<0.0001, the product of arginase activity, were increased in Pseudomonas infected lungs compared to naïve controls. Concentrations of the NOS inhibitor asymmetric dimethylarginine (ADMA were also increased (0.44±0.02 vs. 0.16±0.01 µM, p<0.0001. Arginase inhibition in the infected animals resulted in a significant decrease in L-ornithine (14.6±1.6 µM, p<0.0001 but increase in L-arginine concentration (p<0.001, L-arginine/ADMA ratio (p<0.001, L-arginine availability for NOS (p<0.001, and NO metabolite concentrations (67.3±5.7 µM, p<0.05. Arginase inhibitor treatment also resulted in an increase in NO metabolite levels in animals following intratracheal injection of LPS (p = 0.015. Arginase inhibition was not associated with an increase in inflammatory markers (IFN-γ, IL-1β, IL-6, MIP-2, KC or TNF-α in lung. Concentrations of the L-ornithine-dependent polyamines putrescine, spermidine and spermine were increased

  20. Tyrosine 87 is vital for the activity of human protein arginine methyltransferase 3 (PRMT3)

    Czech Academy of Sciences Publication Activity Database

    Handrková, H.; Petrák, J.; Halada, Petr; Pospíšilová, D.; Čmejla, R.

    2011-01-01

    Roč. 1814, č. 2 (2011), s. 277-282. ISSN 1570-9639 R&D Projects: GA MŠk LC07017 Institutional research plan: CEZ:AV0Z50200510 Keywords : DIAMOND-BLACKFAN ANEMIA * SUBSTRATE-SPECIFICITY * N-METHYLTRANSFERASE Subject RIV: CE - Biochemistry Impact factor: 3.635, year: 2011

  1. Co-operation between different targeting pathways during integration of a membrane protein

    OpenAIRE

    Keller, Rebecca; de Keyzer, Jeanine; Driessen, Arnold J. M.; Palmer, Tracy

    2012-01-01

    Membrane protein assembly is a fundamental process in all cells. The membrane-bound Rieske iron-sulfur protein is an essential component of the cytochrome bc(1) and cytochrome b(6)f complexes, and it is exported across the energy-coupling membranes of bacteria and plants in a folded conformation by the twin arginine protein transport pathway (Tat) transport pathway. Although the Rieske protein in most organisms is a monotopic membrane protein, in actinobacteria, it is a polytopic protein with...

  2. Nitric Oxide Synthase Inhibition by NG-Nitro-l-Arginine Methyl Ester Inhibits Tumor-Induced Angiogenesis in Mammary Tumors

    Science.gov (United States)

    Jadeski, Lorraine C.; Lala, Peeyush K.

    1999-01-01

    Using a murine breast cancer model, we earlier found a positive correlation between the expression of nitric oxide synthase (NOS) and tumor progression; treatment with inhibitors of NOS, NG-methyl-l-arginine (NMMA) and NG-nitro-l-arginine methyl ester (L-NAME), had antitumor and antimetastatic effects that were partly attributed to reduced tumor cell invasiveness. In the present study, we used a novel in vivo model of tumor angiogenesis using subcutaneous implants of tumor cells suspended in growth factor-reduced Matrigel to examine the angiogenic role of NO in a highly metastatic murine mammary adenocarcinoma cell line. This cell line, C3L5, expresses endothelial (e) NOS in vitro and in vivo, and inducible (i) NOS in vitro on stimulation with lipopolysaccharide and interferon-γ. Female C3H/HeJ mice received subcutaneous implants of growth factor-reduced Matrigel inclusive of C3L5 cells on one side, and on the contralateral side, Matrigel alone; L-NAME and D-NAME (inactive enantiomer) were subsequently administered for 14 days using osmotic minipumps. Immediately after sacrifice, implants were removed and processed for immunolocalization of eNOS and iNOS proteins, and measurement of angiogenesis. Neovascularization was quantified in sections stained with Masson’s trichrome or immunostained for the endothelial cell specific CD31 antigen. While most tumor cells and endothelial cells expressed immunoreactive eNOS protein, iNOS was localized in endothelial cells and some macrophages within the tumor-inclusive implants. Measurable angiogenesis occurred only in implants containing tumor cells. Irrespective of the method of quantification used, tumor-induced neovascularization was significantly reduced in L-NAME-treated mice relative to those treated with D-NAME. The quantity of stromal tissue was lower, but the quantity of necrotic tissue higher in L-NAME relative to D-NAME-treated animals. The total mass of viable tissue (ie, stroma and tumor cells) was lower in L

  3. A lysine-to-arginine mutation on NEDD8 markedly reduces the activity of cullin RING E3 ligase through the impairment of neddylation cascades

    International Nuclear Information System (INIS)

    Neural-precursor-cell-expressed developmentally down-regulated 8 (NEDD8) is a ubiquitin-like modifier, which forms covalent conjugates on lysines of its substrates. This post-translational modification, neddylation, plays important roles in tumor cell proliferation and viability. Ubiquitin can form diverse polyubiquitin chains, on its seven lysines, which play important functions in various biological processes. However, the roles of lysines in NEDD8 have not been explored. Here, we generated nine NEDD8 point mutants, each with one lysine replaced by an arginine, to study the putative function of lysines in NEDD8. Our experiments discover that Lys27 in NEDD8 is a critical residue for protein neddylation. Replacement of this residue with arginine almost completely eliminates the conjugation of NEDD8 to its substrates. Furthermore, we find that the K27R mutant impairs NEDD8 conjugation to the E2 enzyme, which normally forms thioester bonds for further transferring NEDD8 to its ligases and substrates. Therefore, this mutation completely inhibits global protein neddylation, including neddylation of cullin family proteins, resulting in decreased activity of cullin-RING E3 ligases. This work sheds new light on the roles of NEDD8 lysines on neddylation cascades and provides a dominant negative mutant for the study of neddylation and its biological functions. - Highlights: • Lys27 in NEDD8 is critical for protein neddylation. • NEDD8 K27R mutant impairs the NEDD8 conjugation. • NEDD8 K27R mutant significantly reduces the activity of cullin-RING E3 ligases

  4. A lysine-to-arginine mutation on NEDD8 markedly reduces the activity of cullin RING E3 ligase through the impairment of neddylation cascades

    Energy Technology Data Exchange (ETDEWEB)

    Sui, Yiyan; Liu, Yaobin; Xu, Guoqiang, E-mail: gux2002@suda.edu.cn

    2015-06-12

    Neural-precursor-cell-expressed developmentally down-regulated 8 (NEDD8) is a ubiquitin-like modifier, which forms covalent conjugates on lysines of its substrates. This post-translational modification, neddylation, plays important roles in tumor cell proliferation and viability. Ubiquitin can form diverse polyubiquitin chains, on its seven lysines, which play important functions in various biological processes. However, the roles of lysines in NEDD8 have not been explored. Here, we generated nine NEDD8 point mutants, each with one lysine replaced by an arginine, to study the putative function of lysines in NEDD8. Our experiments discover that Lys27 in NEDD8 is a critical residue for protein neddylation. Replacement of this residue with arginine almost completely eliminates the conjugation of NEDD8 to its substrates. Furthermore, we find that the K27R mutant impairs NEDD8 conjugation to the E2 enzyme, which normally forms thioester bonds for further transferring NEDD8 to its ligases and substrates. Therefore, this mutation completely inhibits global protein neddylation, including neddylation of cullin family proteins, resulting in decreased activity of cullin-RING E3 ligases. This work sheds new light on the roles of NEDD8 lysines on neddylation cascades and provides a dominant negative mutant for the study of neddylation and its biological functions. - Highlights: • Lys27 in NEDD8 is critical for protein neddylation. • NEDD8 K27R mutant impairs the NEDD8 conjugation. • NEDD8 K27R mutant significantly reduces the activity of cullin-RING E3 ligases.

  5. The effect of L-Arginine on the brain tissue of stressed rats

    Directory of Open Access Journals (Sweden)

    Batoul Ebadi

    2010-01-01

    Full Text Available   Abstract  Introduction: This study was conducted to determine the possible beneficial results of L-arginine on prefrontal cortex of rats which impressed by immobilization stress to define the synchronous impression of stress and nitric oxide (NO on evolution of prefrontal cortex of rats after birth. Methods: Forty-eight one month, male Wistar rats were divided into two groups: stressed and non-stressed. L-Arginine (200 mg/kg as a NO synthase (NOS inducer and L-NAME (2O mg/kg were injected intraperitonealy (IP and 7- nitroindazde (25 mg/kg as non-specific was injected subcutaneously (S.C. for 4 weeks. The kind of stress was immobilization for 4 weeks, every other day. The brain was removed after this period and each brain divided into two parts in a coronal section manner. Anterior part used for histological studies with H&E staining and posterior part used for measurement of NO production using spectrophotometer at 540 nm wavelengh. Results: Statistical analysis of microscopic and light microscopic finding showed that thickness of prefrontal cortex and NO production were significantly decreased in stressed rats and especially in groups which received 7- nitroindazole and L-NAME and L-arginine could reverse these results. Discussion: According to this research, we could say that L-arginine decreases the cortical damages in stressed rats and 7-nitroindazole and L-NAME increase this damage in non-stressed group. Although in non stressed groups, L-arginine, L-NAME and 7- nitroindazole were all non-protective and damaging.

  6. Glutamine, glutamate, and arginine-based acid resistance in Lactobacillus reuteri.

    Science.gov (United States)

    Teixeira, Januana S; Seeras, Arisha; Sanchez-Maldonado, Alma Fernanda; Zhang, Chonggang; Su, Marcia Shu-Wei; Gänzle, Michael G

    2014-09-01

    This study aimed to determine whether glutamine deamidation improves acid resistance of Lactobacillus reuteri, and to assess whether arginine, glutamine, and glutamate-mediated acid resistance are redundant or complementary mechanisms of acid resistance. Three putative glutaminase genes, gls1, gls2, and gls3, were identified in L. reuteri 100-23. All three genes were expressed during growth in mMRS and wheat sourdough. L. reuteri consistently over-expressed gls3 and the glutamate decarboxylase gadB. L. reuteri 100-23ΔgadB over-expressed gls3 and the arginine deiminase gene adi. Analysis of the survival of L. reuteri in acidic conditions revealed that arginine conversion is effective at pH of 3.5 while glutamine or glutamate conversion were effective at pH of 2.5. Arginine conversion increased the pHin but not ΔΨ; glutamate decarboxylation had only a minor effect on the pHin but increased the ΔΨ. This study demonstrates that glutamine deamidation increases the acid resistance of L. reuteri independent of glutamate decarboxylase activity. Arginine and glutamine/glutamate conversions confer resistance to lactate at pH of 3.5 and phosphate at pH of 2.5, respectively. Knowledge of L. reuteri's acid resistance improves the understanding of the adaptation of L. reuteri to intestinal ecosystems, and facilitates the selection of probiotic and starter cultures. PMID:24929734

  7. Genome-wide lentiviral shRNA screen identifies serine/arginine-rich splicing factor 2 as a determinant of oncolytic virus activity in breast cancer cells.

    Science.gov (United States)

    Workenhe, S T; Ketela, T; Moffat, J; Cuddington, B P; Mossman, K L

    2016-05-12

    Oncolytic human herpes simplex virus type 1 (HSV-1) shows promising treatment efficacy in late-stage clinical trials. The anticancer activity of oncolytic viruses relies on deregulated pathways in cancer cells, which make them permissive to oncolysis. To identify pathways that restrict HSV-1 KM100-mediated oncolysis, this study used a pooled genome-wide short hairpin RNA library and found that depletion of the splicing factor arginine-rich splicing factor 2 (SRSF2) leads to enhanced cytotoxicity of breast cancer cells by KM100. Serine/arginine-rich (SR) proteins are a family of RNA-binding phosphoproteins that control both constitutive and alternative pre-mRNA splicing. Further characterization showed that KM100 infection of HS578T cells under conditions of low SRSF2 leads to pronounced apoptosis without a corresponding increase in virus replication. As DNA topoisomerase I inhibitors can limit the phosphorylation of SRSF2, we combined a topoisomerase I inhibitor chemotherapeutic with KM100 and observed synergistic anticancer effect in vitro and prolonged survival of tumor-bearing mice in vivo. PMID:26257065

  8. Bioactive products of arginine in sepsis: tissue and plasma composition after LPS and iNOS blockade.

    Science.gov (United States)

    Lortie, M J; Ishizuka, S; Schwartz, D; Blantz, R C

    2000-06-01

    Blockade or gene deletion of inducible nitric oxide synthase (iNOS) fails to fully abrogate all the sequelae leading to the high morbidity of septicemia. An increase in substrate uptake may be necessary for the increased production of nitric oxide (NO), but arginine is also a precursor for other bioactive products. Herein, we demonstrate an increase in alternate arginine products via arginine and ornithine decarboxylase in rats given lipopolysaccharide (LPS). The expression of iNOS mRNA in renal tissue was evident 60 but not 30 min post-LPS, yet a rapid decrease in blood pressure was obtained within 30 min that was completely inhibited by selective iNOS blockade. Plasma levels of arginine and ornithine decreased by at least 30% within 60 min of LPS administration, an effect not inhibited by the iNOS blocker L-N(6)(1-iminoethyl)lysine (L-NIL). Significant increases in plasma nitrates and citrulline occurred only 3-4 h post-LPS, an effect blocked by L-NIL pretreatment. The intracellular composition of organs harvested 6 h post-LPS reflected tissue-specific profiles of arginine and related metabolites. Tissue arginine concentration, normally an order of magnitude higher than in plasma, did not decrease after LPS. Pretreatment with L-NIL had a significant impact on the disposition of tissue arginine that was organ specific. These data demonstrate changes in arginine metabolism before and after de novo iNOS activity. Selective blockade of iNOS did not prevent uptake and can deregulate the production of other bioactive arginine metabolites. PMID:10837347

  9. The structure and IR signatures of the arginine-glutamate salt bridge. Insights from the classical MD simulations

    Energy Technology Data Exchange (ETDEWEB)

    Vener, M. V., E-mail: mikhail.vener@gmail.com [Mendeleev University of Chemical Technology, Moscow (Russian Federation); Odinokov, A. V. [Photochemistry Center of the Russian Academy of Sciences, Moscow (Russian Federation); Wehmeyer, C. [Free University, Berlin (Germany); Sebastiani, D. [Martin-Luther-Universität Halle-Wittenberg, Halle (Germany)

    2015-06-07

    Salt bridges and ionic interactions play an important role in protein stability, protein-protein interactions, and protein folding. Here, we provide the classical MD simulations of the structure and IR signatures of the arginine (Arg)–glutamate (Glu) salt bridge. The Arg-Glu model is based on the infinite polyalanine antiparallel two-stranded β-sheet structure. The 1 μs NPT simulations show that it preferably exists as a salt bridge (a contact ion pair). Bidentate (the end-on and side-on structures) and monodentate (the backside structure) configurations are localized [Donald et al., Proteins 79, 898–915 (2011)]. These structures are stabilized by the short {sup +}N–H⋯O{sup −} bonds. Their relative stability depends on a force field used in the MD simulations. The side-on structure is the most stable in terms of the OPLS-AA force field. If AMBER ff99SB-ILDN is used, the backside structure is the most stable. Compared with experimental data, simulations using the OPLS all-atom (OPLS-AA) force field describe the stability of the salt bridge structures quite realistically. It decreases in the following order: side-on > end-on > backside. The most stable side-on structure lives several nanoseconds. The less stable backside structure exists a few tenth of a nanosecond. Several short-living species (solvent shared, completely separately solvated ionic groups ion pairs, etc.) are also localized. Their lifetime is a few tens of picoseconds or less. Conformational flexibility of amino acids forming the salt bridge is investigated. The spectral signature of the Arg-Glu salt bridge is the IR-intensive band around 2200 cm{sup −1}. It is caused by the asymmetric stretching vibrations of the {sup +}N–H⋯O{sup −} fragment. Result of the present paper suggests that infrared spectroscopy in the 2000–2800 frequency region may be a rapid and quantitative method for the study of salt bridges in peptides and ionic interactions between proteins. This region is

  10. The structure and IR signatures of the arginine-glutamate salt bridge. Insights from the classical MD simulations

    International Nuclear Information System (INIS)

    Salt bridges and ionic interactions play an important role in protein stability, protein-protein interactions, and protein folding. Here, we provide the classical MD simulations of the structure and IR signatures of the arginine (Arg)–glutamate (Glu) salt bridge. The Arg-Glu model is based on the infinite polyalanine antiparallel two-stranded β-sheet structure. The 1 μs NPT simulations show that it preferably exists as a salt bridge (a contact ion pair). Bidentate (the end-on and side-on structures) and monodentate (the backside structure) configurations are localized [Donald et al., Proteins 79, 898–915 (2011)]. These structures are stabilized by the short +N–H⋯O− bonds. Their relative stability depends on a force field used in the MD simulations. The side-on structure is the most stable in terms of the OPLS-AA force field. If AMBER ff99SB-ILDN is used, the backside structure is the most stable. Compared with experimental data, simulations using the OPLS all-atom (OPLS-AA) force field describe the stability of the salt bridge structures quite realistically. It decreases in the following order: side-on > end-on > backside. The most stable side-on structure lives several nanoseconds. The less stable backside structure exists a few tenth of a nanosecond. Several short-living species (solvent shared, completely separately solvated ionic groups ion pairs, etc.) are also localized. Their lifetime is a few tens of picoseconds or less. Conformational flexibility of amino acids forming the salt bridge is investigated. The spectral signature of the Arg-Glu salt bridge is the IR-intensive band around 2200 cm−1. It is caused by the asymmetric stretching vibrations of the +N–H⋯O− fragment. Result of the present paper suggests that infrared spectroscopy in the 2000–2800 frequency region may be a rapid and quantitative method for the study of salt bridges in peptides and ionic interactions between proteins. This region is usually not considered in

  11. Arginine-based cationic liposomes for efficient in vitro plasmid DNA delivery with low cytotoxicity

    Directory of Open Access Journals (Sweden)

    Sarker SR

    2013-04-01

    Full Text Available Satya Ranjan Sarker, Yumiko Aoshima, Ryosuke Hokama, Takafumi Inoue, Keitaro Sou, Shinji Takeoka Department of Life Science and Medical Bioscience, Graduate School of Advanced Science and Engineering, Waseda University (TWIns, Tokyo, Japan Background: Currently available gene delivery vehicles have many limitations such as low gene delivery efficiency and high cytotoxicity. To overcome these drawbacks, we designed and synthesized two cationic lipids comprised of n-tetradecyl alcohol as the hydrophobic moiety, 3-hydrocarbon chain as the spacer, and different counterions (eg, hydrogen chloride [HCl] salt or trifluoroacetic acid [TFA] salt in the arginine head group. Methods: Cationic lipids were hydrated in 4-(2-hydroxyethyl-1-piperazineethanesulfonic acid (HEPES buffer to prepare cationic liposomes and characterized in terms of their size, zeta potential, phase transition temperature, and morphology. Lipoplexes were then prepared and characterized in terms of their size and zeta potential in the absence or presence of serum. The morphology of the lipoplexes was determined using transmission electron microscopy and atomic force microscopy. The gene delivery efficiency was evaluated in neuronal cells and HeLa cells and compared with that of lysine-based cationic assemblies and Lipofectamine™ 2000. The cytotoxicity level of the cationic lipids was investigated and compared with that of Lipofectamine™ 2000. Results: We synthesized arginine-based cationic lipids having different counterions (ie, HCl-salt or TFA-salt that formed cationic liposomes of around 100 nm in size. In the absence of serum, lipoplexes prepared from the arginine-based cationic liposomes and plasmid (p DNA formed large aggregates and attained a positive zeta potential. However, in the presence of serum, the lipoplexes were smaller in size and negative in zeta potential. The morphology of the lipoplexes was vesicular. Arginine-based cationic liposomes with HCl-salt showed the

  12. Insight on an arginine synthesis metabolon from the tetrameric structure of yeast acetylglutamate kinase.

    Directory of Open Access Journals (Sweden)

    Sergio de Cima

    Full Text Available N-acetyl-L-glutamate kinase (NAGK catalyzes the second, generally controlling, step of arginine biosynthesis. In yeasts, NAGK exists either alone or forming a metabolon with N-acetyl-L-glutamate synthase (NAGS, which catalyzes the first step and exists only within the metabolon. Yeast NAGK (yNAGK has, in addition to the amino acid kinase (AAK domain found in other NAGKs, a ~150-residue C-terminal domain of unclear significance belonging to the DUF619 domain family. We deleted this domain, proving that it stabilizes yNAGK, slows catalysis and modulates feed-back inhibition by arginine. We determined the crystal structures of both the DUF619 domain-lacking yNAGK, ligand-free as well as complexed with acetylglutamate or acetylglutamate and arginine, and of complete mature yNAGK. While all other known arginine-inhibitable NAGKs are doughnut-like hexameric trimers of dimers of AAK domains, yNAGK has as central structure a flat tetramer formed by two dimers of AAK domains. These dimers differ from canonical AAK dimers in the -110° rotation of one subunit with respect to the other. In the hexameric enzymes, an N-terminal extension, found in all arginine-inhibitable NAGKs, forms a protruding helix that interlaces the dimers. In yNAGK, however, it conforms a two-helix platform that mediates interdimeric interactions. Arginine appears to freeze an open inactive AAK domain conformation. In the complete yNAGK structure, two pairs of DUF619 domains flank the AAK domain tetramer, providing a mechanism for the DUF619 domain modulatory functions. The DUF619 domain exhibits the histone acetyltransferase fold, resembling the catalytic domain of bacterial NAGS. However, the putative acetyl CoA site is blocked, explaining the lack of NAGS activity of yNAGK. We conclude that the tetrameric architecture is an adaptation to metabolon formation and propose an organization for this metabolon, suggesting that yNAGK may be a good model also for yeast and human NAGSs.

  13. Insight on an arginine synthesis metabolon from the tetrameric structure of yeast acetylglutamate kinase.

    Science.gov (United States)

    de Cima, Sergio; Gil-Ortiz, Fernando; Crabeel, Marjolaine; Fita, Ignacio; Rubio, Vicente

    2012-01-01

    N-acetyl-L-glutamate kinase (NAGK) catalyzes the second, generally controlling, step of arginine biosynthesis. In yeasts, NAGK exists either alone or forming a metabolon with N-acetyl-L-glutamate synthase (NAGS), which catalyzes the first step and exists only within the metabolon. Yeast NAGK (yNAGK) has, in addition to the amino acid kinase (AAK) domain found in other NAGKs, a ~150-residue C-terminal domain of unclear significance belonging to the DUF619 domain family. We deleted this domain, proving that it stabilizes yNAGK, slows catalysis and modulates feed-back inhibition by arginine. We determined the crystal structures of both the DUF619 domain-lacking yNAGK, ligand-free as well as complexed with acetylglutamate or acetylglutamate and arginine, and of complete mature yNAGK. While all other known arginine-inhibitable NAGKs are doughnut-like hexameric trimers of dimers of AAK domains, yNAGK has as central structure a flat tetramer formed by two dimers of AAK domains. These dimers differ from canonical AAK dimers in the -110° rotation of one subunit with respect to the other. In the hexameric enzymes, an N-terminal extension, found in all arginine-inhibitable NAGKs, forms a protruding helix that interlaces the dimers. In yNAGK, however, it conforms a two-helix platform that mediates interdimeric interactions. Arginine appears to freeze an open inactive AAK domain conformation. In the complete yNAGK structure, two pairs of DUF619 domains flank the AAK domain tetramer, providing a mechanism for the DUF619 domain modulatory functions. The DUF619 domain exhibits the histone acetyltransferase fold, resembling the catalytic domain of bacterial NAGS. However, the putative acetyl CoA site is blocked, explaining the lack of NAGS activity of yNAGK. We conclude that the tetrameric architecture is an adaptation to metabolon formation and propose an organization for this metabolon, suggesting that yNAGK may be a good model also for yeast and human NAGSs. PMID:22529931

  14. Improved Muscle Function in Duchenne Muscular Dystrophy through L-Arginine and Metformin: An Investigator-Initiated, Open-Label, Single-Center, Proof-Of-Concept-Study.

    Directory of Open Access Journals (Sweden)

    Patricia Hafner

    Full Text Available Altered neuronal nitric oxide synthase function in Duchenne muscular dystrophy leads to impaired mitochondrial function which is thought to be one cause of muscle damage in this disease. The study tested if increased intramuscular nitric oxide concentration can improve mitochondrial energy metabolism in Duchenne muscular dystrophy using a novel therapeutic approach through the combination of L-arginine with metformin. Five ambulatory, genetically confirmed Duchenne muscular dystrophy patients aged between 7–10 years were treated with L-arginine (3 x 2.5 g/d and metformin (2 x 250 mg/d for 16 weeks. Treatment effects were assessed using mitochondrial protein expression analysis in muscular biopsies, indirect calorimetry, Dual-Energy X-Ray Absorptiometry, quantitative thigh muscle MRI, and clinical scores of muscle performance. There were no serious side effects and no patient dropped out. Muscle biopsy results showed pre-treatment a significantly reduced mitochondrial protein expression and increased oxidative stress in Duchenne muscular dystrophy patients compared to controls. Post-treatment a significant elevation of proteins of the mitochondrial electron transport chain was observed as well as a reduction in oxidative stress. Treatment also decreased resting energy expenditure rates and energy substrate use shifted from carbohydrates to fatty acids. These changes were associated with improved clinical scores. In conclusion pharmacological stimulation of the nitric oxide pathway leads to improved mitochondria function and clinically a slowing of disease progression in Duchenne muscular dystrophy. This study shall lead to further development of this novel therapeutic approach into a real alternative for Duchenne muscular dystrophy patients.ClinicalTrials.gov NCT02516085.

  15. Double Strand Break Unwinding and Resection by the Mycobacterial Helicase-Nuclease AdnAB in the Presence of Single Strand DNA-binding Protein (SSB)*

    OpenAIRE

    Unciuleac, Mihaela-Carmen; Shuman, Stewart

    2010-01-01

    Mycobacterial AdnAB is a heterodimeric DNA helicase-nuclease and 3′ to 5′ DNA translocase implicated in the repair of double strand breaks (DSBs). The AdnA and AdnB subunits are each composed of an N-terminal motor domain and a C-terminal nuclease domain. Inclusion of mycobacterial single strand DNA-binding protein (SSB) in reactions containing linear plasmid dsDNA allowed us to study the AdnAB helicase under conditions in which the unwound single strands are coated by SSB and thereby prevent...

  16. Dysregulated expression of arginine metabolic enzymes in human intestinal tissues of necrotizing enterocolitis and response of CaCO2 cells to bacterial components.

    Science.gov (United States)

    Leung, Kam Tong; Chan, Kathy Yuen Yee; Ma, Terence Ping Yuen; Yu, Jasmine Wai Sum; Tong, Joanna Hung Man; Tam, Yuk Him; Cheung, Hon Ming; To, Ka Fai; Lam, Hugh Simon; Lee, Kim Hung; Li, Karen; Ng, Pak Cheung

    2016-03-01

    The small intestine is the exclusive site of arginine synthesis in neonates. Low levels of circulating arginine have been associated with the occurrence of necrotizing enterocolitis (NEC) but the mechanism of arginine dysregulation has not been fully elucidated. We aimed to investigate (i) expressional changes of arginine synthesizing and catabolic enzymes in human intestinal tissues of NEC, spontaneous intestinal perforation (SIP) and noninflammatory surgical conditions (Surg-CTL) and to investigate the (ii) mechanisms of arginine dysregulation and enterocyte proliferation upon stimulation by bacterial components, arginine depletion, ARG1 overexpression and nitric oxide (NO) supplementation. Our results showed that expressions of arginine synthesizing enzymes ALDH18A1, ASL, ASS1, CPS1, GLS, OAT and PRODH were significantly decreased in NEC compared with Surg-CTL or SIP tissues. Catabolic enzyme ARG1 was increased (>100-fold) in NEC tissues and histologically demonstrated to be expressed by infiltrating neutrophils. No change in arginine metabolic enzymes was observed between SIP and Surg-CTL tissues. In CaCO2 cells, arginine metabolic enzymes were differentially dysregulated by lipopolysaccharide or lipoteichoic acid. Depletion of arginine reduced cell proliferation and this phenomenon could be partially rescued by NO. Overexpression of ARG1 also reduced enterocyte proliferation. We provided the first expressional profile of arginine metabolic enzymes at the tissue level of NEC. Our findings suggested that arginine homeostasis was severely disturbed and could be triggered by inflammatory responses of enterocytes and infiltrating neutrophils as well as bacterial components. Such reactions could reduce arginine and NO, resulting in mucosal damage. The benefit of arginine supplementation for NEC prophylaxis merits further clinical evaluation. PMID:26895666

  17. Nanoscale characterization of effect of L-arginine on Streptococcus mutans biofilm adhesion by atomic force microscopy.

    Science.gov (United States)

    Sharma, Shivani; Lavender, Stacey; Woo, JungReem; Guo, Lihong; Shi, Wenyuan; Kilpatrick-Liverman, LaTonya; Gimzewski, James K

    2014-07-01

    A major aetiological factor of dental caries is the pathology of the dental plaque biofilms. The amino acid L-arginine (Arg) is found naturally in saliva as a free molecule or as a part of salivary peptides and proteins. Plaque bacteria metabolize Arg to produce alkali and neutralize glycolytic acids, promoting a less cariogenous oral microbiome. Here, we explored an alternative and complementary mechanism of action of Arg using atomic force microscopy. The nanomechanical properties of Streptococcus mutans biofilm extracellular matrix were characterized under physiological buffer conditions. We report the effect of Arg on the adhesive behaviour and structural properties of extracellular polysaccharides in S. mutans biofilms. High-resolution imaging of biofilm surfaces can reveal additional structural information on bacterial cells embedded within the surrounding extracellular matrix. A dense extracellular matrix was observed in biofilms without Arg compared to those grown in the presence of Arg. S. mutans biofilms grown in the presence of Arg could influence the production and/or composition of extracellular membrane glucans and thereby affect their adhesion properties. Our results suggest that the presence of Arg in the oral cavity could influence the adhesion properties of S. mutans to the tooth surface. PMID:24763427

  18. Conductometric and volumetric studies of atorvastatin in aqueous solution of arginine from 298.15 to 313.15 K

    Directory of Open Access Journals (Sweden)

    M. M. R. Meor Mohd Affandi

    2016-01-01

    Full Text Available Categorized as a Biopharmaceutics Classification System Class II drugs, atorvastatin (ATV exhibits low aqueous solubility and bioavailability thus presenting an obstacle and great challenge to formulation researchers. Numerous studies are available in regard to the solubility enhancement of ATV, but very few actually describe this phenomenon in terms of thermodynamics and the solute-solvent interaction. Arginine (ARG is an amino acid that has been reported to enhance the solubility of the highly insoluble wheat protein gluten through hydrogen bonding and π electron-cation interaction. To our knowledge, ARG has never been investigated as a solubility enhancement agent of aqueous insoluble drugs. Thus, this study aimed to elucidate the solute-solvent and solute-cosolute interactions and derive thermodynamic parameters that bolstered the solubility of ATV in the presence of ARG. We examined the electrolytic conductance and densities of ATV-ARG binary system covering the temperature ranging from 298.15 K to 313.15 K. Conductometric and volumetric parameters such as limiting molar conductance, association constants, limiting partial molar volumes, and expansibility values were calculated. Additionally, thermodynamic parameters (ΔG0, ΔH0, ΔS0, and Es involved in the association process of the solute in the aqueous solution of ARG were also determined.

  19. Abrogation of the twin arginine transport system in Salmonella enterica serovar Typhimurium leads to colonization defects during infection.

    Science.gov (United States)

    Reynolds, M Megan; Bogomolnaya, Lydia; Guo, Jinbai; Aldrich, Lindsay; Bokhari, Danial; Santiviago, Carlos A; McClelland, Michael; Andrews-Polymenis, Helene

    2011-01-01

    TatC (STM3975) is a highly conserved component of the Twin Arginine Transport (Tat) systems that is required for transport of folded proteins across the inner membrane in gram-negative bacteria. We previously identified a ΔtatC mutant as defective in competitive infections with wild type ATCC14028 during systemic infection of Salmonella-susceptible BALB/c mice. Here we confirm these results and show that the ΔtatC mutant is internalized poorly by cultured J774-A.1 mouse macrophages a phenotype that may be related to the systemic infection defect. This mutant is also defective for short-term intestinal and systemic colonization after oral infection of BALB/c mice and is shed in reduced numbers in feces from orally infected Salmonella-resistant (CBA/J) mice. We show that the ΔtatC mutant is highly sensitive to bile acids perhaps resulting in the defect in intestinal infection that we observe. Finally, the ΔtatC mutant has an unusual combination of motility phenotypes in Salmonella; it is severely defective for swimming motility but is able to swarm well. The ΔtatC mutant has a lower amount of flagellin on the bacterial surface during swimming motility but normal levels under swarming conditions. PMID:21298091

  20. Topology independent protein structural alignment

    Directory of Open Access Journals (Sweden)

    DasGupta Bhaskar

    2007-10-01

    Full Text Available Abstract Background Identifying structurally similar proteins with different chain topologies can aid studies in homology modeling, protein folding, protein design, and protein evolution. These include circular permuted protein structures, and the more general cases of non-cyclic permutations between similar structures, which are related by non-topological rearrangement beyond circular permutation. We present a method based on an approximation algorithm that finds sequence-order independent structural alignments that are close to optimal. We formulate the structural alignment problem as a special case of the maximum-weight independent set problem, and solve this computationally intensive problem approximately by iteratively solving relaxations of a corresponding integer programming problem. The resulting structural alignment is sequence order independent. Our method is also insensitive to insertions, deletions, and gaps. Results Using a novel similarity score and a statistical model for significance p-value, we are able to discover previously unknown circular permuted proteins between nucleoplasmin-core protein and auxin binding protein, between aspartate rasemase and 3-dehydrogenate dehydralase, as well as between migration inhibition factor and arginine repressor which involves an additional strand-swapping. We also report the finding of non-cyclic permuted protein structures existing in nature between AML1/core binding factor and ribofalvin synthase. Our method can be used for large scale alignment of protein structures regardless of the topology. Conclusion The approximation algorithm introduced in this work can find good solutions for the problem of protein structure alignment. Furthermore, this algorithm can detect topological differences between two spatially similar protein structures. The alignment between MIF and the arginine repressor demonstrates our algorithm's ability to detect structural similarities even when spatial

  1. Amino acids composition and protein quality evaluation of marine species and meals for feed formulations in cephalopods

    OpenAIRE

    Cerezo Valverde, Jesús; Martínez-Llorens, Silvia; TOMÁS VIDAL, ANA; Jover Cerda, Miguel; Rodriguez, Carmen; Estefanell, Juan; Gairin, Joan I.; Domingues, Pedro Miguel; Rodriguez, Carlos J.; Garcia Garcia, Benjamin

    2013-01-01

    The amino acid composition and protein levels of three species of cephalopods (Octopus vulgaris, Loligo gahi and Todarodes sagittatus), the natural diets of common octopus (O. vulgaris) and different kinds of meals were determined in order to optimise the content of these nutrients in artificial feeds. Arginine, leucine and lysine were the most abundant essential amino acids in cephalopods, while glutamate and aspartate represented the main non-essential amino acids. Arginine and leucine were...

  2. Reconstruction and modeling protein translocation and compartmentalization in Escherichia coli at the genome-scale

    DEFF Research Database (Denmark)

    Liu, Joanne K.; O’Brien, Edward J.; Lerman, Joshua A.;

    2014-01-01

    Background: Membranes play a crucial role in cellular functions. Membranes provide a physical barrier, control the trafficking of substances entering and leaving the cell, and are a major determinant of cellular ultra-structure. In addition, components embedded within the membrane participate in...... into genome-scale models. Results: The recent genome-scale model of metabolism and protein expression in Escherichia coli (called a ME-model) computes the complete composition of the proteome required to perform whole cell functions. Here we expand the ME-model to include (1) a reconstruction of...... protein translocation pathways, (2) assignment of all cellular proteins to one of four compartments (cytoplasm, inner membrane, periplasm, and outer membrane) and a translocation pathway, (3) experimentally determined translocase catalytic and porin diffusion rates, and (4) a novel membrane constraint...

  3. High-performance liquid chromatography method with radiochemical detection for measurement of nitric oxide synthase, arginase, and arginine decarboxylase activities

    DEFF Research Database (Denmark)

    Volke, A; Wegener, Gregers; Vasar, E;

    2006-01-01

    Nitric oxide has been shown to be involved in numerous biological processes, and many studies have aimed to measure nitric oxide synthase (NOS) activity. Recently, it has been demonstrated that arginase and arginine decarboxylase (ADC), two enzymes that also employ arginine as a substrate, may...... simple and fast HPLC method with radiochemical detection to separate radiolabeled L-arginine, L-citrulline, L-ornithine, and agmatine. 3H-labeled L-arginine, L-citrulline, agmatine, and 14C-labeled L-citrulline were used as standards. These compounds were separated in the normal phase column (Allure...... Acidix 250 x 4.6 mm i.d.) under isocratic conditions in less than 20 min with good sensitivity. Using the current method, we have shown the formation of L-citrulline and L-ornithine in vitro using brain tissue homogenate of rats and that of agmatine by Escherichia coli ADC. Udgivelsesdato: null-null...

  4. Effects of a food supplement rich in arginine in patients with smear positive pulmonary tuberculosis--a randomised trial

    DEFF Research Database (Denmark)

    Schön, T; Idh, J; Westman, A;

    2011-01-01

    In tuberculosis (TB), the production of nitric oxide (NO) is confirmed but its importance in host defense is debated. Our aim was to investigate whether a food supplement rich in arginine could enhance clinical improvement in TB patients by increased NO production. Smear positive TB patients from...... Gondar, Ethiopia (n = 180) were randomized to a food supplementation rich in arginine (peanuts, equivalent to 1 g of arginine/day) or with a low arginine content (wheat crackers, locally called daboqolo) during four weeks. The primary outcome was cure rate according to the WHO classification and...... secondary outcomes. In the subgroup analysis according to HIV status, peanut supplemented HIV+/TB patients showed increased cure rate (83.8% (31/37) vs 53.1% (17/32), p <0.01). A low baseline eNO (...

  5. One-Pot Green Synthesis and Bioapplication of l-Arginine-Capped Superparamagnetic Fe3O4 Nanoparticles

    Science.gov (United States)

    Lai, Yongchao; Yin, Weiwei; Liu, Jinting; Xi, Rimo; Zhan, Jinhua

    2010-02-01

    Water-soluble l-arginine-capped Fe3O4 nanoparticles were synthesized using a one-pot and green method. Nontoxic, renewable and inexpensive reagents including FeCl3, l-arginine, glycerol and water were chosen as raw materials. Fe3O4 nanoparticles show different dispersive states in acidic and alkaline solutions for the two distinct forms of surface binding l-arginine. Powder X-ray diffraction and X-ray photoelectron spectroscopy were used to identify the structure of Fe3O4 nanocrystals. The products behave like superparamagnetism at room temperature with saturation magnetization of 49.9 emu g-1 and negligible remanence or coercivity. In the presence of 1-ethyl-3-(dimethylaminopropyl) carbodiimide hydrochloride, the anti-chloramphenicol monoclonal antibodies were connected to the l-arginine-capped magnetite nanoparticles. The as-prepared conjugates could be used in immunomagnetic assay.

  6. Oral L-Arginine Stimulates GLP-1 Secretion to Improve Glucose Tolerance in Male Mice

    DEFF Research Database (Denmark)

    Clemmensen, Christoffer; Smajilovic, Sanela; Smith, Eric P;

    2013-01-01

    Pharmacological and surgical interventions that increase glucagon-like peptide 1 (GLP-1) action are effective to improve glucose homeostasis in type 2 diabetes mellitus. In light of this, nutritional strategies to enhance postprandial GLP-1 secretion, particularly in the context of diet-induced o......Pharmacological and surgical interventions that increase glucagon-like peptide 1 (GLP-1) action are effective to improve glucose homeostasis in type 2 diabetes mellitus. In light of this, nutritional strategies to enhance postprandial GLP-1 secretion, particularly in the context of diet......-induced obesity, may provide an alternative therapeutic approach. Importantly, recent evidence suggests the amino acid l-arginine, a well-known insulin secretagogue, can also stimulate release of GLP-1 from isolated rat intestine. Here we tested the hypothesis that oral l-arginine acts as a GLP-1 secretagogue in...

  7. Atomic-Resolution Structure of an N(5) Flavin Adduct in D-Arginine Dehydrogenase

    Energy Technology Data Exchange (ETDEWEB)

    Fu, Guoxing; Yuan, Hongling; Wang, Siming; Gadda, Giovanni; Weber, Irene T. (GSU)

    2011-09-06

    D-Arginine dehydrogenase (DADH) catalyzes the flavin-dependent oxidative deamination of D-arginine and other D-amino acids to the corresponding imino acids. The 1.07 {angstrom} atomic-resolution structure of DADH crystallized with D-leucine unexpectedly revealed a covalent N(5) flavin adduct, instead of the expected iminoleucine product in the active site. This acyl adduct has been successfully reproduced by photoreduction of DADH in the presence of 4-methyl-2-oxopentanoic acid (ketoleucine). The iminoleucine may be released readily because of weak interactions in the binding site, in contrast to iminoarginine, converted to ketoleucine, which reacts with activated FAD to form the covalently linked acyl adduct.

  8. Atomic-resolution structure of an N5 flavin adduct in D-arginine dehydrogenase.

    Science.gov (United States)

    Fu, Guoxing; Yuan, Hongling; Wang, Siming; Gadda, Giovanni; Weber, Irene T

    2011-07-26

    D-Arginine dehydrogenase (DADH) catalyzes the flavin-dependent oxidative deamination of D-arginine and other D-amino acids to the corresponding imino acids. The 1.07 Å atomic-resolution structure of DADH crystallized with D-leucine unexpectedly revealed a covalent N(5) flavin adduct, instead of the expected iminoleucine product in the active site. This acyl adduct has been successfully reproduced by photoreduction of DADH in the presence of 4-methyl-2-oxopentanoic acid (ketoleucine). The iminoleucine may be released readily because of weak interactions in the binding site, in contrast to iminoarginine, converted to ketoleucine, which reacts with activated FAD to form the covalently linked acyl adduct. PMID:21707047

  9. Arginine-assisted synthesis and catalytic properties of single-crystalline palladium tetrapods.

    Science.gov (United States)

    Fu, Geng-Tao; Jiang, Xian; Wu, Rui; Wei, Shao-Hua; Sun, Dong-Mei; Tang, Ya-Wen; Lu, Tian-Hong; Chen, Yu

    2014-12-24

    Noble metallic nanocrystals (NMNCs) with highly branched morphologies are an exciting new class of nanomaterials because of their great potential application in catalysis, sensing, optics, and electronics originating from their unique structures. Herein, we report a facile water-based method to synthesize high-quality palladium (Pd) tetrapods with the assistance of arginine molecule, which is more economical and environmentally friendly than the previous reported carbon monoxide (CO)-assisted synthesis in the organic system. During the synthesis, arginine molecule plays an essential role in controlling the tetrapod-like morphology. The as-synthesized Pd tetrapods have a potential application in the formic acid (HCOOH)-induced reduction of highly toxic hexavalent chromium (Cr(VI)) owing to their improved catalytic performance for the HCOOH decomposition. PMID:25469763

  10. Solid-state properties and dissolution behaviour of tablets containing co-amorphous indomethacin-arginine

    DEFF Research Database (Denmark)

    Lenz, Elisabeth; Jensen, Katrine Birgitte Tarp; Blaabjerg, Lasse Ingerslev;

    2015-01-01

    Co-amorphous drug formulations provide the possibility to stabilize a drug in its amorphous form by interactions with low molecular weight compounds, e.g. amino acids. Recent studies have shown the feasibility of spray drying as a technique to manufacture co-amorphous indomethacin–arginine in a...... larger production scale. In this work, a tablet formulation was developed for a co-amorphous salt, namely spray dried indomethacin–arginine (SD IND–ARG). The effects of compaction pressure on tablet properties, physical stability and dissolution profiles under non-sink conditions were examined....... Dissolution profiles of tablets with SD IND–ARG (TAB SD IND–ARG) were compared to those of tablets containing a physical mixture of crystalline IND and ARG (TAB PM IND–ARG) and to the dissolution of pure spray dried powder. Concerning tableting, the developed formulation allowed for the preparation of tablets...

  11. Arginine Vasopressin V1a Receptor Antagonist Impairs Maternal Memory in Rats

    OpenAIRE

    Nephew, Benjamin C.; Bridges, Robert S.

    2008-01-01

    Primiparous female rats rapidly respond to foster pups following an extended separation from pups after an initial maternal experience. This consolidation of maternal behavior has been referred to as maternal memory. The neurochemical regulation of maternal memory is not clearly understood. One neuropeptide that may mediate maternal memory is arginine vasopressin (AVP), a neuropeptide which is modulated around the time of parturition and has an established role in learning and memory processe...

  12. Comparison of L-Arginine and Hydroxyurea Interactractions with Transitional Metal Ions.

    Directory of Open Access Journals (Sweden)

    Sangeeta Parab

    2015-11-01

    Full Text Available No* is a free radical with one free electron and as such it is very highly reactive and particularly it interacts with transitional metals. Nitric oxide, gas is an important signaling molecule in the body of mammals, including humans and is an extremely important intermediate in chemical industry In biological systems there are many enzymes, which contain transitional elements like iron, copper and manganese, which are the most probable sites for nitric oxide to react. Such type of interactions results in considerable mod ification of the enzyme functions resulting pathological and even genetic disorders. This needs a critical amount of nitric oxide in the system for proper functioning. To observe the effects of NO*, various NO* donor compounds are used. Hydroxyurea (HU is shown to increase the levels of NO*.L- arginine is one of the non - essential amino acids. In the body L- arginine is used to make nitric oxide, which reduces blood vessel stiffness, increases blood flow and improves blood vessel function. The visible spectra of some transitional metals Cu, Fe(II,Fe(III,Cr, Mn, Ni have been studied individually in presence of hydroxyurea (HU with varying amounts .The spectra are also studied for the effect of varying amounts of metal ion on hydroxyurea. To observe how arginine itself acts on transitional metal ions. Even effect of Hydroxyurea on metal - arginine binding is also studied. The evaluation of these spectra is carried out for its binding parameters with the help of scatchard plots. The work has revealed certain very significant and interesting data which can have a lot of bearing on many chemical, biological and environmental aspects.

  13. Effect of L- Arginine On Electrocardiographic Changes Induced By Hypercholesterolemia And Isoproterenol In Rabbits

    OpenAIRE

    Kumar, Pradeep; Goyal, Manish; Agarwal, J L

    2009-01-01

    Hypercholesterolemia, a well-known cardiovascular risk factor, is associated with prolonged action potential duration, longer QTc intervals (rate controlled QT interval), suggested that Hypercholesterolemia may have a direct effect on ventricular repolarization. Hypercholesterolemia was induced in rabbits and L-arginine was given orally to animals for sixteen weeks. The isoproterenol was injected in all the animals to produce electrocardiographic changes. ECG was recorded in lead II at start ...

  14. Arginine vasopressin versus norepinephrine: will the stronger one win the race?

    OpenAIRE

    Ertmer, Christian; Bone, Hans-Georg; Westphal, Martin

    2006-01-01

    In the current issue of Critical Care, Friesenecker and colleagues present a well-designed comparative study on the microvascular effects of arginine vasopressin (AVP) and norepinephrine (NE) in a physiological, unanesthetized hamster model. The authors clearly demonstrate that AVP, but not NE, has marked vasoconstrictive effects on large arterioles, whereas the impact on small arterioles is comparable for both vasopressors. However, it remains unclear if these results, per se, reflect a stro...

  15. Levels of arginine-vasopressin in cerebrospinal fluid during passive avoidance behavior in rats

    OpenAIRE

    Kloet, E.R. de; Laczi, F.; Gaffori, O.; Fekete, M.; Wied, D. de

    1984-01-01

    The concentration of immunoreactive arginine-vasopressin (IR-AVP) was measured in the cerebrospinal fluid (CSF) during acquisition and retention of passive avoidance behavior. IR-AVP level in CSF of male Wistar rats immediately after the learning trial was increased; the rate of which was related to the intensity of the electric footschock during the learning trial and the avoidance latency as measured 1 day after the learning trial. Immediately after the 24 h retention test IR-AVP levels wer...

  16. Arteriolar vasoconstrictive response: comparing the effects of arginine vasopressin and norepinephrine

    OpenAIRE

    Friesenecker, Barbara E; Tsai, Amy G; Martini, Judith; Ulmer, Hanno; Wenzel, Volker; Hasibeder, Walter R; Intaglietta, Marcos; Dünser, Martin W

    2006-01-01

    INTRODUCTION: This study was designed to examine differences in the arteriolar vasoconstrictive response between arginine vasopressin (AVP) and norepinephrine (NE) on the microcirculatory level in the hamster window chamber model in unanesthetized, normotonic hamsters using intravital microscopy. It is known from patients with advanced vasodilatory shock that AVP exerts strong additional vasoconstriction when incremental dosage increases of NE have no further effect on mean arterial blood pre...

  17. Degradation of Arginine and Other Amino Acids by Eubacterium nodatum ATCC 33099

    OpenAIRE

    Uematsu, H.; Hoshino, E.

    2011-01-01

    The utilisation of a total of 20 amino acids by Eubacterium nodatum, a predominant asaccharolytic anaerobe isolated from human periodontal pockets, was studied. Washed cells of the microorganism produced substantial amounts of acetate, butyrate and ammonia from lysine, and butyrate and ammonia from arginine as main products under anaerobic conditions. They also produced a small amount of formate from histidine. Metabolic products were not detected from any of the other 17 amino acids. These r...

  18. The development of poly-L-arginine-coated liposomes for gene delivery

    OpenAIRE

    Opanasopit P; Tragulpakseerojn J; Apirakaramwong A; Ngawhirunpat T; Rojanarata T; Ruktanonchai U

    2011-01-01

    Praneet Opanasopit1, Jintana Tragulpakseerojn1, Auayporn Apirakaramwong1, Tanasait Ngawhirunpat1, Theerasak Rojanarata1, Uracha Ruktanonchai21Faculty of Pharmacy, Silpakorn University, Nakhon Pathom, Thailand; 2National Nanotechnology Center, Thailand Science Park, Pathumthani, Thailand Abstract: In this study, liposomes coated with cationic polymers, poly-L-arginine (PLA), were assessed as a promising gene transfer system in human cervical carcinoma (HeLa) cells and human h...

  19. Efficacy L-Arginine In Patients With Nonalcoholic Steatohepatitis Associated With Metabolic Syndrome

    OpenAIRE

    Oleksandr Fediv; Volodymyr Shevchuk; Oksana Olinyk

    2015-01-01

    Abstract Background and Purpose Recent research in the field of hematology indicate that among the many pathogenic mechanisms of development and progression of nonalcoholic steatohepatitis NASH which occurs on the background of the metabolic syndrome an important role is played by endothelial dysfunction and violations of haemocoagulation. The aim of this research was to study the effectiveness of L-arginine as it corrects endothelial dysfunction and disorders of homeostasis haemocoagulation ...

  20. A novel virulence strategy for Pseudomonas aeruginosa mediated by an autotransporter with arginine-specific aminopeptidase activity

    OpenAIRE

    Luckett, Jeni C. A.; Owen Darch; Chase Watters; Manal Abuoun; Victoria Wright; Esteban Paredes-Osses; Jenny Ward; Hana Goto; Stephan Heeb; Stéphanie Pommier; Rumbaugh, Kendra P.; Miguel Cámara; Hardie, Kim R.

    2012-01-01

    Author Summary We present a new Pseudomonas aeruginosa virulence factor that promotes chronic skin wound infections. We propose the name AaaA for this cell-surface tethered autotransporter. This arginine-specific aminopeptidase confers a growth advantage upon P. aeruginosa, providing a fitness advantage by creating a supply of arginine in chronic wounds where oxygen availability is limited and biofilm formation is involved. To our knowledge, this is the first mechanistic evidence linking the ...