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Sample records for core metabolic proteins

  1. Random close packing in protein cores.

    Science.gov (United States)

    Gaines, Jennifer C; Smith, W Wendell; Regan, Lynne; O'Hern, Corey S

    2016-03-01

    Shortly after the determination of the first protein x-ray crystal structures, researchers analyzed their cores and reported packing fractions ϕ ≈ 0.75, a value that is similar to close packing of equal-sized spheres. A limitation of these analyses was the use of extended atom models, rather than the more physically accurate explicit hydrogen model. The validity of the explicit hydrogen model was proved in our previous studies by its ability to predict the side chain dihedral angle distributions observed in proteins. In contrast, the extended atom model is not able to recapitulate the side chain dihedral angle distributions, and gives rise to large atomic clashes at side chain dihedral angle combinations that are highly probable in protein crystal structures. Here, we employ the explicit hydrogen model to calculate the packing fraction of the cores of over 200 high-resolution protein structures. We find that these protein cores have ϕ ≈ 0.56, which is similar to results obtained from simulations of random packings of individual amino acids. This result provides a deeper understanding of the physical basis of protein structure that will enable predictions of the effects of amino acid mutations to protein cores and interfaces of known structure.

  2. Core-shell microparticles for protein sequestration and controlled release of a protein-laden core.

    Science.gov (United States)

    Rinker, Torri E; Philbrick, Brandon D; Temenoff, Johnna S

    2017-07-01

    Development of multifunctional biomaterials that sequester, isolate, and redeliver cell-secreted proteins at a specific timepoint may be required to achieve the level of temporal control needed to more fully regulate tissue regeneration and repair. In response, we fabricated core-shell heparin-poly(ethylene-glycol) (PEG) microparticles (MPs) with a degradable PEG-based shell that can temporally control delivery of protein-laden heparin MPs. Core-shell MPs were fabricated via a re-emulsification technique and the number of heparin MPs per PEG-based shell could be tuned by varying the mass of heparin MPs in the precursor PEG phase. When heparin MPs were loaded with bone morphogenetic protein-2 (BMP-2) and then encapsulated into core-shell MPs, degradable core-shell MPs initiated similar C2C12 cell alkaline phosphatase (ALP) activity as the soluble control, while non-degradable core-shell MPs initiated a significantly lower response (85+19% vs. 9.0+4.8% of the soluble control, respectively). Similarly, when degradable core-shell MPs were formed and then loaded with BMP-2, they induced a ∼7-fold higher C2C12 ALP activity than the soluble control. As C2C12 ALP activity was enhanced by BMP-2, these studies indicated that degradable core-shell MPs were able to deliver a bioactive, BMP-2-laden heparin MP core. Overall, these dynamic core-shell MPs have the potential to sequester, isolate, and then redeliver proteins attached to a heparin core to initiate a cell response, which could be of great benefit to tissue regeneration applications requiring tight temporal control over protein presentation. Tissue repair requires temporally controlled presentation of potent proteins. Recently, biomaterial-mediated binding (sequestration) of cell-secreted proteins has emerged as a strategy to harness the regenerative potential of naturally produced proteins, but this strategy currently only allows immediate amplification and re-delivery of these signals. The multifunctional, dynamic

  3. Differential Stoichiometry among Core Ribosomal Proteins

    Directory of Open Access Journals (Sweden)

    Nikolai Slavov

    2015-11-01

    Full Text Available Understanding the regulation and structure of ribosomes is essential to understanding protein synthesis and its dysregulation in disease. While ribosomes are believed to have a fixed stoichiometry among their core ribosomal proteins (RPs, some experiments suggest a more variable composition. Testing such variability requires direct and precise quantification of RPs. We used mass spectrometry to directly quantify RPs across monosomes and polysomes of mouse embryonic stem cells (ESC and budding yeast. Our data show that the stoichiometry among core RPs in wild-type yeast cells and ESC depends both on the growth conditions and on the number of ribosomes bound per mRNA. Furthermore, we find that the fitness of cells with a deleted RP-gene is inversely proportional to the enrichment of the corresponding RP in polysomes. Together, our findings support the existence of ribosomes with distinct protein composition and physiological function.

  4. Random close packing in protein cores

    OpenAIRE

    Gaines, Jennifer C.; Smith, W. Wendell; Regan, Lynne; O'Hern, Corey S.

    2015-01-01

    Shortly after the determination of the first protein x-ray crystal structures, researchers analyzed their cores and reported packing fractions $\\phi \\approx 0.75$, a value that is similar to close packing equal-sized spheres. A limitation of these analyses was the use of `extended atom' models, rather than the more physically accurate `explicit hydrogen' model. The validity of using the explicit hydrogen model is proved by its ability to predict the side chain dihedral angle distributions obs...

  5. HCV core protein induces hepatic lipid accumulation by activating SREBP1 and PPARγ

    International Nuclear Information System (INIS)

    Kim, Kook Hwan; Hong, Sung Pyo; Kim, KyeongJin; Park, Min Jung; Kim, Kwang Jin; Cheong, JaeHun

    2007-01-01

    Hepatic steatosis is a common feature in patients with chronic hepatitis C virus (HCV) infection. HCV core protein plays an important role in the development of hepatic steatosis in HCV infection. Because SREBP1 (sterol regulatory element binding protein 1) and PPARγ (peroxisome proliferators-activated receptor γ) are involved in the regulation of lipid metabolism of hepatocyte, we sought to determine whether HCV core protein may impair the expression and activity of SREBP1 and PPARγ. In this study, it was demonstrated that HCV core protein increases the gene expression of SREBP1 not only in Chang liver, Huh7, and HepG2 cells transiently transfected with HCV core protein expression plasmid, but also in Chang liver-core stable cells. Furthermore, HCV core protein enhanced the transcriptional activity of SREBP1. In addition, HCV core protein elevated PPARγ transcriptional activity. However, HCV core protein had no effect on PPARγ gene expression. Finally, we showed that HCV core protein stimulates the genes expression of lipogenic enzyme and fatty acid uptake associated protein. Therefore, our finding provides a new insight into the mechanism of hepatic steatosis by HCV infection

  6. Tissue protein metabolism in parasitized animals

    International Nuclear Information System (INIS)

    Symons, L.E.A.; Steel, J.W.; Jones, W.O.

    1981-01-01

    The effects of gastrointestinal nematode infection of mammals, particularly of the small intestine of the sheep, on protein metabolism of skeletal muscle, liver, the gastrointestinal tract and wool are described. These changes have been integrated to explain poor growth and production in the sheep heavily infected with Trichostrongylus colubriformis. The rates of both synthesis and catabolism of muscle protein are depressed, but nitrogen is lost from this tissue because the depression of synthesis exceeds that of catabolism. Anorexia is the major cause of these changes. Although the effect on liver protein synthesis is unclear, it is probable that the leakage of plasma proteins into the gastrointestinal tract stimulates an early increase in the rate of synthesis of these proteins, but this eventually declines and is insufficient to correct developing hypoalbuminaemia. Changes in the intestinal tract are complex. Exogenous nitrogen is reduced by anorexia, but the flow of nitrogen through the tract from abomasum to faeces is above normal because of the increase of endogenous protein from leakage of plasma protein and, presumably, from exfoliated epithelial cells. There is evidence that protein metabolism of intestinal tissue, particularly in the uninfected distal two-thirds, is increased. Synthesis of wool protein is decreased. As the result of anorexia, intestinal loss of endogenous protein and an increased rate of intestinal protein metabolism there is a net movement of amino nitrogen from muscle, liver and possibly skin to the intestine of the heavily infected sheep. Thus, the availability of amino nitrogen for growth and wool production is reduced. (author)

  7. Cytokines: muscle protein and amino acid metabolism

    DEFF Research Database (Denmark)

    van Hall, Gerrit

    2012-01-01

    raises TNF-α and IL-6 to moderate levels, has only identified IL-6 as a potent cytokine, decreasing systemic amino acid levels and muscle protein metabolism. The marked decrease in circulatory and muscle amino acid concentrations was observed with a concomitant reduction in both the rates of muscle...... of IL-6 on the regulation of muscle protein metabolism but indirectly via IL-6 reducing amino acid availability. SUMMARY: Recent studies suggest that the best described cytokines TNF-α and IL-6 are unlikely to be the major direct mediators of muscle protein loss in inflammatory diseases. However...

  8. Amino acid metabolism conflicts with protein diversity

    OpenAIRE

    Krick, Teresa; Shub, David A.; Verstraete, Nina; Ferreiro, Diego U.; Alonso, Leonardo G.; Shub, Michael; Sanchez, Ignacio E.

    2014-01-01

    The 20 protein-coding amino acids are found in proteomes with different relative abundances. The most abundant amino acid, leucine, is nearly an order of magnitude more prevalent than the least abundant amino acid, cysteine. Amino acid metabolic costs differ similarly, constraining their incorporation into proteins. On the other hand, a diverse set of protein sequences is necessary to build functional proteomes. Here, we present a simple model for a cost-diversity trade-off postulating that n...

  9. Hepatitis C Virus Core Protein Decreases Lipid Droplet Turnover

    Science.gov (United States)

    Harris, Charles; Herker, Eva; Farese, Robert V.; Ott, Melanie

    2011-01-01

    Steatosis is a frequent complication of hepatitis C virus infection. In mice, this condition is recapitulated by the expression of a single viral protein, the nucleocapsid core. Core localizes to the surface of lipid droplets (LDs) in infected liver cells through a process dependent on host diacylglycerol acyltransferase 1 (DGAT1), an enzyme that synthesizes triglycerides in the endoplasmic reticulum. Whether DGAT1 also plays a role in core-induced steatosis is uncertain. Here, we show that mouse embryonic fibroblasts isolated from DGAT1−/− mice are protected from core-induced steatosis, as are livers of DGAT1−/− mice expressing core, demonstrating that the steatosis is DGAT1-dependent. Surprisingly, core expression did not increase DGAT1 activity or triglyceride synthesis, thus excluding the possibility that core activates DGAT1 to cause steatosis. Instead, we find that DGAT1-dependent localization of core to LDs is a prerequisite for the steatogenic properties of the core. Using biochemical and immunofluorescence microscopy techniques, we show that the turnover of lipids in core-coated droplets is decreased, providing a physiological mechanism for core-induced steatosis. Our results support a bipartite model in which core first requires DGAT1 to gain access to LDs, and then LD-localized core interferes with triglyceride turnover, thus stabilizing lipid droplets and leading to steatosis. PMID:21984835

  10. Mitochondrial uncoupling proteins and energy metabolism

    Directory of Open Access Journals (Sweden)

    Rosa Anna Busiello

    2015-02-01

    Full Text Available Understanding the metabolic factors that contribute to energy metabolism (EM is critical for the development of new treatments for obesity and related diseases. Mitochondrial oxidative phosphorylation is not perfectly coupled to ATP synthesis, and the process of proton-leak plays a crucial role. Proton-leak accounts for a significant part of the resting metabolic rate and therefore enhancement of this process represents a potential target for obesity treatment. Since their discovery, uncoupling proteins have stimulated great interest due to their involvement in mitochondrial-inducible proton-leak. Despite the widely accepted uncoupling/thermogenic effect of uncoupling protein one (UCP1, which was the first in this family to be discovered, the reactions catalyzed by its homologue UCP3 and the physiological role remain under debate.This review provides an overview of the role played by UCP1 and UCP3 in mitochondrial uncoupling/functionality as well as EM and suggests that they are a potential therapeutic target for treating obesity and its related diseases such as type II diabetes mellitus.

  11. Protein metabolism in marine animals: the underlying mechanism of growth.

    Science.gov (United States)

    Fraser, Keiron P P; Rogers, Alex D

    2007-01-01

    Growth is a fundamental process within all marine organisms. In soft tissues, growth is primarily achieved by the synthesis and retention of proteins as protein growth. The protein pool (all the protein within the organism) is highly dynamic, with proteins constantly entering the pool via protein synthesis or being removed from the pool via protein degradation. Any net change in the size of the protein pool, positive or negative, is termed protein growth. The three inter-related processes of protein synthesis, degradation and growth are together termed protein metabolism. Measurement of protein metabolism is vital in helping us understand how biotic and abiotic factors affect growth and growth efficiency in marine animals. Recently, the developing fields of transcriptomics and proteomics have started to offer us a means of greatly increasing our knowledge of the underlying molecular control of protein metabolism. Transcriptomics may also allow us to detect subtle changes in gene expression associated with protein synthesis and degradation, which cannot be detected using classical methods. A large literature exists on protein metabolism in animals; however, this chapter concentrates on what we know of marine ectotherms; data from non-marine ectotherms and endotherms are only discussed when the data are of particular relevance. We first consider the techniques available to measure protein metabolism, their problems and what validation is required. Protein metabolism in marine organisms is highly sensitive to a wide variety of factors, including temperature, pollution, seasonality, nutrition, developmental stage, genetics, sexual maturation and moulting. We examine how these abiotic and biotic factors affect protein metabolism at the level of whole-animal (adult and larval), tissue and cellular protein metabolism. Available gene expression data, which help us understand the underlying control of protein metabolism, are also discussed. As protein metabolism appears to

  12. Structural characterization of Mumps virus fusion protein core

    International Nuclear Information System (INIS)

    Liu Yueyong; Xu Yanhui; Lou Zhiyong; Zhu Jieqing; Hu Xuebo; Gao, George F.; Qiu Bingsheng; Rao Zihe; Tien, Po

    2006-01-01

    The fusion proteins of enveloped viruses mediating the fusion between the viral and cellular membranes comprise two discontinuous heptad repeat (HR) domains located at the ectodomain of the enveloped glycoproteins. The crystal structure of the fusion protein core of Mumps virus (MuV) was determined at 2.2 A resolution. The complex is a six-helix bundle in which three HR1 peptides form a central highly hydrophobic coiled-coil and three HR2 peptides pack against the hydrophobic grooves on the surface of central coiled-coil in an oblique antiparallel manner. Fusion core of MuV, like those of simian virus 5 and human respiratory syncytium virus, forms typical 3-4-4-4-3 spacing. The similar charecterization in HR1 regions, as well as the existence of O-X-O motif in extended regions of HR2 helix, suggests a basic rule for the formation of the fusion core of viral fusion proteins

  13. Modulation of mitogen-activated protein kinase-activated protein kinase 3 by hepatitis C virus core protein

    DEFF Research Database (Denmark)

    Ngo, HT; Pham, Long; Kim, JW

    2013-01-01

    Hepatitis C virus (HCV) is highly dependent on cellular proteins for its own propagation. In order to identify the cellular factors involved in HCV propagation, we performed protein microarray assays using the HCV core protein as a probe. Of ~9,000 host proteins immobilized in a microarray...... inducers. Binding of HCV core to MAPKAPK3 was confirmed by in vitro pulldown assay and further verified by coimmunoprecipitation assay. HCV core protein interacted with MAPKAPK3 through amino acid residues 41 to 75 of core and the N-terminal half of kinase domain of MAPKAPK3. In addition, both RNA...... increased HCV IRES-mediated translation and MAPKAPK3-dependent HCV IRES activity was further increased by core protein. These data suggest that HCV core may modulate MAPKAPK3 to facilitate its own propagation....

  14. Prediction of Hydrophobic Cores of Proteins Using Wavelet Analysis.

    Science.gov (United States)

    Hirakawa; Kuhara

    1997-01-01

    Information concerning the secondary structures, flexibility, epitope and hydrophobic regions of amino acid sequences can be extracted by assigning physicochemical indices to each amino acid residue, and information on structure can be derived using the sliding window averaging technique, which is in wide use for smoothing out raw functions. Wavelet analysis has shown great potential and applicability in many fields, such as astronomy, radar, earthquake prediction, and signal or image processing. This approach is efficient for removing noise from various functions. Here we employed wavelet analysis to smooth out a plot assigned to a hydrophobicity index for amino acid sequences. We then used the resulting function to predict hydrophobic cores in globular proteins. We calculated the prediction accuracy for the hydrophobic cores of 88 representative set of proteins. Use of wavelet analysis made feasible the prediction of hydrophobic cores at 6.13% greater accuracy than the sliding window averaging technique.

  15. Colorful packages : fluorescent proteins in complex coacervate core micelles

    NARCIS (Netherlands)

    Nolles, Antsje

    2018-01-01

    This thesis explores the encapsulation of fluorescent proteins (FPs) into complex coacervate core micelles (C3Ms) and features the impact of this encapsulation on the biophysical properties of the FPs. In total eight different FPs were investigated originating from two different classes

  16. Protein design in systems metabolic engineering for industrial strain development.

    Science.gov (United States)

    Chen, Zhen; Zeng, An-Ping

    2013-05-01

    Accelerating the process of industrial bacterial host strain development, aimed at increasing productivity, generating new bio-products or utilizing alternative feedstocks, requires the integration of complementary approaches to manipulate cellular metabolism and regulatory networks. Systems metabolic engineering extends the concept of classical metabolic engineering to the systems level by incorporating the techniques used in systems biology and synthetic biology, and offers a framework for the development of the next generation of industrial strains. As one of the most useful tools of systems metabolic engineering, protein design allows us to design and optimize cellular metabolism at a molecular level. Here, we review the current strategies of protein design for engineering cellular synthetic pathways, metabolic control systems and signaling pathways, and highlight the challenges of this subfield within the context of systems metabolic engineering. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Homogeneous protein analysis by magnetic core-shell nanorod probes

    KAUST Repository

    Schrittwieser, Stefan

    2016-03-29

    Studying protein interactions is of vital importance both to fundamental biology research and to medical applications. Here, we report on the experimental proof of a universally applicable label-free homogeneous platform for rapid protein analysis. It is based on optically detecting changes in the rotational dynamics of magnetically agitated core-shell nanorods upon their specific interaction with proteins. By adjusting the excitation frequency, we are able to optimize the measurement signal for each analyte protein size. In addition, due to the locking of the optical signal to the magnetic excitation frequency, background signals are suppressed, thus allowing exclusive studies of processes at the nanoprobe surface only. We study target proteins (soluble domain of the human epidermal growth factor receptor 2 - sHER2) specifically binding to antibodies (trastuzumab) immobilized on the surface of our nanoprobes and demonstrate direct deduction of their respective sizes. Additionally, we examine the dependence of our measurement signal on the concentration of the analyte protein, and deduce a minimally detectable sHER2 concentration of 440 pM. For our homogeneous measurement platform, good dispersion stability of the applied nanoprobes under physiological conditions is of vital importance. To that end, we support our measurement data by theoretical modeling of the total particle-particle interaction energies. The successful implementation of our platform offers scope for applications in biomarker-based diagnostics as well as for answering basic biology questions.

  18. Radioactive Lysine in Protein Metabolism Studies

    Science.gov (United States)

    Miller, L. L.; Bale, W. F.; Yuile, C. L.; Masters, R. E.; Tishkoff, G. H.; Whipple,, G. H.

    1950-01-09

    Studies of incorporation of DL-lysine in various body proteins of the dog; the time course of labeled blood proteins; and apparent rate of disappearance of labeled plasma proteins for comparison of behavior of the plasma albumin and globulin fractions; shows more rapid turn over of globulin fraction.

  19. Metabolic behavior of cell surface biotinylated proteins

    International Nuclear Information System (INIS)

    Hare, J.F.; Lee, E.

    1989-01-01

    The turnover of proteins on the surface of cultured mammalian cells was measured by a new approach. Reactive free amino or sulfhydryl groups on surface-accessible proteins were derivatized with biotinyl reagents and the proteins solubilized from culture dishes with detergent. Solubilized, biotinylated proteins were then adsorbed onto streptavidin-agarose, released with sodium dodecyl sulfate and mercaptoethanol, and separated on polyacrylamide gels. Biotin-epsilon-aminocaproic acid N-hydroxysuccinimide ester (BNHS) or N-biotinoyl-N'-(maleimidohexanoyl)hydrazine (BM) were the derivatizing agents. Only 10-12 bands were adsorbed onto streptavidin-agarose from undervatized cells or from derivatized cells treated with free avidin at 4 degrees C. Two-dimensional isoelectric focusing-sodium dodecyl sulfate gel electrophoresis resolved greater than 100 BNHS-derivatized proteins and greater than 40 BM-derivatized proteins. There appeared to be little overlap between the two groups of derivatized proteins. Short-term pulse-chase studies showed an accumulation of label into both groups of biotinylated proteins up until 1-2 h of chase and a rapid decrease over the next 1-5 h. Delayed appearance of labeled protein at the cell surface was attributed to transit time from site of synthesis. The unexpected and unexplained rapid disappearance of pulse-labeled proteins from the cell surface was invariant for all two-dimensionally resolved proteins and was sensitive to temperature reduction to 18 degrees C. Long-term pulse-chase experiments beginning 4-8 h after the initiation of chase showed the disappearance of derivatized proteins to be a simple first-order process having a half-life of 115 h in the case of BNHS-derivatized proteins and 30 h in the case of BM-derivatized proteins

  20. Effect of dietary protein restriction on renal ammonia metabolism

    Science.gov (United States)

    Lee, Hyun-Wook; Osis, Gunars; Handlogten, Mary E.; Guo, Hui; Verlander, Jill W.

    2015-01-01

    Dietary protein restriction has multiple benefits in kidney disease. Because protein intake is a major determinant of endogenous acid production, it is important that net acid excretion change in parallel during protein restriction. Ammonia is the primary component of net acid excretion, and inappropriate ammonia excretion can lead to negative nitrogen balance. Accordingly, we examined ammonia excretion in response to protein restriction and then we determined the molecular mechanism of the changes observed. Wild-type C57Bl/6 mice fed a 20% protein diet and then changed to 6% protein developed an 85% reduction in ammonia excretion within 2 days, which persisted during a 10-day study. The expression of multiple proteins involved in renal ammonia metabolism was altered, including the ammonia-generating enzymes phosphate-dependent glutaminase (PDG) and phosphoenolpyruvate carboxykinase (PEPCK) and the ammonia-metabolizing enzyme glutamine synthetase. Rhbg, an ammonia transporter, increased in expression in the inner stripe of outer medullary collecting duct intercalated cell (OMCDis-IC). However, collecting duct-specific Rhbg deletion did not alter the response to protein restriction. Rhcg deletion did not alter ammonia excretion in response to dietary protein restriction. These results indicate 1) dietary protein restriction decreases renal ammonia excretion through coordinated regulation of multiple components of ammonia metabolism; 2) increased Rhbg expression in the OMCDis-IC may indicate a biological role in addition to ammonia transport; and 3) Rhcg expression is not necessary to decrease ammonia excretion during dietary protein restriction. PMID:25925252

  1. Leucine and protein metabolism in obese zucker rats

    Science.gov (United States)

    Branched-chain amino acids (BCAAs) are circulating nutrient signals for protein accretion, however they increase in obesity and appear to prognosticate diabetes onset. To understand the mechanisms whereby obesity affects BCAAs and protein metabolism, we employed metabolomics and measured rates of [1...

  2. Liver and muscle protein metabolism in cachexia

    NARCIS (Netherlands)

    Peters, J.A.C.

    2009-01-01

    Up to 50% of cancer patients suffer from progressive weight loss (cachexia). Cachexia is induced by proinflammatory mediators (cytokines), induced by the tumor’s presence. These cytokines induce so-called acute phase protein synthesis by the liver, followed by skeletal muscle protein breakdown.

  3. Hepatitis B core protein as a therapeutic target.

    Science.gov (United States)

    Mak, Lung-Yi; Wong, Danny Ka-Ho; Seto, Wai-Kay; Lai, Ching-Lung; Yuen, Man Fung

    2017-12-01

    Chronic hepatitis B virus (HBV) infection is difficult to cure, due to the presence of covalently-closed-circular DNA and virus-mediated blunting of host immune response. Existing therapies with nucleos(t)ide analogue or pegylated-interferon are not sufficient to achieve a high rate of HBV surface antigen seroclearance, a more desirable treatment outcome. Novel therapeutic agents targeting alternative viral replication steps are being developed. In this review, we will discuss the hepatitis B core antigen (HBcAg) as a therapeutic target. Areas covered: The basic structure and fundamental functions of HBcAg including nucleocapsid assembly, pre-genomic RNA encapsidation, reverse transcription, virion formation, cccDNA amplification, immune response regulation, and HBx protein interaction will be reviewed. Most of these are identified as therapeutic targets and tested in in vitro and in vivo studies, although clinical trials are scanty. Among the different components, the core protein allosteric modulators (CpAM) have been most widely investigated and appear promising in clinical trials. Expert opinion: The multiple and essential functions of HBcAg for HBV life cycle are important and attractive targets for HBV therapeutic interventions. Controlled trials involving CpAM are awaited. Apart from CpAM, drugs directed against different functions of HBcAg may be further explored to maximize the chance of cure.

  4. Hepatitis B Virus Core Protein Dephosphorylation Occurs during Pregenomic RNA Encapsidation.

    Science.gov (United States)

    Zhao, Qiong; Hu, Zhanying; Cheng, Junjun; Wu, Shuo; Luo, Yue; Chang, Jinhong; Hu, Jianming; Guo, Ju-Tao

    2018-07-01

    Hepatitis B virus (HBV) core protein consists of an N-terminal assembly domain and a C-terminal domain (CTD) with seven conserved serines or threonines that are dynamically phosphorylated/dephosphorylated during the viral replication cycle. Sulfamoylbenzamide derivatives are small molecular core protein allosteric modulators (CpAMs) that bind to the heteroaryldihydropyrimidine (HAP) pocket between the core protein dimer-dimer interfaces. CpAM binding alters the kinetics and pathway of capsid assembly and can result in the formation of morphologically "normal" capsids devoid of viral pregenomic RNA (pgRNA) and DNA polymerase. In order to investigate the mechanism underlying CpAM inhibition of pgRNA encapsidation, we developed an immunoblotting assay that can resolve core protein based on its phosphorylation status and demonstrated, for the first time, that core protein is hyperphosphorylated in free dimers and empty capsids from both mock-treated and CpAM-treated cells but is hypophosphorylated in pgRNA- and DNA-containing nucleocapsids. Interestingly, inhibition of pgRNA encapsidation by a heat shock protein 90 (HSP90) inhibitor prevented core protein dephosphorylation. Moreover, core proteins with point mutations at the wall of the HAP pocket, V124A and V124W, assembled empty capsids and nucleocapsids with altered phosphorylation status. The results thus suggest that core protein dephosphorylation occurs in the assembly of pgRNA and that interference with the interaction between core protein subunits at dimer-dimer interfaces during nucleocapsid assembly alters not only capsid structure, but also core protein dephosphorylation. Hence, inhibition of pgRNA encapsidation by CpAMs might be due to disruption of core protein dephosphorylation during nucleocapsid assembly. IMPORTANCE Dynamic phosphorylation of HBV core protein regulates multiple steps of viral replication. However, the regulatory function was mainly investigated by phosphomimetic mutagenesis, which

  5. Adjustments of Protein Metabolism in Fasting Arctic Charr, Salvelinus alpinus.

    Directory of Open Access Journals (Sweden)

    Alicia A Cassidy

    Full Text Available Protein metabolism, including the interrelated processes of synthesis and degradation, mediates the growth of an animal. In ectothermic animals, protein metabolism is responsive to changes in both biotic and abiotic conditions. This study aimed to characterise responses of protein metabolism to food deprivation that occur in the coldwater salmonid, Arctic charr, Salvelinus alpinus. We compared two groups of Arctic charr: one fed continuously and the other deprived of food for 36 days. We measured the fractional rate of protein synthesis (KS in individuals from the fed and fasted groups using a flooding dose technique modified for the use of deuterium-labelled phenylalanine. The enzyme activities of the three major protein degradation pathways (ubiquitin proteasome, lysosomal cathepsins and the calpain systems were measured in the same fish. This study is the first to measure both KS and the enzymatic activity of protein degradation in the same fish, allowing us to examine the apparent contribution of different protein degradation pathways to protein turnover in various tissues (red and white muscle, liver, heart and gills. KS was lower in the white muscle and in liver of the fasted fish compared to the fed fish. There were no observable effects of food deprivation on the protease activities in any of the tissues with the exception of liver, where the ubiquitin proteasome pathway seemed to be activated during fasting conditions. Lysosomal proteolysis appears to be the primary degradation pathway for muscle protein, while the ubiquitin proteasome pathway seems to predominate in the liver. We speculate that Arctic charr regulate protein metabolism during food deprivation to conserve proteins.

  6. Both core and F proteins of hepatitis C virus could enhance cell proliferation in transgenic mice

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Wen-Ta [Graduate Institute of Medical Biotechnology, Tzu Chi University, Hualien, Taiwan (China); Li, Hui-Chun [Department of Biochemistry, Tzu Chi University, Hualien, Taiwan (China); Lee, Shen-Kao; Ma, Hsin-Chieh; Yang, Chee-Hing; Chen, Hung-Ling [Graduate Institute of Medical Biotechnology, Tzu Chi University, Hualien, Taiwan (China); Lo, Shih-Yen, E-mail: losylo@mail.tcu.edu.tw [Graduate Institute of Medical Biotechnology, Tzu Chi University, Hualien, Taiwan (China); Department of Laboratory Medicine, Buddhist Tzu Chi General Hospital, Hualien, Taiwan (China)

    2013-05-24

    Highlights: •HCV core and F proteins could induce hepatocyte proliferation in the transgenic mice. •β-Catenin signaling pathway was activated by core protein in the transgenic mice. •β-Catenin signaling pathway was activated by myc-F protein in the transgenic mice. •Expression of SMA protein was enhanced by core but not myc-F protein. -- Abstract: The role of the protein encoded by the alternative open reading frame (ARF/F/core+1) of the Hepatitis C virus (HCV) genome in viral pathogenesis remains unknown. The different forms of ARF/F/core+1 protein were labile in cultured cells, a myc-tag fused at the N-terminus of the F protein made it more stable. To determine the role of core and F proteins in HCV pathogenesis, transgenic mice with either protein expression under the control of Albumin promoter were generated. Expression of core protein and F protein with myc tag (myc-F) could be detected by Western blotting analysis in the livers of these mice. The ratio of liver to body weight is increased for both core and myc-F transgenic mice compared to that of wild type mice. Indeed, the proliferating cell nuclear antigen protein, a proliferation marker, was up-regulated in the transgenic mice with core or myc-F protein. Further analyses by microarray and Western blotting suggested that β-catenin signaling pathway was activated by either core or myc-F protein in the transgenic mice. These transgenic mice were further treated with either Diethynitrosamine (a tumor initiator) or Phenobarbital (a tumor promoter). Phenobarbital but not Diethynitrosamine treatment could increase the liver/body weight ratio of these mice. However, no tumor formation was observed in these mice. In conclusion, HCV core and myc-F proteins could induce hepatocyte proliferation in the transgenic mice possibly through β-catenin signaling pathway.

  7. Both core and F proteins of hepatitis C virus could enhance cell proliferation in transgenic mice

    International Nuclear Information System (INIS)

    Hu, Wen-Ta; Li, Hui-Chun; Lee, Shen-Kao; Ma, Hsin-Chieh; Yang, Chee-Hing; Chen, Hung-Ling; Lo, Shih-Yen

    2013-01-01

    Highlights: •HCV core and F proteins could induce hepatocyte proliferation in the transgenic mice. •β-Catenin signaling pathway was activated by core protein in the transgenic mice. •β-Catenin signaling pathway was activated by myc-F protein in the transgenic mice. •Expression of SMA protein was enhanced by core but not myc-F protein. -- Abstract: The role of the protein encoded by the alternative open reading frame (ARF/F/core+1) of the Hepatitis C virus (HCV) genome in viral pathogenesis remains unknown. The different forms of ARF/F/core+1 protein were labile in cultured cells, a myc-tag fused at the N-terminus of the F protein made it more stable. To determine the role of core and F proteins in HCV pathogenesis, transgenic mice with either protein expression under the control of Albumin promoter were generated. Expression of core protein and F protein with myc tag (myc-F) could be detected by Western blotting analysis in the livers of these mice. The ratio of liver to body weight is increased for both core and myc-F transgenic mice compared to that of wild type mice. Indeed, the proliferating cell nuclear antigen protein, a proliferation marker, was up-regulated in the transgenic mice with core or myc-F protein. Further analyses by microarray and Western blotting suggested that β-catenin signaling pathway was activated by either core or myc-F protein in the transgenic mice. These transgenic mice were further treated with either Diethynitrosamine (a tumor initiator) or Phenobarbital (a tumor promoter). Phenobarbital but not Diethynitrosamine treatment could increase the liver/body weight ratio of these mice. However, no tumor formation was observed in these mice. In conclusion, HCV core and myc-F proteins could induce hepatocyte proliferation in the transgenic mice possibly through β-catenin signaling pathway

  8. Regulation of intestinal protein metabolism by amino acids.

    Science.gov (United States)

    Bertrand, Julien; Goichon, Alexis; Déchelotte, Pierre; Coëffier, Moïse

    2013-09-01

    Gut homeostasis plays a major role in health and may be regulated by quantitative and qualitative food intake. In the intestinal mucosa, an intense renewal of proteins occurs, at approximately 50% per day in humans. In some pathophysiological conditions, protein turnover is altered and may contribute to intestinal or systemic diseases. Amino acids are key effectors of gut protein turnover, both as constituents of proteins and as regulatory molecules limiting intestinal injury and maintaining intestinal functions. Many studies have focused on two amino acids: glutamine, known as the preferential substrate of rapidly dividing cells, and arginine, another conditionally essential amino acid. The effects of glutamine and arginine on protein synthesis appear to be model and condition dependent, as are the involved signaling pathways. The regulation of gut protein degradation by amino acids has been minimally documented until now. This review will examine recent data, helping to better understand how amino acids regulate intestinal protein metabolism, and will explore perspectives for future studies.

  9. Efficient protein production by yeast requires global tuning of metabolism

    DEFF Research Database (Denmark)

    Huang, Mingtao; Bao, Jichen; Hallstrom, Bjorn M.

    2017-01-01

    The biotech industry relies on cell factories for production of pharmaceutical proteins, of which several are among the top-selling medicines. There is, therefore, considerable interest in improving the efficiency of protein production by cell factories. Protein secretion involves numerous...... intracellular processes with many underlying mechanisms still remaining unclear. Here, we use RNA-seq to study the genome-wide transcriptional response to protein secretion in mutant yeast strains. We find that many cellular processes have to be attuned to support efficient protein secretion. In particular...... that by tuning metabolism cells are able to efficiently secrete recombinant proteins. Our findings provide increased understanding of which cellular regulations and pathways are associated with efficient protein secretion....

  10. The Expanded FindCore Method for Identification of a Core Atom Set for Assessment of Protein Structure Prediction

    Science.gov (United States)

    Snyder, David A.; Grullon, Jennifer; Huang, Yuanpeng J.; Tejero, Roberto; Montelione, Gaetano T.

    2014-01-01

    Maximizing the scientific impact of NMR-based structure determination requires robust and statistically sound methods for assessing the precision of NMR-derived structures. In particular, a method to define a core atom set for calculating superimpositions and validating structure predictions is critical to the use of NMR-derived structures as targets in the CASP competition. FindCore (D.A. Snyder and G.T. Montelione PROTEINS 2005;59:673–686) is a superimposition independent method for identifying a core atom set, and partitioning that set into domains. However, as FindCore optimizes superimposition by sensitively excluding not-well-defined atoms, the FindCore core may not comprise all atoms suitable for use in certain applications of NMR structures, including the CASP assessment process. Adapting the FindCore approach to assess predicted models against experimental NMR structures in CASP10 required modification of the FindCore method. This paper describes conventions and a standard protocol to calculate an “Expanded FindCore” atom set suitable for validation and application in biological and biophysical contexts. A key application of the Expanded FindCore method is to identify a core set of atoms in the experimental NMR structure for which it makes sense to validate predicted protein structure models. We demonstrate the application of this Expanded FindCore method in characterizing well-defined regions of 18 NMR-derived CASP10 target structures. The Expanded FindCore protocol defines “expanded core atom sets” that match an expert’s intuition of which parts of the structure are sufficiently well-defined to use in assessing CASP model predictions. We also illustrate the impact of this analysis on the CASP GDT assessment scores. PMID:24327305

  11. Effects of reducing dietary crude protein and metabolic energy in ...

    African Journals Online (AJOL)

    The objective of this experiment was to determine the effects of a pure reduction in the dietary crude protein (CP) and metabolic energy (ME) contents on growth performance, nutrient digestibility, blood profile, faecal microflora and odour gas emission in weaned pigs. A total of 80 weaned piglets ((Landrace × Yorkshire) ...

  12. FANCM-FAAP24 and FANCJ: FA proteins that metabolize DNA

    Energy Technology Data Exchange (ETDEWEB)

    Ali, Abdullah Mahmood; Singh, Thiyam Ramsing [Division of Experimental Hematology and Cancer Biology, Cincinnati Children' s Research Foundation, Cincinnati Children' s Hospital Medical Center, Cincinnati, OH 45229 (United States); Meetei, Amom Ruhikanta, E-mail: Ruhikanta.Meetei@cchmc.org [Division of Experimental Hematology and Cancer Biology, Cincinnati Children' s Research Foundation, Cincinnati Children' s Hospital Medical Center, Cincinnati, OH 45229 (United States); Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, OH 45229 (United States)

    2009-07-31

    Fanconi anemia (FA) is a rare autosomal recessive or X-linked disorder characterized by aplastic anemia, cancer susceptibility and cellular sensitivity to DNA-crosslinking agents. Eight FA proteins (FANCA, -B, -C, -E, -F, -G, -L and -M) and three non-FA proteins (FAAP100, FAAP24 and HES1) form the FA nuclear core complex that is required for monoubiquitination of the FANCD2-FANCI dimer upon DNA damage. The other three FA proteins, FANCD1/BRCA2, FANCJ/BACH1/BRIP1 and FANCN/PALB2, act in parallel or downstream of the FANCD2-FANCI dimer. Despite the isolation and characterization of several FA proteins, the mechanism by which these proteins protect cells from DNA interstrand crosslinking agents has been unclear. This is because a majority of the FA proteins lack any recognizable functional domains that can provide insight into their function. The recently discovered FANCM (Hef) and FANCJ (BRIP1/BACH1) proteins contain helicase domains, providing potential insight into the role of FA proteins in DNA repair. FANCM with its partner, FAAP24, and FANCJ bind and metabolize a variety of DNA substrates. In this review, we focus on the discovery, structure, and function of the FANCM-FAAP24 and FANCJ proteins.

  13. FANCM-FAAP24 and FANCJ: FA proteins that metabolize DNA

    International Nuclear Information System (INIS)

    Ali, Abdullah Mahmood; Singh, Thiyam Ramsing; Meetei, Amom Ruhikanta

    2009-01-01

    Fanconi anemia (FA) is a rare autosomal recessive or X-linked disorder characterized by aplastic anemia, cancer susceptibility and cellular sensitivity to DNA-crosslinking agents. Eight FA proteins (FANCA, -B, -C, -E, -F, -G, -L and -M) and three non-FA proteins (FAAP100, FAAP24 and HES1) form the FA nuclear core complex that is required for monoubiquitination of the FANCD2-FANCI dimer upon DNA damage. The other three FA proteins, FANCD1/BRCA2, FANCJ/BACH1/BRIP1 and FANCN/PALB2, act in parallel or downstream of the FANCD2-FANCI dimer. Despite the isolation and characterization of several FA proteins, the mechanism by which these proteins protect cells from DNA interstrand crosslinking agents has been unclear. This is because a majority of the FA proteins lack any recognizable functional domains that can provide insight into their function. The recently discovered FANCM (Hef) and FANCJ (BRIP1/BACH1) proteins contain helicase domains, providing potential insight into the role of FA proteins in DNA repair. FANCM with its partner, FAAP24, and FANCJ bind and metabolize a variety of DNA substrates. In this review, we focus on the discovery, structure, and function of the FANCM-FAAP24 and FANCJ proteins.

  14. The use of LeptiCore® in reducing fat gain and managing weight loss in patients with metabolic syndrome

    Directory of Open Access Journals (Sweden)

    Ngondi Judith L

    2010-02-01

    Full Text Available Abstract Background LeptiCore® is a proprietary combination of various ingredients which have been shown to have properties which could be beneficial to weight loss in obese and overweight human subjects. This study evaluates the effect of Lepticore® on bodyweight as well as parameters associated with obesity and metabolic syndrome. Methods The study was an 8 week randomized, double-blind, placebo-controlled design involving 92 obese (mean BMI > 30 kg/m2 participants (37 males; 55 females; ages 19-52; mean age = 30.7. The participants were randomly divided into three groups: placebo (n = 30, LeptiCore® formula A (low dose (n = 31 and LeptiCore® formula B (high dose (n = 31. Capsules containing the placebo or active formulations were administered twice daily before meals with 300 ml of water. None of the participants followed any specific diet nor took any weight-reducing medications for the duration of the study. A total of 12 anthropomorphic and serological measurements were taken at the beginning of the study and after 2, 4, 6, and 8 weeks of treatment. Results Compared to the placebo group, the two active groups showed statistically significant differences on all 12 variables by week 8. These included four anthropomorphic variables (body weight, body fat, waist and hip size and eight measures of serological levels (plasma total cholesterol, LDL, HDL, triglycerides, blood glucose, serotonin, leptin, C-reactive protein. The two active groups also showed significant intra-group differences on all 12 variables between study onset and week 8. Conclusion The LeptiCore® formulation at both the low and high dosages appears to be helpful in the management of fat gain and its related complications. The higher dosage resulted in significantly greater reductions in body weight and triglyceride, blood glucose, and C-reactive protein levels, as well as increased serotonin levels.

  15. Protein and leucine metabolism in maple syrup urine disease

    International Nuclear Information System (INIS)

    Thompson, G.N.; Bresson, J.L.; Pacy, P.J.; Bonnefont, J.P.; Walter, J.H.; Leonard, J.V.; Saudubray, J.M.; Halliday, D.

    1990-01-01

    Constant infusions of [13C]leucine and [2H5]phenylalanine were used to trace leucine and protein kinetics, respectively, in seven children with maple syrup urine disease (MSUD) and eleven controls matched for age and dietary protein intake. Despite significant elevations of plasma leucine (mean 351 mumol/l, range 224-477) in MSUD subjects, mean whole body protein synthesis [3.78 +/- 0.42 (SD) g.kg-1. 24 h-1] and catabolism (4.07 +/- 0.46) were similar to control values (3.69 +/- 0.50 and 4.09 +/- 0.50, respectively). The relationship between phenylalanine and leucine fluxes was also similar in MSUD subjects (mean phenylalanine-leucine flux ratio 0.35 +/- 0.07) and previously reported adult controls (0.33 +/- 0.02). Leucine oxidation was undetectable in four of the MSUD subjects and very low in the other three (less than 4 mumol.kg-1.h-1; controls 13-20). These results show that persistent elevation in leucine concentration has no effect on protein synthesis. The marked disturbance in leucine metabolism in MSUD did not alter the relationship between rates of catabolism of protein to phenylalanine and leucine, which provides further support for the validity of the use of a single amino acid to trace whole body protein metabolism. The minimal leucine oxidation in MSUD differs from findings in other inborn metabolic errors and indicates that in patients with classical MSUD there is no significant route of leucine disposal other than through protein synthesis

  16. The retrovirus MA and PreTM proteins follow immature MVL cores

    DEFF Research Database (Denmark)

    Andersen, Klaus Bahl

    2013-01-01

    Detergent can dissolve retrovirus, exept the immature core. Here we show that the Matrix protein (MA) and the Transmembrane protein in its immature form (PreTM) bind to the retrovirus core. These attachments explain the attachment in the virus particle and the dynamics of the ability to fuse with...

  17. Identification of a functional, CRM-1-dependent nuclear export signal in hepatitis C virus core protein.

    Directory of Open Access Journals (Sweden)

    Andrea Cerutti

    Full Text Available Hepatitis C virus (HCV infection is a major cause of chronic liver disease worldwide. HCV core protein is involved in nucleocapsid formation, but it also interacts with multiple cytoplasmic and nuclear molecules and plays a crucial role in the development of liver disease and hepatocarcinogenesis. The core protein is found mostly in the cytoplasm during HCV infection, but also in the nucleus in patients with hepatocarcinoma and in core-transgenic mice. HCV core contains nuclear localization signals (NLS, but no nuclear export signal (NES has yet been identified.We show here that the aa(109-133 region directs the translocation of core from the nucleus to the cytoplasm by the CRM-1-mediated nuclear export pathway. Mutagenesis of the three hydrophobic residues (L119, I123 and L126 in the identified NES or in the sequence encoding the mature core aa(1-173 significantly enhanced the nuclear localisation of the corresponding proteins in transfected Huh7 cells. Both the NES and the adjacent hydrophobic sequence in domain II of core were required to maintain the core protein or its fragments in the cytoplasmic compartment. Electron microscopy studies of the JFH1 replication model demonstrated that core was translocated into the nucleus a few minutes after the virus entered the cell. The blockade of nucleocytoplasmic export by leptomycin B treatment early in infection led to the detection of core protein in the nucleus by confocal microscopy and coincided with a decrease in virus replication.Our data suggest that the functional NLS and NES direct HCV core protein shuttling between the cytoplasmic and nuclear compartments, with at least some core protein transported to the nucleus. These new properties of HCV core may be essential for virus multiplication and interaction with nuclear molecules, influence cell signaling and the pathogenesis of HCV infection.

  18. Identification of a functional, CRM-1-dependent nuclear export signal in hepatitis C virus core protein.

    Science.gov (United States)

    Cerutti, Andrea; Maillard, Patrick; Minisini, Rosalba; Vidalain, Pierre-Olivier; Roohvand, Farzin; Pecheur, Eve-Isabelle; Pirisi, Mario; Budkowska, Agata

    2011-01-01

    Hepatitis C virus (HCV) infection is a major cause of chronic liver disease worldwide. HCV core protein is involved in nucleocapsid formation, but it also interacts with multiple cytoplasmic and nuclear molecules and plays a crucial role in the development of liver disease and hepatocarcinogenesis. The core protein is found mostly in the cytoplasm during HCV infection, but also in the nucleus in patients with hepatocarcinoma and in core-transgenic mice. HCV core contains nuclear localization signals (NLS), but no nuclear export signal (NES) has yet been identified.We show here that the aa(109-133) region directs the translocation of core from the nucleus to the cytoplasm by the CRM-1-mediated nuclear export pathway. Mutagenesis of the three hydrophobic residues (L119, I123 and L126) in the identified NES or in the sequence encoding the mature core aa(1-173) significantly enhanced the nuclear localisation of the corresponding proteins in transfected Huh7 cells. Both the NES and the adjacent hydrophobic sequence in domain II of core were required to maintain the core protein or its fragments in the cytoplasmic compartment. Electron microscopy studies of the JFH1 replication model demonstrated that core was translocated into the nucleus a few minutes after the virus entered the cell. The blockade of nucleocytoplasmic export by leptomycin B treatment early in infection led to the detection of core protein in the nucleus by confocal microscopy and coincided with a decrease in virus replication.Our data suggest that the functional NLS and NES direct HCV core protein shuttling between the cytoplasmic and nuclear compartments, with at least some core protein transported to the nucleus. These new properties of HCV core may be essential for virus multiplication and interaction with nuclear molecules, influence cell signaling and the pathogenesis of HCV infection.

  19. Radioisotope techniques in the study of protein metabolism

    International Nuclear Information System (INIS)

    1965-01-01

    The International Atomic Energy Agency (IAEA) held a panel meeting on June 1-5, 1964. The purpose of the panel was to discuss the present status of radioactive tracer techniques for the study of protein metabolism and to suggest ways of extending an co-ordinating the Agency's research programme in this field. The meeting was attended by 13 invited experts from ten different countries, and three representatives of the World Health Organization (WHO). Sessions of the panel were devoted to methods of preparation of labelled proteins and protein-like substances, to techniques of measurement of gastro-intestinal protein absorption and loss and to the clinical applications of these techniques. At each session, working papers were presented by various participants and then discussed in detail. This report gives the full texts of the working papers together with extensive summaries of the discussions and provides a detailed picture of the present situation and likely future developments in this field of work. It is hoped that its publication will be of interest to all concerned with problems of protein metabolism, whether in clinical medicine or the basic medical sciences. 349 refs, figs and tabs

  20. 5' adenosine monophosphate-activated protein kinase, metabolism and exercise.

    Science.gov (United States)

    Aschenbach, William G; Sakamoto, Kei; Goodyear, Laurie J

    2004-01-01

    The 5' adenosine monophosphate-activated protein kinase (AMPK) is a member of a metabolite-sensing protein kinase family that functions as a metabolic 'fuel gauge' in skeletal muscle. AMPK is a ubiquitous heterotrimeric protein, consisting of an alpha catalytic, and beta and gamma regulatory subunits that exist in multiple isoforms and are all required for full enzymatic activity. During exercise, AMPK becomes activated in skeletal muscle in response to changes in cellular energy status (e.g. increased adenosine monophosphate [AMP]/adenosine triphosphate [ATP] and creatine/phosphocreatine ratios) in an intensity-dependent manner, and serves to inhibit ATP-consuming pathways, and activate pathways involved in carbohydrate and fatty-acid metabolism to restore ATP levels. Recent evidence shows that although AMPK plays this key metabolic role during acute bouts of exercise, it is also an important component of the adaptive response of skeletal muscles to endurance exercise training because of its ability to alter muscle fuel reserves and expression of several exercise-responsive genes. This review discusses the putative roles of AMPK in acute and chronic exercise responses, and suggests avenues for future AMPK research in exercise physiology and biochemistry.

  1. Bidirectional lipid droplet velocities are controlled by differential binding strengths of HCV core DII protein.

    Directory of Open Access Journals (Sweden)

    Rodney K Lyn

    Full Text Available Host cell lipid droplets (LD are essential in the hepatitis C virus (HCV life cycle and are targeted by the viral capsid core protein. Core-coated LDs accumulate in the perinuclear region and facilitate viral particle assembly, but it is unclear how mobility of these LDs is directed by core. Herein we used two-photon fluorescence, differential interference contrast imaging, and coherent anti-Stokes Raman scattering microscopies, to reveal novel core-mediated changes to LD dynamics. Expression of core protein's lipid binding domain II (DII-core induced slower LD speeds, but did not affect directionality of movement on microtubules. Modulating the LD binding strength of DII-core further impacted LD mobility, revealing the temporal effects of LD-bound DII-core. These results for DII-core coated LDs support a model for core-mediated LD localization that involves core slowing down the rate of movement of LDs until localization at the perinuclear region is accomplished where LD movement ceases. The guided localization of LDs by HCV core protein not only is essential to the viral life cycle but also poses an interesting target for the development of antiviral strategies against HCV.

  2. RNA chaperoning and intrinsic disorder in the core proteins of Flaviviridae.

    Science.gov (United States)

    Ivanyi-Nagy, Roland; Lavergne, Jean-Pierre; Gabus, Caroline; Ficheux, Damien; Darlix, Jean-Luc

    2008-02-01

    RNA chaperone proteins are essential partners of RNA in living organisms and viruses. They are thought to assist in the correct folding and structural rearrangements of RNA molecules by resolving misfolded RNA species in an ATP-independent manner. RNA chaperoning is probably an entropy-driven process, mediated by the coupled binding and folding of intrinsically disordered protein regions and the kinetically trapped RNA. Previously, we have shown that the core protein of hepatitis C virus (HCV) is a potent RNA chaperone that can drive profound structural modifications of HCV RNA in vitro. We now examined the RNA chaperone activity and the disordered nature of core proteins from different Flaviviridae genera, namely that of HCV, GBV-B (GB virus B), WNV (West Nile virus) and BVDV (bovine viral diarrhoea virus). Despite low-sequence similarities, all four proteins demonstrated general nucleic acid annealing and RNA chaperone activities. Furthermore, heat resistance of core proteins, as well as far-UV circular dichroism spectroscopy suggested that a well-defined 3D protein structure is not necessary for core-induced RNA structural rearrangements. These data provide evidence that RNA chaperoning-possibly mediated by intrinsically disordered protein segments-is conserved in Flaviviridae core proteins. Thus, besides nucleocapsid formation, core proteins may function in RNA structural rearrangements taking place during virus replication.

  3. HCV core protein promotes hepatocyte proliferation and chemoresistance by inhibiting NR4A1

    Energy Technology Data Exchange (ETDEWEB)

    Tan, Yongsheng, E-mail: yongshengtanwhu@126.com; Li, Yan, E-mail: liyansd2@163.com

    2015-10-23

    This study investigated the effect of HCV core protein on the proliferation of hepatocytes and hepatocellular carcinoma cells (HCC), the influence of HCV core protein on HCC apoptosis induced by the chemotherapeutic agent cisplatin, and the mechanism through which HCV core protein acts as a potential oncoprotein in HCV-related HCC by measuring the levels of NR4A1 and Runt-related transcription factor 3 (RUNX3), which are associated with tumor suppression and chemotherapy resistance. In the present study, PcDNA3.1-core and RUNX3 siRNA were transfected into LO2 and HepG2 cells using Lipofectamine 2000. LO2-core, HepG2-core, LO2-RUNX3 {sup low} and control cells were treated with different concentrations of cisplatin for 72 h, and cell proliferation and apoptosis were assayed using the CellTiter 96{sup ®}Aqueous Non-Radioactive Cell Proliferation Assay Kit. Western blot and real time PCR analyses were used to detect NR4A1, RUNX3, smad7, Cyclin D1 and BAX. Confocal microscopy was used to determine the levels of NR4A1 in HepG2 and HepG2-core cells. The growth rate of HepG2-core cells was considerably greater than that of HepG2 cells. HCV core protein increased the expression of cyclin D1 and decreased the expressions of NR4A1 and RUNX3. In LO2 – RUNX3 {sup low}, the rate of cell proliferation and the level of cisplatin resistance were the same as in the LO2 -core. These results suggest that HCV core protein decreases the sensitivity of hepatocytes to cisplatin by inhibiting the expression of NR4A1 and promoting the expression of smad7, which negatively regulates the TGF-β pathway. This effect results in down regulation of RUNX3, a target of the TGF-β pathway. Taken together, these findings indicate that in hepatocytes, HCV core protein increases drug resistance and inhibits cell apoptosis by inhibiting the expressions of NR4A1 and RUNX3. - Highlights: • HCV core protein inhibits HepG2 cell sensitivity to cisplatin. • Core expression in HepG2 decreases

  4. HCV core protein promotes hepatocyte proliferation and chemoresistance by inhibiting NR4A1

    International Nuclear Information System (INIS)

    Tan, Yongsheng; Li, Yan

    2015-01-01

    This study investigated the effect of HCV core protein on the proliferation of hepatocytes and hepatocellular carcinoma cells (HCC), the influence of HCV core protein on HCC apoptosis induced by the chemotherapeutic agent cisplatin, and the mechanism through which HCV core protein acts as a potential oncoprotein in HCV-related HCC by measuring the levels of NR4A1 and Runt-related transcription factor 3 (RUNX3), which are associated with tumor suppression and chemotherapy resistance. In the present study, PcDNA3.1-core and RUNX3 siRNA were transfected into LO2 and HepG2 cells using Lipofectamine 2000. LO2-core, HepG2-core, LO2-RUNX3 "l"o"w and control cells were treated with different concentrations of cisplatin for 72 h, and cell proliferation and apoptosis were assayed using the CellTiter 96"®Aqueous Non-Radioactive Cell Proliferation Assay Kit. Western blot and real time PCR analyses were used to detect NR4A1, RUNX3, smad7, Cyclin D1 and BAX. Confocal microscopy was used to determine the levels of NR4A1 in HepG2 and HepG2-core cells. The growth rate of HepG2-core cells was considerably greater than that of HepG2 cells. HCV core protein increased the expression of cyclin D1 and decreased the expressions of NR4A1 and RUNX3. In LO2 – RUNX3 "l"o"w, the rate of cell proliferation and the level of cisplatin resistance were the same as in the LO2 -core. These results suggest that HCV core protein decreases the sensitivity of hepatocytes to cisplatin by inhibiting the expression of NR4A1 and promoting the expression of smad7, which negatively regulates the TGF-β pathway. This effect results in down regulation of RUNX3, a target of the TGF-β pathway. Taken together, these findings indicate that in hepatocytes, HCV core protein increases drug resistance and inhibits cell apoptosis by inhibiting the expressions of NR4A1 and RUNX3. - Highlights: • HCV core protein inhibits HepG2 cell sensitivity to cisplatin. • Core expression in HepG2 decreases expression of NR4A1

  5. Homogeneous protein analysis by magnetic core-shell nanorod probes

    KAUST Repository

    Schrittwieser, Stefan; Pelaz, Beatriz; Parak, Wolfgang J; Lentijo Mozo, Sergio; Soulantica, Katerina; Dieckhoff, Jan; Ludwig, Frank; Altantzis, Thomas; Bals, Sara; Schotter, Joerg

    2016-01-01

    analyte protein size. In addition, due to the locking of the optical signal to the magnetic excitation frequency, background signals are suppressed, thus allowing exclusive studies of processes at the nanoprobe surface only. We study target proteins

  6. Regulation of Lipid and Glucose Metabolism by Phosphatidylcholine Transfer Protein

    Science.gov (United States)

    Kang, Hye Won; Wei, Jie; Cohen, David E.

    2010-01-01

    Phosphatidylcholine transfer protein (PC-TP, a.k.a. StARD2) binds phosphatidylcholines and catalyzes their intermembrane transfer and exchange in vitro. The structure of PC-TP comprises a hydrophobic pocket and a well-defined head-group binding site, and its gene expression is regulated by peroxisome proliferator activated receptor α. Recent studies have revealed key regulatory roles for PC-TP in lipid and glucose metabolism. Notably, Pctp−/− mice are sensitized to insulin action and exhibit more efficient brown fat-mediated thermogenesis. PC-TP appears to limit access of fatty acids to mitochondria by stimulating the activity of thioesterase superfamily member 2, a newly characterized long-chain fatty acyl-CoA thioesterase. Because PC-TP discriminates among phosphatidylcholines within lipid bilayers, it may function as a sensor that links metabolic regulation to membrane composition. PMID:20338778

  7. The effect of HCV Core protein on the expression of miR-150

    Directory of Open Access Journals (Sweden)

    Sayad Khanizadeh

    2016-09-01

    Full Text Available Background : Hepatitis C virus (HCV is considered as one of the major pathogenic agents of chronic liver diseases. Previous studies have shown that HCV proteins can interaction with gene regulatory networks such as microRNAs. The aim of this study was to investigate the effect of HCV core protein on the expression of miR-150 in a cell culture model. Materials and Methods: Plasmids expressing full HCV core protein was transfected into Huh7 cell lines while a GFP expressing plasmid employed as negative control. Subsequently, total RNA extracted and Real-Time PCR performed to measure the expression level of miR-150 expression. Moreover, trypan blue exclusion assay was performed to investigate the effect of core protein on cell viability. Results: The gene expression analysis of miR-150 in Huh7 cells showed that endogenous HCV core protein could significantly down regulation of miR-150 when compared to GFP control plasmid and normal cells (P<0.01. Beside, core protein induced no significant proliferative or cytotoxic effects on hepatic cells as determined by trypan blue exclusion assay (P<0.05. Conclusion: Our study suggests that HCV core protein can led to down regulation of miR-150 expression. This data revealed that HCV protein interactions with cell regulatory machinery may contribute to pathogenesis of chronic liver diseases.

  8. Internalisation of hepatitis C virus core protein by human conjunctival fibroblasts.

    Science.gov (United States)

    Rajalakshmy, A R; Malathi, J; Madhavan, H N; Bhaskar, S; Iyer, G K

    2016-01-01

    Recent studies indicate that hepatitis C virus (HCV) proteins can mediate innate immune response and inflammation in conjunctival fibroblasts which contributes to the pathology of dry eye condition associated with chronic HCV infection. The present study investigates the phagocytic potential of human conjunctival fibroblasts (HCFj) for HCV core protein. HCFj cells were incubated with HCV core antigen for different periods of time, and fluorescent micrographs were taken to observe protein internalisation. HCFj cells were capable of internalising HCV core antigen within 1 h; this gives an insight into another molecular mechanism which may contribute towards HCV-associated conjunctival inflammation.

  9. Hepatitis C virus core protein potentiates proangiogenic activity of hepatocellular carcinoma cells.

    Science.gov (United States)

    Shao, Yu-Yun; Hsieh, Min-Shu; Wang, Han-Yu; Li, Yong-Shi; Lin, Hang; Hsu, Hung-Wei; Huang, Chung-Yi; Hsu, Chih-Hung; Cheng, Ann-Lii

    2017-10-17

    Increased angiogenic activity has been demonstrated in hepatitis C virus (HCV)-related hepatocellular carcinoma (HCC), but the mechanism was unclear. To study the role of HCV core protein, we used tube formation and Matrigel plug assays to assess the proangiogenic activity of an HCC cell line, HuH7, and 2 of its stable clones-HuH7-core-high and HuH7-core-low, with high and low HCV core protein expression, respectively. In both assays, HuH7-core-high and HuH7-core-low cells dose-dependently induced stronger angiogenesis than control cells. HuH7 cells with HCV core protein expression showed increased mRNA and protein expression of vascular endothelial growth factor (VEGF). VEGF inhibition by bevacizumab reduced the proangiogenic activity of HuH7-core-high cells. The promotor region of VEGF contains the binding site of activator protein-1 (AP-1). Compared with controls, HuH7-core-high cells had an increased AP-1 activity and nuclear localization of phospho-c-jun. AP-1 inhibition using either RNA knockdown or AP-1 inhibitors reduced the VEGF mRNA expression and the proangiogenic activity of HuH7-core-high cells. Among 131 tissue samples from HCC patients, HCV-related HCC revealed stronger VEGF expression than did hepatitis B virus-related HCC. In conclusion, increased VEGF expression through AP-1 activation is a crucial mechanism underlying the proangiogenic activity of the HCV core protein in HCC cells.

  10. Leucine and protein metabolism in obese Zucker rats.

    Directory of Open Access Journals (Sweden)

    Pengxiang She

    Full Text Available Branched-chain amino acids (BCAAs are circulating nutrient signals for protein accretion, however, they increase in obesity and elevations appear to be prognostic of diabetes. To understand the mechanisms whereby obesity affects BCAAs and protein metabolism, we employed metabolomics and measured rates of [1-(14C]-leucine metabolism, tissue-specific protein synthesis and branched-chain keto-acid (BCKA dehydrogenase complex (BCKDC activities. Male obese Zucker rats (11-weeks old had increased body weight (BW, 53%, liver (107% and fat (∼300%, but lower plantaris and gastrocnemius masses (-21-24%. Plasma BCAAs and BCKAs were elevated 45-69% and ∼100%, respectively, in obese rats. Processes facilitating these rises appeared to include increased dietary intake (23%, leucine (Leu turnover and proteolysis [35% per g fat free mass (FFM, urinary markers of proteolysis: 3-methylhistidine (183% and 4-hydroxyproline (766%] and decreased BCKDC per g kidney, heart, gastrocnemius and liver (-47-66%. A process disposing of circulating BCAAs, protein synthesis, was increased 23-29% by obesity in whole-body (FFM corrected, gastrocnemius and liver. Despite the observed decreases in BCKDC activities per gm tissue, rates of whole-body Leu oxidation in obese rats were 22% and 59% higher normalized to BW and FFM, respectively. Consistently, urinary concentrations of eight BCAA catabolism-derived acylcarnitines were also elevated. The unexpected increase in BCAA oxidation may be due to a substrate effect in liver. Supporting this idea, BCKAs were elevated more in liver (193-418% than plasma or muscle, and per g losses of hepatic BCKDC activities were completely offset by increased liver mass, in contrast to other tissues. In summary, our results indicate that plasma BCKAs may represent a more sensitive metabolic signature for obesity than BCAAs. Processes supporting elevated BCAA]BCKAs in the obese Zucker rat include increased dietary intake, Leu and protein

  11. Early postnatal low-protein nutrition, metabolic programming and the autonomic nervous system in adult life

    OpenAIRE

    de Oliveira, Júlio Cezar; Grassiolli, Sabrina; Gravena, Clarice; de Mathias, Paulo Cezar Freitas

    2012-01-01

    Abstract Protein restriction during lactation has been used as a rat model of metabolic programming to study the impact of perinatal malnutrition on adult metabolism. In contrast to protein restriction during fetal life, protein restriction during lactation did not appear to cause either obesity or the hallmarks of metabolic syndrome, such as hyperinsulinemia, when individuals reached adulthood. However, protein restriction provokes body underweight and hypoinsulinemia. This review is focused...

  12. Amino acid metabolism and whole-body protein turnover in lambs ...

    African Journals Online (AJOL)

    The effect of protein supplementation of a wheat straw diet on the metabolism of lysine, leucine, methionine and urea, and on whole-body protein turnover rate was investigated in lambs. The metabolism of lysine and leucine is reported elsewhere (Cronje et aI., 1992); in this paper methionine metabolism is discussed, and ...

  13. Production of biopharmaceutical proteins by yeast: Advances through metabolic engineering

    DEFF Research Database (Denmark)

    Nielsen, Jens

    2013-01-01

    Production of recombinant proteins for use as pharmaceuticals, so-called biopharmaceuticals, is a multi-billion dollar industry. Many different cell factories are used for the production of biopharmaceuticals, but the yeast Saccharomyces cerevisiae is an important cell factory as it is used for p...... production. The involvement of directed metabolic engineering through the integration of tools from genetic engineering, systems biology and mathematical modeling, is also discussed....... by yeast are human serum albumin, hepatitis vaccines and virus like particles used for vaccination against human papillomavirus. Here is given a brief overview of biopharmaceutical production by yeast and it is discussed how the secretory pathway can be engineered to ensure more efficient protein...

  14. The role of mitochondria in cellular iron-sulfur protein biogenesis and iron metabolism.

    Science.gov (United States)

    Lill, Roland; Hoffmann, Bastian; Molik, Sabine; Pierik, Antonio J; Rietzschel, Nicole; Stehling, Oliver; Uzarska, Marta A; Webert, Holger; Wilbrecht, Claudia; Mühlenhoff, Ulrich

    2012-09-01

    Mitochondria play a key role in iron metabolism in that they synthesize heme, assemble iron-sulfur (Fe/S) proteins, and participate in cellular iron regulation. Here, we review the latter two topics and their intimate connection. The mitochondrial Fe/S cluster (ISC) assembly machinery consists of 17 proteins that operate in three major steps of the maturation process. First, the cysteine desulfurase complex Nfs1-Isd11 as the sulfur donor cooperates with ferredoxin-ferredoxin reductase acting as an electron transfer chain, and frataxin to synthesize an [2Fe-2S] cluster on the scaffold protein Isu1. Second, the cluster is released from Isu1 and transferred toward apoproteins with the help of a dedicated Hsp70 chaperone system and the glutaredoxin Grx5. Finally, various specialized ISC components assist in the generation of [4Fe-4S] clusters and cluster insertion into specific target apoproteins. Functional defects of the core ISC assembly machinery are signaled to cytosolic or nuclear iron regulatory systems resulting in increased cellular iron acquisition and mitochondrial iron accumulation. In fungi, regulation is achieved by iron-responsive transcription factors controlling the expression of genes involved in iron uptake and intracellular distribution. They are assisted by cytosolic multidomain glutaredoxins which use a bound Fe/S cluster as iron sensor and additionally perform an essential role in intracellular iron delivery to target metalloproteins. In mammalian cells, the iron regulatory proteins IRP1, an Fe/S protein, and IRP2 act in a post-transcriptional fashion to adjust the cellular needs for iron. Thus, Fe/S protein biogenesis and cellular iron metabolism are tightly linked to coordinate iron supply and utilization. This article is part of a Special Issue entitled: Cell Biology of Metals. Copyright © 2012 Elsevier B.V. All rights reserved.

  15. Effect of altitude on the protein metabolism of Bolivian children

    International Nuclear Information System (INIS)

    San Miguel Simron, J.L.; Berger, J.; Spielvogel, H.; Tellez Castellon, W.; Lujan Medina, C.; Caceres, E.

    1996-01-01

    The malnutrition is prevalent and is a major problem among Bolivian children. It is caused by several interacting factors: (1) inadequate protein energy intake due to low socio-economic status; (ii) exposure to acute, repeated and chronic bacterial infections; (iii) exposure to multiple and chronic parasitic infections; (iv) high altitude of the capital, La Paz, 3600 m, with a numerous populations compared to the rest of the country. The research objectives in the first phase are: (i) determination of protein utilization with a non-invasive method using stable isotope tracer among children living at high and low altitude; (ii) determination of protein metabolism among eutrophic children without parasitic or acute bacterial infections at both altitudes; (iii) determination of protein requirement among these children. Two groups of 10 pubertal children, matched for age and sex, of same socio-economic status, eutrophic, without malnutrition, infections or intestinal parasites will be studied; the different status being arrived by anthropometric, nutritional intake, biochemical and pediatrical evaluation. For the metabolic study, stable isotopes L-[1-13C] leucine labelled casein will be used and 13 CO 2 excreted will be measured. All the basic nutritional assessment and VCO 2 measurements will be performed in Bolivia, while the samples of expired gas will be stored in Vacutainers for further analysis by isotope radio mass spectrometer (IRMS), in Clermont-Ferrand, France. The plans for future work is based on the study of the effects of the different variables and their interactions. The following will be evaluated: (i) the socio-economic status; (ii) the bacterial infections: (iii) the parasitic infections; (iv) the altitude. As published by Obert, et al., the socio-economic variable is more connected with the nutritional status than with the altitude. 12 refs, 1 fig., 1 tab

  16. Effect of altitude on the protein metabolism of Bolivian children

    Energy Technology Data Exchange (ETDEWEB)

    San Miguel Simron, J L; Berger, J; Spielvogel, H; Tellez Castellon, W; Lujan Medina, C; Caceres, E [Instituto Boliviano de Boliviano de Biologia de Altura, La Paz (Bolivia). Dept. de Nutricion; Beaufrere, B; Gachons, P; Coudert, J [Laboratoire de Nutrition Humaine, Clermont-Ferrand (France)

    1997-12-31

    The malnutrition is prevalent and is a major problem among Bolivian children. It is caused by several interacting factors: (1) inadequate protein energy intake due to low socio-economic status; (ii) exposure to acute, repeated and chronic bacterial infections; (iii) exposure to multiple and chronic parasitic infections; (iv) high altitude of the capital, La Paz, 3600 m, with a numerous populations compared to the rest of the country. The research objectives in the first phase are: (i) determination of protein utilization with a non-invasive method using stable isotope tracer among children living at high and low altitude; (ii) determination of protein metabolism among eutrophic children without parasitic or acute bacterial infections at both altitudes; (iii) determination of protein requirement among these children. Two groups of 10 pubertal children, matched for age and sex, of same socio-economic status, eutrophic, without malnutrition, infections or intestinal parasites will be studied; the different status being arrived by anthropometric, nutritional intake, biochemical and pediatrical evaluation. For the metabolic study, stable isotopes L-[1-13C] leucine labelled casein will be used and {sup 13}CO{sub 2} excreted will be measured. All the basic nutritional assessment and VCO{sub 2} measurements will be performed in Bolivia, while the samples of expired gas will be stored in Vacutainers for further analysis by isotope radio mass spectrometer (IRMS), in Clermont-Ferrand, France. The plans for future work is based on the study of the effects of the different variables and their interactions. The following will be evaluated: (i) the socio-economic status; (ii) the bacterial infections: (iii) the parasitic infections; (iv) the altitude. As published by Obert, et al., the socio-economic variable is more connected with the nutritional status than with the altitude. 12 refs, 1 fig., 1 tab.

  17. Quantitative analysis of the interaction between the envelope protein domains and the core protein of human hepatitis B virus

    International Nuclear Information System (INIS)

    Choi, Kyoung-Jae; Lim, Chun-Woo; Yoon, Moon-Young; Ahn, Byung-Yoon; Yu, Yeon Gyu

    2004-01-01

    Interaction between preformed nucleocapsids and viral envelope proteins is critical for the assembly of virus particles in infected cells. The pre-S1 and pre-S2 and cytosolic regions of the human hepatitis B virus envelope protein had been implicated in the interaction with the core protein of nucleocapsids. The binding affinities of specific subdomains of the envelope protein to the core protein were quantitatively measured by both ELISA and BIAcore assay. While a marginal binding was detected with the pre-S1 or pre-S2, the core protein showed high affinities to pre-S with apparent dissociation constants (K D app ) of 7.3 ± 0.9 and 8.2 ± 0.4 μM by ELISA and BIAcore assay, respectively. The circular dichroism analysis suggested that conformational change occurs in pre-S through interaction with core protein. These results substantiate the importance of specific envelope domains in virion assembly, and demonstrate that the interaction between viral proteins can be quantitatively measured in vitro

  18. Quantitative analysis of core fucosylation of serum proteins in liver diseases by LC-MS-MRM.

    Science.gov (United States)

    Ma, Junfeng; Sanda, Miloslav; Wei, Renhuizi; Zhang, Lihua; Goldman, Radoslav

    2018-02-07

    Aberrant core fucosylation of proteins has been linked to liver diseases. In this study, we carried out multiple reaction monitoring (MRM) quantification of core fucosylated N-glycopeptides of serum proteins partially deglycosylated by a combination of endoglycosidases (endoF1, endoF2, and endoF3). To minimize variability associated with the preparatory steps, the analysis was performed without enrichment of glycopeptides or fractionation of serum besides the nanoRP chromatography. Specifically, we quantified core fucosylation of 22 N-glycopeptides derived from 17 proteins together with protein abundance of these glycoproteins in a cohort of 45 participants (15 disease-free control, 15 fibrosis and 15 cirrhosis patients) using a multiplex nanoUPLC-MS-MRM workflow. We find increased core fucosylation of 5 glycopeptides at the stage of liver fibrosis (i.e., N630 of serotransferrin, N107 of alpha-1-antitrypsin, N253 of plasma protease C1 inhibitor, N397 of ceruloplasmin, and N86 of vitronectin), increase of additional 6 glycopeptides at the stage of cirrhosis (i.e., N138 and N762 of ceruloplasmin, N354 of clusterin, N187 of hemopexin, N71 of immunoglobulin J chain, and N127 of lumican), while the degree of core fucosylation of 10 glycopeptides did not change. Interestingly, although we observe an increase in the core fucosylation at N86 of vitronectin in liver fibrosis, core fucosylation decreases on the N169 glycopeptide of the same protein. Our results demonstrate that the changes in core fucosylation are protein and site specific during the progression of fibrotic liver disease and independent of the changes in the quantity of N-glycoproteins. It is expected that the fully optimized multiplex LC-MS-MRM assay of core fucosylated glycopeptides will be useful for the serologic assessment of the fibrosis of liver. We have quantified the difference in core fucosylation among three comparison groups (healthy control, fibrosis and cirrhosis patients) using a sensitive and

  19. Protein metabolism in malnourished children with acute lower respiratory infection

    International Nuclear Information System (INIS)

    Manary, M.; Broadhead, R.

    1996-01-01

    We studied 19 subjects and 15 controls from November 1994 to February 1995. HIV infection is common among this population and HIV testing was done by ELISA of most subjects and controls in the course of their routine clinical care. To determine how HIV infection effects protein metabolism all HIV infected subjects and controls were grouped into a third category and compared to the subjects and controls. After the HIV subgrouping we were left with 13 subjects, 13 controls, and 8 HIV positive patients. KIC enrichments were used to calculate protein synthesis and breakdown, as KIC is believed to reflect intracellular leucine concentrations. Of note in Table 2 is the KIC/Leucine ratio is consistently greater than 1, averaging 1.3 over 16 samples. This is an unexpected finding as the KIC/Leucine ratio has been shown to be constant with a value of about 0.75 over a wide range of conditions. Samples for these eight patients have been evaluated under six different GCMS conditions to verify this unexpected observation. This ratio > 1.0 has been consistently found under all of these conditions. We are not certain what biological phenomenon can explain this, but it calls into question the validity of the four compartment model upon which these calculations are based. It is not unreasonable to expect that children with kwashiorkor metabolize ketoacids differently, and this difference could account for the increased KIC/Leucine ratio. 19 refs, 4 tabs

  20. Changes in contralateral protein metabolism following unilateral sciatic nerve section

    International Nuclear Information System (INIS)

    Menendez, J.A.; Cubas, S.C.

    1990-01-01

    Changes in nerve biochemistry, anatomy, and function following injuries to the contralateral nerve have been repeatedly reported, though their significance is unknown. The most likely mechanisms for their development are either substances carried by axoplasmic flow or electrically transmitted signals. This study analyzes which mechanism underlies the development of a contralateral change in protein metabolism. The incorporation of labelled amino acids (AA) into proteins of both sciatic nerves was assessed by liquid scintillation after an unilateral section. AA were offered locally for 30 min to the distal stump of the sectioned nerves and at homologous levels of the intact contralateral nerves. At various times, from 1 to 24 h, both sciatic nerves were removed and the proteins extracted with trichloroacetic acid (TCA). An increase in incorporation was found in both nerves 14-24 h after section. No difference existed between sectioned and intact nerves, which is consistent with the contralateral effect. Lidocaine, but not colchicine, when applied previously to the nerves midway between the sectioning site and the spinal cord, inhibited the contralateral increase in AA incorporation. It is concluded that electrical signals, crossing through the spinal cord, are responsible for the development of the contralateral effect. Both the nature of the proteins and the significance of the contralateral effect are matters for speculation

  1. Kinetic parameters of protein metabolism in rats during protein-free feeding

    International Nuclear Information System (INIS)

    Krawielitzki, K.; Schadereit, R.; Wuensche, J.

    1987-01-01

    16 male rats of 100 g live weight were given 50 mg of a mixture containing 15 N-labelled amino acids as a single dose within a protein-free feeding period. Following this the 15 N excretion in feces and urine as well as the development of the 15 N excess in different organs and tissues were estimated over 3 days by slaughtering the animals within given 7 time intervals. Using a 3 pool model and the computer program for the interpretation of 15 N tracer experiments by Toewe et al. (1984), kinetic parameters such as the rate of protein synthesis, protein breakdown and the rate of reutilization were calculated. Despite a negative N balance (- 41.8 mg N/d) under protein-free conditions the protein metabolism of the rat shows high dynamics characterized by a high flux rate (225 mg N/d) and a high rate of body protein synthesis (181 mg/d). The reutilization was 85 %. Depending on time the 15 N excess in the tested organs and tissues showed significant differences and seems to demonstrate that under these conditions protein synthesis mainly takes place in the most important organs (e.g. intestinal tract, liver). Under protein-free feeding conditions protein synthesis and protein breakdown of the whole body seems to be slightly increased in comparison to N balanced feeding conditions. (author)

  2. Protein and amino acid metabolism in skeletal muscle

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Guoyao.

    1989-01-01

    Isolated chick extensor digitorum communis (EDC) muscles and, in some experiments, rat skeletal muscles were used to study a number of aspects of protein and amino acid metabolism. (1) Chick EDC muscles synthesize and release large amounts of alanine and glutamine, which indirectly obtain their amino groups from branched-chain amino acids (BCAA). (2) Acetoacetate or DL-{beta}-hydroxybutyrate (4 mM) decrease (P < 0.01) alanine synthesis and BCAA transamination in EDC muscles from 24-h fasted chicks by decreasing (P < 0.01) intracellular concentrations of pyruvate due to inhibition of glycolysis. (3) Glutamine is extensively degraded in skeletal muscles from both chicks and rats, thus challenging the traditional view that glutamine oxidation is negligible in skeletal muscle. The cytosolic glutamine aminotransferases L and K in the rat and the mitochondrial phosphate-activated glutaminase in the chick play important roles in the conversion of glutamine to {alpha}-ketoglutarate for further oxidation. (4) Although methionine has been reported to be extensively transaminated in rat skeletal muscle preparations in the absence of other amino acids, transamination of methionine is absent or negligible in chick and rat skeletal muscles in the presence of physiological concentrations of amino acids. (5) Glutamine at 1.0-15 mM increases (P < 0.01) protein synthesis ({sup 3}H-phenylalanine incorporation), and at 10.0-15.0 mM decreases (P < 0.05) protein degradation ({sup 3}H-phenylalanine release from prelabelled protein in vivo) in EDC muscles from fed chicks as compared to muscles incubated in the absence of glutamine. (6) Acetoacetate or DL-{beta}-hydroxybutyrate (4 mM) has a small but significant inhibitory effect (P < 0.05) on the rate of protein synthesis, but has no effect (P > 0.05) on the rate of protein degradation in EDC muscles from fed chicks.

  3. Metatranscriptomic analysis of diverse microbial communities reveals core metabolic pathways and microbiome-specific functionality.

    Science.gov (United States)

    Jiang, Yue; Xiong, Xuejian; Danska, Jayne; Parkinson, John

    2016-01-12

    Metatranscriptomics is emerging as a powerful technology for the functional characterization of complex microbial communities (microbiomes). Use of unbiased RNA-sequencing can reveal both the taxonomic composition and active biochemical functions of a complex microbial community. However, the lack of established reference genomes, computational tools and pipelines make analysis and interpretation of these datasets challenging. Systematic studies that compare data across microbiomes are needed to demonstrate the ability of such pipelines to deliver biologically meaningful insights on microbiome function. Here, we apply a standardized analytical pipeline to perform a comparative analysis of metatranscriptomic data from diverse microbial communities derived from mouse large intestine, cow rumen, kimchi culture, deep-sea thermal vent and permafrost. Sequence similarity searches allowed annotation of 19 to 76% of putative messenger RNA (mRNA) reads, with the highest frequency in the kimchi dataset due to its relatively low complexity and availability of closely related reference genomes. Metatranscriptomic datasets exhibited distinct taxonomic and functional signatures. From a metabolic perspective, we identified a common core of enzymes involved in amino acid, energy and nucleotide metabolism and also identified microbiome-specific pathways such as phosphonate metabolism (deep sea) and glycan degradation pathways (cow rumen). Integrating taxonomic and functional annotations within a novel visualization framework revealed the contribution of different taxa to metabolic pathways, allowing the identification of taxa that contribute unique functions. The application of a single, standard pipeline confirms that the rich taxonomic and functional diversity observed across microbiomes is not simply an artefact of different analysis pipelines but instead reflects distinct environmental influences. At the same time, our findings show how microbiome complexity and availability of

  4. Kinetic variation of protein metabolism in pregnant rats

    International Nuclear Information System (INIS)

    Kubo, Katsuharu

    1980-01-01

    Kinetic variation of nitrogen metabolism in the skeletal muscle and liver of rats during the course of pregnancy was studied by the use of 15 N-amino nitrogen during acclimatization on a protein-free diet. 15 N from 15 N-glycine given on day 1 of pregnancy decreased from the 1st to 2nd trimester in the liver, suggesting contribution to the N metabolic pool. In the muscle, the rate of 15 N showed a marked decrease in the 2nd trimester, indicating, along with an increased accumulation of the total muscular N content, N accumulation in muscle protein in the 2nd trimester and promoted decomposition of mobiler muscular protein in the 2nd trimester. The marked decrease in the muscle 15 N content from the 2nd trimester and the decrease in the total N content in the 3rd trimester support the serious involvement of muscular N in fetal growth. The level of 15 N from 15 N-ammonium during the course of pregnancy was significantly high in the 2nd trimester and low in the 3rd. The 2nd trimester showed amino N accumulation in the muscle, and the 3rd, a decrease in N accumulation and amino N release. In regard to the kinetics of 15 N-lysine in the cell fraction, the muscular microsomes showed a high 15 N accumulation in the 2nd trimester and a voluminous release in the 3rd trimester. In contrast, the liver microsomes showed a linear decrease of 15 N up to 2nd trimester, followed by no change. (Chiba, N.)

  5. Acute Phase Proteins and Variables of Protein Metabolism in Dairy Cows during the Pre- and Postpartal Period

    Directory of Open Access Journals (Sweden)

    Cs. Tóthová

    2008-01-01

    Full Text Available The objective of the present study was to compare the concentrations of acute phase proteins and selected variables of protein metabolism in dairy cows of the Slovak Spotted breed from 4 weeks before parturition to 10 weeks after parturition. Acute phase proteins - haptoglobin (Hp and serum amyloid A (SAA - and variables of protein metabolism - total proteins, albumin, urea, creatinine, total immunoglobulins - were evaluated in blood serum. Significant differences were found in average values of the Hp and SAA concentrations in several groups during the monitored period (P P P P P P P P P < 0.001. The above mentioned results indicate that in the time around parturition there are significant changes in concentrations of acute phase proteins, as well as in the whole protein metabolism of dairy cows. These facts suggest that the postparturient period is a critical biological phase, throughout which there is the highest incidence of metabolic disorders.

  6. Hepatitis C virus core protein induces hepatic steatosis via Sirt1-dependent pathway.

    Science.gov (United States)

    Zhang, Chuanhai; Wang, Jingjing; Zhang, Hanlin; Liu, Shunai; Lee, Hyuek Jong; Jin, Wanzhu; Cheng, Jun

    2018-05-01

    Hepatic steatosis is a common feature of patients with chronic hepatitis C. Previous reports have shown that the overexpression of hepatitis C virus core-encoding sequences (hepatitis C virus genotypes 3a and 1b) significantly induces intracellular triglyceride accumulation. However, the underlying mechanism has not yet been revealed. To investigate whether Sirt1 is involved in hepatitis C virus-mediated hepatic steatosis, the overexpression of hepatitis C virus core 1b protein and Sirt1 and the knockdown of Sirt1 in HepG2 cells were performed. To confirm the results of the cellular experiment liver-specific Sirt1 KO mice with lentivirus-mediated hepatitis C virus core 1b overexpression were studied. Our results show that hepatitis C virus core 1b protein overexpression led to the accumulation of triglycerides in HepG2 cells. Notably the expression of PPARγ2 was dramatically increased at both the mRNA and protein levels by hepatitis C virus core 1b overexpression. The protein expression of Sirt1 is an upstream regulator of PPARγ2 and was also significantly increased after core 1b overexpression. In addition, the overexpression or knockdown of Sirt1 expression alone was sufficient to modulate p300-mediated PPARγ2 deacetylation. In vivo studies showed that hepatitis C virus core protein 1b-induced hepatic steatosis was attenuated in liver-specific Sirt1 KO mice by downregulation of PPARγ2 expression. Sirt1 mediates hepatitis C virus core protein 1b-induced hepatic steatosis by regulation of PPARγ2 expression. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  7. Identifying protein complex by integrating characteristic of core-attachment into dynamic PPI network.

    Directory of Open Access Journals (Sweden)

    Xianjun Shen

    Full Text Available How to identify protein complex is an important and challenging task in proteomics. It would make great contribution to our knowledge of molecular mechanism in cell life activities. However, the inherent organization and dynamic characteristic of cell system have rarely been incorporated into the existing algorithms for detecting protein complexes because of the limitation of protein-protein interaction (PPI data produced by high throughput techniques. The availability of time course gene expression profile enables us to uncover the dynamics of molecular networks and improve the detection of protein complexes. In order to achieve this goal, this paper proposes a novel algorithm DCA (Dynamic Core-Attachment. It detects protein-complex core comprising of continually expressed and highly connected proteins in dynamic PPI network, and then the protein complex is formed by including the attachments with high adhesion into the core. The integration of core-attachment feature into the dynamic PPI network is responsible for the superiority of our algorithm. DCA has been applied on two different yeast dynamic PPI networks and the experimental results show that it performs significantly better than the state-of-the-art techniques in terms of prediction accuracy, hF-measure and statistical significance in biology. In addition, the identified complexes with strong biological significance provide potential candidate complexes for biologists to validate.

  8. Mitochondrial energy metabolism is required for lifespan extension by the spastic paraplegia-associated protein spartin

    Directory of Open Access Journals (Sweden)

    Julia Ring

    2017-11-01

    Full Text Available Hereditary spastic paraplegias, a group of neurodegenerative disorders, can be caused by loss-of-function mutations in the protein spartin. However, the physiological role of spartin remains largely elusive. Here we show that heterologous expression of human or Drosophila spartin extends chronological lifespan of yeast, reducing age-associated ROS production, apoptosis, and necrosis. We demonstrate that spartin localizes to the proximity of mitochondria and physically interacts with proteins related to mitochondrial and respiratory metabolism. Interestingly, Nde1, the mitochondrial external NADH dehydrogenase, and Pda1, the core enzyme of the pyruvate dehydrogenase complex, are required for spartin-mediated cytoprotection. Furthermore, spartin interacts with the glycolysis enhancer phospo-fructo-kinase-2,6 (Pfk26 and is sufficient to complement for PFK26-deficiency at least in early aging. We conclude that mitochondria-related energy metabolism is crucial for spartin’s vital function during aging and uncover a network of specific interactors required for this function.

  9. Virus-producing cells determine the host protein profiles of HIV-1 virion cores

    Science.gov (United States)

    2012-01-01

    Background Upon HIV entry into target cells, viral cores are released and rearranged into reverse transcription complexes (RTCs), which support reverse transcription and also protect and transport viral cDNA to the site of integration. RTCs are composed of viral and cellular proteins that originate from both target and producer cells, the latter entering the target cell within the viral core. However, the proteome of HIV-1 viral cores in the context of the type of producer cells has not yet been characterized. Results We examined the proteomic profiles of the cores purified from HIV-1 NL4-3 virions assembled in Sup-T1 cells (T lymphocytes), PMA and vitamin D3 activated THP1 (model of macrophages, mMΦ), and non-activated THP1 cells (model of monocytes, mMN) and assessed potential involvement of identified proteins in the early stages of infection using gene ontology information and data from genome-wide screens on proteins important for HIV-1 replication. We identified 202 cellular proteins incorporated in the viral cores (T cells: 125, mMΦ: 110, mMN: 90) with the overlap between these sets limited to 42 proteins. The groups of RNA binding (29), DNA binding (17), cytoskeleton (15), cytoskeleton regulation (21), chaperone (18), vesicular trafficking-associated (12) and ubiquitin-proteasome pathway-associated proteins (9) were most numerous. Cores of the virions from SupT1 cells contained twice as many RNA binding proteins as cores of THP1-derived virus, whereas cores of virions from mMΦ and mMN were enriched in components of cytoskeleton and vesicular transport machinery, most probably due to differences in virion assembly pathways between these cells. Spectra of chaperones, cytoskeletal proteins and ubiquitin-proteasome pathway components were similar between viral cores from different cell types, whereas DNA-binding and especially RNA-binding proteins were highly diverse. Western blot analysis showed that within the group of overlapping proteins, the level of

  10. Identification of Protein Complexes Using Weighted PageRank-Nibble Algorithm and Core-Attachment Structure.

    Science.gov (United States)

    Peng, Wei; Wang, Jianxin; Zhao, Bihai; Wang, Lusheng

    2015-01-01

    Protein complexes play a significant role in understanding the underlying mechanism of most cellular functions. Recently, many researchers have explored computational methods to identify protein complexes from protein-protein interaction (PPI) networks. One group of researchers focus on detecting local dense subgraphs which correspond to protein complexes by considering local neighbors. The drawback of this kind of approach is that the global information of the networks is ignored. Some methods such as Markov Clustering algorithm (MCL), PageRank-Nibble are proposed to find protein complexes based on random walk technique which can exploit the global structure of networks. However, these methods ignore the inherent core-attachment structure of protein complexes and treat adjacent node equally. In this paper, we design a weighted PageRank-Nibble algorithm which assigns each adjacent node with different probability, and propose a novel method named WPNCA to detect protein complex from PPI networks by using weighted PageRank-Nibble algorithm and core-attachment structure. Firstly, WPNCA partitions the PPI networks into multiple dense clusters by using weighted PageRank-Nibble algorithm. Then the cores of these clusters are detected and the rest of proteins in the clusters will be selected as attachments to form the final predicted protein complexes. The experiments on yeast data show that WPNCA outperforms the existing methods in terms of both accuracy and p-value. The software for WPNCA is available at "http://netlab.csu.edu.cn/bioinfomatics/weipeng/WPNCA/download.html".

  11. Central regulation of metabolism by protein tyrosine phosphatases

    Directory of Open Access Journals (Sweden)

    Ryan eTsou

    2013-01-01

    Full Text Available Protein tyrosine phosphatases (PTPs are important regulators of intracellular signaling pathways via the dephosphorylation of phosphotyrosyl residues on various receptor and non-receptor substrates. The phosphorylation state of central nervous system (CNS signaling components underlies the molecular mechanisms of a variety of physiological functions including the control of energy balance and glucose homeostasis. In this review, we summarize the current evidence implicating PTPs as central regulators of metabolism, specifically highlighting their interactions with the neuronal leptin and insulin signaling pathways. We discuss the role of a number of PTPs (PTP1B, SHP2, TCPTP, RPTPe, and PTEN, reviewing the findings from genetic mouse models and in vitro studies which highlight these phosphatases as key central regulators of energy homeostasis.

  12. Uncoupling proteins, dietary fat and the metabolic syndrome

    Directory of Open Access Journals (Sweden)

    Warden Craig H

    2006-09-01

    Full Text Available Abstract There has been intense interest in defining the functions of UCP2 and UCP3 during the nine years since the cloning of these UCP1 homologues. Current data suggest that both UCP2 and UCP3 proteins share some features with UCP1, such as the ability to reduce mitochondrial membrane potential, but they also have distinctly different physiological roles. Human genetic studies consistently demonstrate the effect of UCP2 alleles on type-2 diabetes. Less clear is whether UCP2 alleles influence body weight or body mass index (BMI with many studies showing a positive effect while others do not. There is strong evidence that both UCP2 and UCP3 protect against mitochondrial oxidative damage by reducing the production of reactive oxygen species. The evidence that UCP2 protein is a negative regulator of insulin secretion by pancreatic β-cells is also strong: increased UCP2 decreases glucose stimulated insulin secretion ultimately leading to β-cell dysfunction. UCP2 is also neuroprotective, reducing oxidative stress in neurons. UCP3 may also transport fatty acids out of mitochondria thereby protecting the mitochondria from fatty acid anions or peroxides. Current data suggest that UCP2 plays a role in the metabolic syndrome through down-regulation of insulin secretion and development of type-2 diabetes. However, UCP2 may protect against atherosclerosis through reduction of oxidative stress and both UCP2 and UCP3 may protect against obesity. Thus, these UCP1 homologues may both contribute to and protect from the markers of the metabolic syndrome.

  13. Metabolic syndrome and C-reactive protein in bank employees

    Directory of Open Access Journals (Sweden)

    Cattafesta M

    2016-05-01

    Full Text Available Monica Cattafesta,1 Nazaré Souza Bissoli,2 Luciane Bresciani Salaroli,1,31Postgraduate Program in Nutrition and Health, 2Postgraduate Program in Physiological Sciences, 3Postgraduate Program in Public Health, Department of Health Integrated Education, Federal University of Espírito Santo, Vitória, Espírito Santo, Brazil Background: The ultrasensitive C-reactive protein (us-CRP is used for the diagnosis of cardiovascular disease, but it is not well described as a marker for the diagnosis of metabolic syndrome (MS. Methods: An observational and transversal study of bank employees evaluated anthropometric, hemodynamic, and biochemical data. CRP values were determined using commercial kits from Roche Diagnostics Ltd, and MS criteria were analyzed according to National Cholesterol Education Program’s – Adult Treatment Panel III (NCEP/ATP III. Results: A total of 88 individuals had MS, and 77.3% (n=68 of these showed alterations of us-CRP (P=0.0001, confidence interval [CI] 0.11–0.34. Individuals with MS had higher mean values of us-CRP in global measures (P=0.0001 and stratified by sex (P=0.004 than individuals without the syndrome. This marker exhibited significant differences with varying criteria for MS, such as waist circumference (P=0.0001, triglycerides (P=0.002, and diastolic blood pressure (P=0.007, and the highest levels of us-CRP were found in individuals with more MS criteria. Conclusion: us-CRP was strongly associated with the presence of MS and MS criteria in this group of workers. us-CRP is a useful and effective marker for identifying the development of MS and may be used as a reference in routine care. Keywords: C-reactive protein, bank employees, metabolic syndrome, inflammation mediators, occupational health

  14. HCV Core Protein Uses Multiple Mechanisms to Induce Oxidative Stress in Human Hepatoma Huh7 Cells

    Science.gov (United States)

    Ivanov, Alexander V.; Smirnova, Olga A.; Petrushanko, Irina Y.; Ivanova, Olga N.; Karpenko, Inna L.; Alekseeva, Ekaterina; Sominskaya, Irina; Makarov, Alexander A.; Bartosch, Birke; Kochetkov, Sergey N.; Isaguliants, Maria G.

    2015-01-01

    Hepatitis C virus (HCV) infection is accompanied by the induction of oxidative stress, mediated by several virus proteins, the most prominent being the nucleocapsid protein (HCV core). Here, using the truncated forms of HCV core, we have delineated several mechanisms by which it induces the oxidative stress. The N-terminal 36 amino acids of HCV core induced TGFβ1-dependent expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidases 1 and 4, both of which independently contributed to the production of reactive oxygen species (ROS). The same fragment also induced the expression of cyclo-oxygenase 2, which, however, made no input into ROS production. Amino acids 37–191 of HCV core up-regulated the transcription of a ROS generating enzyme cytochrome P450 2E1. Furthermore, the same fragment induced the expression of endoplasmic reticulum oxidoreductin 1α. The latter triggered efflux of Ca2+ from ER to mitochondria via mitochondrial Ca2+ uniporter, leading to generation of superoxide anions, and possibly also H2O2. Suppression of any of these pathways in cells expressing the full-length core protein led to a partial inhibition of ROS production. Thus, HCV core causes oxidative stress via several independent pathways, each mediated by a distinct region of the protein. PMID:26035647

  15. Study of protein and metabolic profile of sugarcane workers

    Energy Technology Data Exchange (ETDEWEB)

    Polachini, G.M.; Tajara, E.H. [Faculdade de Medicina de Sao Jose do Rio Preto (FAMERP), SP (Brazil); Santos, U.P. [Universidade de Sao Paulo (USP), SP (Brazil); Zeri, A.C.M.; Paes Leme, A.F. [Laboratorio Nacional de Luz Sincrotron (LNLS), Campinas, SP (Brazil)

    2012-07-01

    Full text: The National Alcohol Program (Proalcool) is a successful Brazilian renewable fuel initiative aiming to reduce the country's oil dependence. Producing ethanol from sugar cane, the program has shown positive results although accompanied by potential damage. The environmental impact mainly derives from the particulate matter emissions due to sugarcane burning, which is potentially harmful to human health. The physical activity of sugarcane workers is repetitive and exhaustive and is carried out in presence of dust, smoke and soot. The efforts by the sugarcane workers during the labor process result in increased risks of nervous, respiratory and cardiovascular system diseases and also in premature death. The aim of the present study was to investigate the effect of occupational stress on protein and metabolic profile of sugarcane workers. Forty serum samples were analyzed by 1-DE and LC MS/MS proteomic shotgun strategy and nuclear magnetic resonance (NMR). A set of proteins was found to be altered in workers after crops when compared with controls. The analysis of NMR spectra by Chenomx also showed differences in the expression of metabolites. For example, lactate displayed higher levels in control subjects than in sugarcane workers, and vice versa for the acetate. The concentrations of the two metabolites were lower after the crop, except in the case of acetate, which remained uniform in the control subjects before and after the crop. The present findings can have important application for rational designs of preventive measures and early disease detection in sugarcane workers. (author)

  16. Study of protein and metabolic profile of sugarcane workers

    International Nuclear Information System (INIS)

    Polachini, G.M.; Tajara, E.H.; Santos, U.P.; Zeri, A.C.M.; Paes Leme, A.F.

    2012-01-01

    Full text: The National Alcohol Program (Proalcool) is a successful Brazilian renewable fuel initiative aiming to reduce the country's oil dependence. Producing ethanol from sugar cane, the program has shown positive results although accompanied by potential damage. The environmental impact mainly derives from the particulate matter emissions due to sugarcane burning, which is potentially harmful to human health. The physical activity of sugarcane workers is repetitive and exhaustive and is carried out in presence of dust, smoke and soot. The efforts by the sugarcane workers during the labor process result in increased risks of nervous, respiratory and cardiovascular system diseases and also in premature death. The aim of the present study was to investigate the effect of occupational stress on protein and metabolic profile of sugarcane workers. Forty serum samples were analyzed by 1-DE and LC MS/MS proteomic shotgun strategy and nuclear magnetic resonance (NMR). A set of proteins was found to be altered in workers after crops when compared with controls. The analysis of NMR spectra by Chenomx also showed differences in the expression of metabolites. For example, lactate displayed higher levels in control subjects than in sugarcane workers, and vice versa for the acetate. The concentrations of the two metabolites were lower after the crop, except in the case of acetate, which remained uniform in the control subjects before and after the crop. The present findings can have important application for rational designs of preventive measures and early disease detection in sugarcane workers. (author)

  17. Systems biology definition of the core proteome of metabolism and expression is consistent with high-throughput data

    DEFF Research Database (Denmark)

    Yang, Laurence; Tan, Justin; O'Brien, Edward J.

    2015-01-01

    based on proteomics data. This systems biology core proteome includes 212 genes not found in previous comparative genomics-based core proteome definitions, accounts for 65% of known essential genes in E. coli, and has 78% gene function overlap with minimal genomes (Buchnera aphidicola and Mycoplasma......Finding the minimal set of gene functions needed to sustain life is of both fundamental and practical importance. Minimal gene lists have been proposed by using comparative genomics-based core proteome definitions. A definition of a core proteome that is supported by empirical data, is understood...... at the systems-level, and provides a basis for computing essential cell functions is lacking. Here, we use a systems biology-based genome-scale model of metabolism and expression to define a functional core proteome consisting of 356 gene products, accounting for 44% of the Escherichia coli proteome by mass...

  18. Lipoic acid entrains the hepatic circadian clock and lipid metabolic proteins that have been desynchronized with advanced age

    International Nuclear Information System (INIS)

    Keith, Dove; Finlay, Liam; Butler, Judy; Gómez, Luis; Smith, Eric; Moreau, Régis; Hagen, Tory

    2014-01-01

    Highlights: • 24 month old rats were supplemented with 0.2% lipoic acid in the diet for 2 weeks. • Lipoic acid shifts phase of core circadian clock proteins. • Lipoic acid corrects age-induced desynchronized lipid metabolism rhythms. - Abstract: It is well established that lipid metabolism is controlled, in part, by circadian clocks. However, circadian clocks lose temporal precision with age and correlates with elevated incidence in dyslipidemia and metabolic syndrome in older adults. Because our lab has shown that lipoic acid (LA) improves lipid homeostasis in aged animals, we hypothesized that LA affects the circadian clock to achieve these results. We fed 24 month old male F344 rats a diet supplemented with 0.2% (w/w) LA for 2 weeks prior to sacrifice and quantified hepatic circadian clock protein levels and clock-controlled lipid metabolic enzymes. LA treatment caused a significant phase-shift in the expression patterns of the circadian clock proteins Period (Per) 2, Brain and Muscle Arnt-Like1 (BMAL1), and Reverse Erythroblastosis virus (Rev-erb) β without altering the amplitude of protein levels during the light phase of the day. LA also significantly altered the oscillatory patterns of clock-controlled proteins associated with lipid metabolism. The level of peroxisome proliferator-activated receptor (PPAR) α was significantly increased and acetyl-CoA carboxylase (ACC) and fatty acid synthase (FAS) were both significantly reduced, suggesting that the LA-supplemented aged animals are in a catabolic state. We conclude that LA remediates some of the dyslipidemic processes associated with advanced age, and this mechanism may be at least partially through entrainment of circadian clocks

  19. Lipoic acid entrains the hepatic circadian clock and lipid metabolic proteins that have been desynchronized with advanced age

    Energy Technology Data Exchange (ETDEWEB)

    Keith, Dove; Finlay, Liam; Butler, Judy [Linus Pauling Institute, Oregon State University (United States); Gómez, Luis; Smith, Eric [Linus Pauling Institute, Oregon State University (United States); Biochemistry Biophysics Department, Oregon State University (United States); Moreau, Régis [Linus Pauling Institute, Oregon State University (United States); Hagen, Tory [Linus Pauling Institute, Oregon State University (United States); Biochemistry Biophysics Department, Oregon State University (United States)

    2014-07-18

    Highlights: • 24 month old rats were supplemented with 0.2% lipoic acid in the diet for 2 weeks. • Lipoic acid shifts phase of core circadian clock proteins. • Lipoic acid corrects age-induced desynchronized lipid metabolism rhythms. - Abstract: It is well established that lipid metabolism is controlled, in part, by circadian clocks. However, circadian clocks lose temporal precision with age and correlates with elevated incidence in dyslipidemia and metabolic syndrome in older adults. Because our lab has shown that lipoic acid (LA) improves lipid homeostasis in aged animals, we hypothesized that LA affects the circadian clock to achieve these results. We fed 24 month old male F344 rats a diet supplemented with 0.2% (w/w) LA for 2 weeks prior to sacrifice and quantified hepatic circadian clock protein levels and clock-controlled lipid metabolic enzymes. LA treatment caused a significant phase-shift in the expression patterns of the circadian clock proteins Period (Per) 2, Brain and Muscle Arnt-Like1 (BMAL1), and Reverse Erythroblastosis virus (Rev-erb) β without altering the amplitude of protein levels during the light phase of the day. LA also significantly altered the oscillatory patterns of clock-controlled proteins associated with lipid metabolism. The level of peroxisome proliferator-activated receptor (PPAR) α was significantly increased and acetyl-CoA carboxylase (ACC) and fatty acid synthase (FAS) were both significantly reduced, suggesting that the LA-supplemented aged animals are in a catabolic state. We conclude that LA remediates some of the dyslipidemic processes associated with advanced age, and this mechanism may be at least partially through entrainment of circadian clocks.

  20. Protein and energy metabolism in two lines of chickens selected for growth on high or low protein diets

    DEFF Research Database (Denmark)

    Chwalibog, André; Eggum, B O; Sørensen, Peter

    1983-01-01

    Genetic adaptation was investigated in broilers selected for seven generations on a normal (A) or a low (B) protein diet. Protein and energy metabolism were studied in males from these selected lines fed on a diet of intermediate protein content. All selected birds retained more nitrogen than those...

  1. A critical role for protein degradation in the nucleus accumbens core in cocaine reward memory.

    Science.gov (United States)

    Ren, Zhen-Yu; Liu, Meng-Meng; Xue, Yan-Xue; Ding, Zeng-Bo; Xue, Li-Fen; Zhai, Suo-Di; Lu, Lin

    2013-04-01

    The intense associative memories that develop between cocaine-paired contexts and rewarding stimuli contribute to cocaine seeking and relapse. Previous studies have shown impairment in cocaine reward memories by manipulating a labile state induced by memory retrieval, but the mechanisms that underlie the destabilization of cocaine reward memory are unknown. In this study, using a Pavlovian cocaine-induced conditioned place preference (CPP) procedure in rats, we tested the contribution of ubiquitin-proteasome system-dependent protein degradation in destabilization of cocaine reward memory. First, we found that polyubiquitinated protein expression levels and polyubiquitinated N-ethylmaleimide-sensitive fusion (NSF) markedly increased 15 min after retrieval while NSF protein levels decreased 1 h after retrieval in the synaptosomal membrane fraction in the nucleus accumbens (NAc) core. We then found that infusion of the proteasome inhibitor lactacystin into the NAc core prevented the impairment of memory reconsolidation induced by the protein synthesis inhibitor anisomycin and reversed the effects of anisomycin on NSF and glutamate receptor 2 (GluR2) protein levels in the synaptosomal membrane fraction in the NAc core. We also found that lactacystin infusion into the NAc core but not into the shell immediately after extinction training sessions inhibited CPP extinction and reversed the extinction training-induced decrease in NSF and GluR2 in the synaptosomal membrane fraction in the NAc core. Finally, infusions of lactacystin by itself into the NAc core immediately after each training session or before the CPP retrieval test had no effect on the consolidation and retrieval of cocaine reward memory. These findings suggest that ubiquitin-proteasome system-dependent protein degradation is critical for retrieval-induced memory destabilization.

  2. Casein and soy protein meals differentially affect whole-body and splanchnic protein metabolism in healthy humans.

    Science.gov (United States)

    Luiking, Yvette C; Deutz, Nicolaas E P; Jäkel, Martin; Soeters, Peter B

    2005-05-01

    Dietary protein quality is considered to be dependent on the degree and velocity with which protein is digested, absorbed as amino acids, and retained in the gut as newly synthesized protein. Metabolic animal studies suggest that the quality of soy protein is inferior to that of casein protein, but confirmatory studies in humans are lacking. The study objective was to assess the quality of casein and soy protein by comparing their metabolic effects in healthy human subjects. Whole-body protein kinetics, splanchnic leucine extraction, and urea production rates were measured in the postabsorptive state and during 8-h enteral intakes of isonitrogenous [0.42 g protein/(kg body weight . 8 h)] protein-based test meals, which contained either casein (CAPM; n = 12) or soy protein (SOPM; n = 10) in 2 separate groups. Stable isotope techniques were used to study metabolic effects. With enteral food intake, protein metabolism changed from net protein breakdown to net protein synthesis. Net protein synthesis was greater in the CAPM group than in the SOPM group [52 +/- 14 and 17 +/- 14 nmol/(kg fat-free mass (FFM) . min), respectively; P CAPM (P = 0.07). Absolute splanchnic extraction of leucine was higher in the subjects that consumed CAPM [306 +/- 31 nmol/(kg FFM . min)] vs. those that consumed SOPM [235 +/- 29 nmol/(kg FFM . min); P < 0.01]. In conclusion, a significantly larger portion of soy protein is degraded to urea, whereas casein protein likely contributes to splanchnic utilization (probably protein synthesis) to a greater extent. The biological value of soy protein must be considered inferior to that of casein protein in humans.

  3. Mitochondrial iron accumulation exacerbates hepatic toxicity caused by hepatitis C virus core protein

    Energy Technology Data Exchange (ETDEWEB)

    Sekine, Shuichi; Ito, Konomi; Watanabe, Haruna; Nakano, Takafumi [Laboratory of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8675 (Japan); Moriya, Kyoji; Shintani, Yoshizumi; Fujie, Hajime; Tsutsumi, Takeya; Miyoshi, Hideyuki; Fujinaga, Hidetake; Shinzawa, Seiko; Koike, Kazuhiko [Department of Internal Medicine, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655 (Japan); Horie, Toshiharu, E-mail: t.horie@thu.ac.jp [Laboratory of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University, 1-8-1 Inohana, Chuo-ku, Chiba 260-8675 (Japan)

    2015-02-01

    Patients with long-lasting hepatitis C virus (HCV) infection are at major risk of hepatocellular carcinoma (HCC). Iron accumulation in the livers of these patients is thought to exacerbate conditions of oxidative stress. Transgenic mice that express the HCV core protein develop HCC after the steatosis stage and produce an excess of hepatic reactive oxygen species (ROS). The overproduction of ROS in the liver is the net result of HCV core protein-induced dysfunction of the mitochondrial respiratory chain. This study examined the impact of ferric nitrilacetic acid (Fe-NTA)-mediated iron overload on mitochondrial damage and ROS production in HCV core protein-expressing HepG2 (human HCC) cells (Hep39b cells). A decrease in mitochondrial membrane potential and ROS production were observed following Fe-NTA treatment. After continuous exposure to Fe-NTA for six days, cell toxicity was observed in Hep39b cells, but not in mock (vector-transfected) HepG2 cells. Moreover, mitochondrial iron ({sup 59}Fe) uptake was increased in the livers of HCV core protein-expressing transgenic mice. This increase in mitochondrial iron uptake was inhibited by Ru360, a mitochondrial Ca{sup 2+} uniporter inhibitor. Furthermore, the Fe-NTA-induced augmentation of mitochondrial dysfunction, ROS production, and cell toxicity were also inhibited by Ru360 in Hep39b cells. Taken together, these results indicate that Ca{sup 2+} uniporter-mediated mitochondrial accumulation of iron exacerbates hepatocyte toxicity caused by the HCV core protein. - Highlights: • Iron accumulation in the livers of patients with hepatitis C virus (HCV) infection is thought to exacerbate oxidative stress. • The impact of iron overload on mitochondrial damage and ROS production in HCV core protein-expressing cells were examined. • Mitochondrial iron uptake was increased in the livers of HCV core protein-expressing transgenic mice. • Ca{sup 2+} uniporter-mediated mitochondrial accumulation of iron exacerbates

  4. Protein kinases responsible for the phosphorylation of the nuclear egress core complex of human cytomegalovirus.

    Science.gov (United States)

    Sonntag, Eric; Milbradt, Jens; Svrlanska, Adriana; Strojan, Hanife; Häge, Sigrun; Kraut, Alexandra; Hesse, Anne-Marie; Amin, Bushra; Sonnewald, Uwe; Couté, Yohann; Marschall, Manfred

    2017-10-01

    Nuclear egress of herpesvirus capsids is mediated by a multi-component nuclear egress complex (NEC) assembled by a heterodimer of two essential viral core egress proteins. In the case of human cytomegalovirus (HCMV), this core NEC is defined by the interaction between the membrane-anchored pUL50 and its nuclear cofactor, pUL53. NEC protein phosphorylation is considered to be an important regulatory step, so this study focused on the respective role of viral and cellular protein kinases. Multiply phosphorylated pUL50 varieties were detected by Western blot and Phos-tag analyses as resulting from both viral and cellular kinase activities. In vitro kinase analyses demonstrated that pUL50 is a substrate of both PKCα and CDK1, while pUL53 can also be moderately phosphorylated by CDK1. The use of kinase inhibitors further illustrated the importance of distinct kinases for core NEC phosphorylation. Importantly, mass spectrometry-based proteomic analyses identified five major and nine minor sites of pUL50 phosphorylation. The functional relevance of core NEC phosphorylation was confirmed by various experimental settings, including kinase knock-down/knock-out and confocal imaging, in which it was found that (i) HCMV core NEC proteins are not phosphorylated solely by viral pUL97, but also by cellular kinases; (ii) both PKC and CDK1 phosphorylation are detectable for pUL50; (iii) no impact of PKC phosphorylation on NEC functionality has been identified so far; (iv) nonetheless, CDK1-specific phosphorylation appears to be required for functional core NEC interaction. In summary, our findings provide the first evidence that the HCMV core NEC is phosphorylated by cellular kinases, and that the complex pattern of NEC phosphorylation has functional relevance.

  5. Impact of Dietary Carbohydrate and Protein Levels on Carbohydrate Metabolism

    Science.gov (United States)

    Lasker, Denise Ann

    2009-01-01

    The goal of this dissertation was to investigate the impact of changing dietary carbohydrate (CARB) intakes within recommended dietary guidelines on metabolic outcomes specifically associated with glycemic regulations and carbohydrate metabolism. This research utilized both human and animal studies to examine changes in metabolism across a wide…

  6. The Effect of Oral Leucine on Protein Metabolism in Adolescents with Type 1 Diabetes Mellitus

    OpenAIRE

    Vardhini Desikan; Izolda Mileva; Jeremy Garlick; Andrew H. Lane; Thomas A. Wilson; Margaret A. McNurlan

    2010-01-01

    Lack of insulin results in a catabolic state in subjects with insulin-dependent diabetes mellitus which is reversed by insulin treatment. Amino acid supply, especially branched chain amino acids such as leucine, enhances protein synthesis in both animal and human studies. This small study was undertaken to assess the acute effect of supplemental leucine on protein metabolism in adolescents with type 1 diabetes. L-[1-13C] Leucine was used to assess whole-body protein metabolism in six adolesc...

  7. Metabolic responses to high protein diet in Korean elite bodybuilders with high-intensity resistance exercise

    OpenAIRE

    Kim, Hyerang; Lee, Saningun; Choue, Ryowon

    2011-01-01

    Abstract Background High protein diet has been known to cause metabolic acidosis, which is manifested by increased urinary excretion of nitrogen and calcium. Bodybuilders habitually consumed excessive dietary protein over the amounts recommended for them to promote muscle mass accretion. This study investigated the metabolic response to high protein consumption in the elite bodybuilders. Methods Eight elite Korean bodybuilders within the age from 18 to 25, mean age 21.5 ± 2.6. For data collec...

  8. Cell-free protein synthesis enabled rapid prototyping for metabolic engineering and synthetic biology

    Directory of Open Access Journals (Sweden)

    Lihong Jiang

    2018-06-01

    Full Text Available Advances in metabolic engineering and synthetic biology have facilitated the manufacturing of many valuable-added compounds and commodity chemicals using microbial cell factories in the past decade. However, due to complexity of cellular metabolism, the optimization of metabolic pathways for maximal production represents a grand challenge and an unavoidable barrier for metabolic engineering. Recently, cell-free protein synthesis system (CFPS has been emerging as an enabling alternative to address challenges in biomanufacturing. This review summarizes the recent progresses of CFPS in rapid prototyping of biosynthetic pathways and genetic circuits (biosensors to speed up design-build-test (DBT cycles of metabolic engineering and synthetic biology. Keywords: Cell-free protein synthesis, Metabolic pathway optimization, Genetic circuits, Metabolic engineering, Synthetic biology

  9. Preparation and recognition of surface molecularly imprinted core-shell microbeads for protein in aqueous solutions

    International Nuclear Information System (INIS)

    Lu Yan; Yan Changling; Gao Shuyan

    2009-01-01

    In this paper, a surface molecular imprinting technique was reported for preparing core-shell microbeads of protein imprinting, and bovine hemoglobin or bovine serum albumin were used as model proteins for studying the imprinted core-shell microbeads. 3-Aminophenylboronic acid (APBA) was polymerized onto the surface of polystyrene microbead in the presence of the protein templates to create protein-imprinted core-shell microbeads. The various samples were characterized using scanning electron microscopy (SEM), transmission electron microscopy (TEM), Raman spectroscopy, X-ray photoelectron spectroscopy (XPS) and Brunauer-Emmett-Teller (BET) methods. The effect of pH on rebinding of the template hemoglobin, the specific binding and selective recognition were studied for the imprinted microbeads. The results show that the bovine hemoglobin-imprinted core-shell microbeads were successfully created. The shell was a sort of imprinted thin films with porous structure and larger surface areas. The imprinted microbeads have good selectivity for templates and high stability. Due to the recognition sites locating at or closing to the surface, these imprinted microbeads have good property of mass-transport. Unfortunately, the imprint technology was not successfully applied to imprinting bovine serum albumin (BSA).

  10. Preparation and recognition of surface molecularly imprinted core-shell microbeads for protein in aqueous solutions

    Energy Technology Data Exchange (ETDEWEB)

    Lu Yan, E-mail: yanlu2001@sohu.com [College of Chemistry and Environmental Science, Henan Normal University, 46 Jlanshe Road, Xinxiang 453007 (China); Yan Changling; Gao Shuyan [College of Chemistry and Environmental Science, Henan Normal University, 46 Jlanshe Road, Xinxiang 453007 (China)

    2009-04-01

    In this paper, a surface molecular imprinting technique was reported for preparing core-shell microbeads of protein imprinting, and bovine hemoglobin or bovine serum albumin were used as model proteins for studying the imprinted core-shell microbeads. 3-Aminophenylboronic acid (APBA) was polymerized onto the surface of polystyrene microbead in the presence of the protein templates to create protein-imprinted core-shell microbeads. The various samples were characterized using scanning electron microscopy (SEM), transmission electron microscopy (TEM), Raman spectroscopy, X-ray photoelectron spectroscopy (XPS) and Brunauer-Emmett-Teller (BET) methods. The effect of pH on rebinding of the template hemoglobin, the specific binding and selective recognition were studied for the imprinted microbeads. The results show that the bovine hemoglobin-imprinted core-shell microbeads were successfully created. The shell was a sort of imprinted thin films with porous structure and larger surface areas. The imprinted microbeads have good selectivity for templates and high stability. Due to the recognition sites locating at or closing to the surface, these imprinted microbeads have good property of mass-transport. Unfortunately, the imprint technology was not successfully applied to imprinting bovine serum albumin (BSA).

  11. Dietary carbohydrate deprivation increases 24-hour nitrogen excretion without affecting postabsorptive hepatic or whole body protein metabolism in healthy men

    NARCIS (Netherlands)

    Bisschop, PH; de Sain-van der Velden, MGM; Stellaard, F; Kuipers, F; Meijer, AJ; Sauerwein, HP; Romijn, JA

    Because insulin is an important regulator of protein metabolism, we hypothesized that physiological modulation of insulin secretion, by means of extreme variations in dietary carbohydrate content, affects postabsorptive protein metabolism. Therefore, we studied the effects of three isocaloric diets

  12. Dietary carbohydrate deprivation increases 24-hour nitrogen excretion without affecting postabsorptive hepatic or whole body protein metabolism in healthy men

    NARCIS (Netherlands)

    Bisschop, P. H.; de Sain-van der Velden, M. G. M.; Stellaard, F.; Kuipers, F.; Meijer, A. J.; Sauerwein, H. P.; Romijn, J. A.

    2003-01-01

    Because insulin is an important regulator of protein metabolism, we hypothesized that physiological modulation of insulin secretion, by means of extreme variations in dietary carbohydrate content, affects postabsorptive protein metabolism. Therefore, we studied the effects of three isocaloric diets

  13. Metabolic Turnover of Synaptic Proteins: Kinetics, Interdependencies and Implications for Synaptic Maintenance

    Science.gov (United States)

    Cohen, Laurie D.; Zuchman, Rina; Sorokina, Oksana; Müller, Anke; Dieterich, Daniela C.; Armstrong, J. Douglas; Ziv, Tamar; Ziv, Noam E.

    2013-01-01

    Chemical synapses contain multitudes of proteins, which in common with all proteins, have finite lifetimes and therefore need to be continuously replaced. Given the huge numbers of synaptic connections typical neurons form, the demand to maintain the protein contents of these connections might be expected to place considerable metabolic demands on each neuron. Moreover, synaptic proteostasis might differ according to distance from global protein synthesis sites, the availability of distributed protein synthesis facilities, trafficking rates and synaptic protein dynamics. To date, the turnover kinetics of synaptic proteins have not been studied or analyzed systematically, and thus metabolic demands or the aforementioned relationships remain largely unknown. In the current study we used dynamic Stable Isotope Labeling with Amino acids in Cell culture (SILAC), mass spectrometry (MS), Fluorescent Non–Canonical Amino acid Tagging (FUNCAT), quantitative immunohistochemistry and bioinformatics to systematically measure the metabolic half-lives of hundreds of synaptic proteins, examine how these depend on their pre/postsynaptic affiliation or their association with particular molecular complexes, and assess the metabolic load of synaptic proteostasis. We found that nearly all synaptic proteins identified here exhibited half-lifetimes in the range of 2–5 days. Unexpectedly, metabolic turnover rates were not significantly different for presynaptic and postsynaptic proteins, or for proteins for which mRNAs are consistently found in dendrites. Some functionally or structurally related proteins exhibited very similar turnover rates, indicating that their biogenesis and degradation might be coupled, a possibility further supported by bioinformatics-based analyses. The relatively low turnover rates measured here (∼0.7% of synaptic protein content per hour) are in good agreement with imaging-based studies of synaptic protein trafficking, yet indicate that the metabolic load

  14. Characterization of the fusion core in zebrafish endogenous retroviral envelope protein

    Energy Technology Data Exchange (ETDEWEB)

    Shi, Jian [State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072 (China); State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, Hubei 430071 (China); Zhang, Huaidong [CAS Key Laboratory of Special Pathogens and Biosafety, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, Hubei 430071 (China); Gong, Rui, E-mail: gongr@wh.iov.cn [CAS Key Laboratory of Special Pathogens and Biosafety, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, Hubei 430071 (China); Xiao, Gengfu, E-mail: xiaogf@wh.iov.cn [State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, Hubei 430072 (China); State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, Hubei 430071 (China)

    2015-05-08

    Zebrafish endogenous retrovirus (ZFERV) is the unique endogenous retrovirus in zebrafish, as yet, containing intact open reading frames of its envelope protein gene in zebrafish genome. Similarly, several envelope proteins of endogenous retroviruses in human and other mammalian animal genomes (such as syncytin-1 and 2 in human, syncytin-A and B in mouse) were identified and shown to be functional in induction of cell–cell fusion involved in placental development. ZFERV envelope protein (Env) gene appears to be also functional in vivo because it is expressible. After sequence alignment, we found ZFERV Env shares similar structural profiles with syncytin and other type I viral envelopes, especially in the regions of N- and C-terminal heptad repeats (NHR and CHR) which were crucial for membrane fusion. We expressed the regions of N + C protein in the ZFERV Env (residues 459–567, including predicted NHR and CHR) to characterize the fusion core structure. We found N + C protein could form a stable coiled-coil trimer that consists of three helical NHR regions forming a central trimeric core, and three helical CHR regions packing into the grooves on the surface of the central core. The structural characterization of the fusion core revealed the possible mechanism of fusion mediated by ZFERV Env. These results gave comprehensive explanation of how the ancient virus infects the zebrafish and integrates into the genome million years ago, and showed a rational clue for discovery of physiological significance (e.g., medicate cell–cell fusion). - Highlights: • ZFERV Env shares similar structural profiles with syncytin and other type I viral envelopes. • The fusion core of ZFERV Env forms stable coiled-coil trimer including three NHRs and three CHRs. • The structural mechanism of viral entry mediated by ZFERV Env is disclosed. • The results are helpful for further discovery of physiological function of ZFERV Env in zebrafish.

  15. Deep Proteomics of Mouse Skeletal Muscle Enables Quantitation of Protein Isoforms, Metabolic Pathways, and Transcription Factors*

    Science.gov (United States)

    Deshmukh, Atul S.; Murgia, Marta; Nagaraj, Nagarjuna; Treebak, Jonas T.; Cox, Jürgen; Mann, Matthias

    2015-01-01

    Skeletal muscle constitutes 40% of individual body mass and plays vital roles in locomotion and whole-body metabolism. Proteomics of skeletal muscle is challenging because of highly abundant contractile proteins that interfere with detection of regulatory proteins. Using a state-of-the art MS workflow and a strategy to map identifications from the C2C12 cell line model to tissues, we identified a total of 10,218 proteins, including skeletal muscle specific transcription factors like myod1 and myogenin and circadian clock proteins. We obtain absolute abundances for proteins expressed in a muscle cell line and skeletal muscle, which should serve as a valuable resource. Quantitation of protein isoforms of glucose uptake signaling pathways and in glucose and lipid metabolic pathways provides a detailed metabolic map of the cell line compared with tissue. This revealed unexpectedly complex regulation of AMP-activated protein kinase and insulin signaling in muscle tissue at the level of enzyme isoforms. PMID:25616865

  16. Protein engineering for metabolic engineering: current and next-generation tools

    Science.gov (United States)

    Marcheschi, Ryan J.; Gronenberg, Luisa S.; Liao, James C.

    2014-01-01

    Protein engineering in the context of metabolic engineering is increasingly important to the field of industrial biotechnology. As the demand for biologically-produced food, fuels, chemicals, food additives, and pharmaceuticals continues to grow, the ability to design and modify proteins to accomplish new functions will be required to meet the high productivity demands for the metabolism of engineered organisms. This article reviews advances of selecting, modeling, and engineering proteins to improve or alter their activity. Some of the methods have only recently been developed for general use and are just beginning to find greater application in the metabolic engineering community. We also discuss methods of generating random and targeted diversity in proteins to generate mutant libraries for analysis. Recent uses of these techniques to alter cofactor use, produce non-natural amino acids, alcohols, and carboxylic acids, and alter organism phenotypes are presented and discussed as examples of the successful engineering of proteins for metabolic engineering purposes. PMID:23589443

  17. Analysis of core-periphery organization in protein contact networks reveals groups of structurally and functionally critical residues.

    Science.gov (United States)

    Isaac, Arnold Emerson; Sinha, Sitabhra

    2015-10-01

    The representation of proteins as networks of interacting amino acids, referred to as protein contact networks (PCN), and their subsequent analyses using graph theoretic tools, can provide novel insights into the key functional roles of specific groups of residues. We have characterized the networks corresponding to the native states of 66 proteins (belonging to different families) in terms of their core-periphery organization. The resulting hierarchical classification of the amino acid constituents of a protein arranges the residues into successive layers - having higher core order - with increasing connection density, ranging from a sparsely linked periphery to a densely intra-connected core (distinct from the earlier concept of protein core defined in terms of the three-dimensional geometry of the native state, which has least solvent accessibility). Our results show that residues in the inner cores are more conserved than those at the periphery. Underlining the functional importance of the network core, we see that the receptor sites for known ligand molecules of most proteins occur in the innermost core. Furthermore, the association of residues with structural pockets and cavities in binding or active sites increases with the core order. From mutation sensitivity analysis, we show that the probability of deleterious or intolerant mutations also increases with the core order. We also show that stabilization centre residues are in the innermost cores, suggesting that the network core is critically important in maintaining the structural stability of the protein. A publicly available Web resource for performing core-periphery analysis of any protein whose native state is known has been made available by us at http://www.imsc.res.in/ ~sitabhra/proteinKcore/index.html.

  18. Hepatitis C Virus Core Protein Modulates Endoglin (CD105) Signaling Pathway for Liver Pathogenesis.

    Science.gov (United States)

    Kwon, Young-Chan; Sasaki, Reina; Meyer, Keith; Ray, Ranjit

    2017-11-01

    Endoglin is part of the TGF-β receptor complex and has a crucial role in fibrogenesis and angiogenesis. It is also an important protein for tumor growth, survival, and cancer cell metastasis. In a previous study, we have shown that hepatitis C virus (HCV) infection induces epithelial-mesenchymal transition (EMT) state and cancer stem-like cell (CSC) properties in human hepatocytes. Our array data suggested that endoglin (CD105) mRNA is significantly upregulated in HCV-associated CSCs. In this study, we have observed increased endoglin expression on the cell surface of an HCV core-expressing hepatocellular carcinoma (HepG2) cell line or immortalized human hepatocytes (IHH) and activation of its downstream signaling molecules. The status of phospho-SMAD1/5 and the expression of inhibitor of DNA binding protein 1 (ID1) were upregulated in HCV-infected cells or viral core gene-transfected cells. Additionally, we observed upregulation of endoglin/ID1 mRNA expression in chronic HCV patient liver biopsy samples. CSC generation by HCV core protein was dependent on the endoglin signaling pathway using activin receptor-like kinase 1 (ALK1) Fc blocking peptide and endoglin small interfering RNA (siRNA). Further, follow-up from in vitro analysis suggested that the antiapoptosis Bcl2 protein, proliferation-related cyclin D1 protein, and CSC-associated Hes1, Notch1, Nanog, and Sox2 proteins are enhanced during infection or ectopic expression of HCV core protein. IMPORTANCE Endoglin plays a crucial role in fibrogenesis and angiogenesis and is an important protein for tumor growth, survival, and cancer cell metastasis. Endoglin enhances ALK1-SMAD1/5 signaling in different cell types, leading to increased proliferation and migration responses. We have observed endoglin expression on the HCV core-expressing cell surface of human hepatocyte origin and activation of phospho-SMAD1/5 and ID1 downstream signaling molecules. ID1 protein plays a role in CSC properties, and we found that

  19. Simultaneous inhibition of aberrant cancer kinome using rationally designed polymer-protein core-shell nanomedicine.

    Science.gov (United States)

    Chandran, Parwathy; Gupta, Neha; Retnakumari, Archana Payickattu; Malarvizhi, Giridharan Loghanathan; Keechilat, Pavithran; Nair, Shantikumar; Koyakutty, Manzoor

    2013-11-01

    Simultaneous inhibition of deregulated cancer kinome using rationally designed nanomedicine is an advanced therapeutic approach. Herein, we have developed a polymer-protein core-shell nanomedicine to inhibit critically aberrant pro-survival kinases (mTOR, MAPK and STAT5) in primitive (CD34(+)/CD38(-)) Acute Myeloid Leukemia (AML) cells. The nanomedicine consists of poly-lactide-co-glycolide core (~250 nm) loaded with mTOR inhibitor, everolimus, and albumin shell (~25 nm thick) loaded with MAPK/STAT5 inhibitor, sorafenib and the whole construct was surface conjugated with monoclonal antibody against CD33 receptor overexpressed in AML. Electron microscopy confirmed formation of core-shell nanostructure (~290 nm) and flow cytometry and confocal studies showed enhanced cellular uptake of targeted nanomedicine. Simultaneous inhibition of critical kinases causing synergistic lethality against leukemic cells, without affecting healthy blood cells, was demonstrated using immunoblotting, cytotoxicity and apoptosis assays. This cell receptor plus multi-kinase targeted core-shell nanomedicine was found better specific and tolerable compared to current clinical regime of cytarabine and daunorubicin. These authors demonstrate simultaneous inhibition of critical kinases causing synergistic lethality against leukemic cells, without affecting healthy blood cells by using rationally designed polymer-protein core-shell nanomedicine, provoding an advanced method to eliminate cancer cells, with the hope of future therapeutic use. Copyright © 2013 Elsevier Inc. All rights reserved.

  20. Dengue Virus Capsid Protein Binds Core Histones and Inhibits Nucleosome Formation in Human Liver Cells

    Science.gov (United States)

    Colpitts, Tonya M.; Barthel, Sebastian; Wang, Penghua; Fikrig, Erol

    2011-01-01

    Dengue virus (DENV) is a member of the Flaviviridae and a globally (re)emerging pathogen that causes serious human disease. There is no specific antiviral or vaccine for dengue virus infection. Flavivirus capsid (C) is a structural protein responsible for gathering the viral RNA into a nucleocapsid that forms the core of a mature virus particle. Flaviviral replication is known to occur in the cytoplasm yet a large portion of capsid protein localizes to the nucleus during infection. The reasons for the nuclear presences of capsid are not completely understood. Here, we expressed mature DENV C in a tandem affinity purification assay to identify potential binding partners in human liver cells. DENV C targeted the four core histones, H2A, H2B, H3 and H4. DENV C bound recombinant histones in solution and colocalized with histones in the nucleus and cytoplasm of liver cells during DENV infection. We show that DENV C acts as a histone mimic, forming heterodimers with core histones, binding DNA and disrupting nucleosome formation. We also demonstrate that DENV infection increases the amounts of core histones in livers cells, which may be a cellular response to C binding away the histone proteins. Infection with DENV additionally alters levels of H2A phosphorylation in a time-dependent manner. The interactions of C and histones add an interesting new role for the presence of C in the nucleus during DENV infection. PMID:21909430

  1. First principles design of a core bioenergetic transmembrane electron-transfer protein

    Energy Technology Data Exchange (ETDEWEB)

    Goparaju, Geetha; Fry, Bryan A.; Chobot, Sarah E.; Wiedman, Gregory; Moser, Christopher C.; Leslie Dutton, P.; Discher, Bohdana M.

    2016-05-01

    Here we describe the design, Escherichia coli expression and characterization of a simplified, adaptable and functionally transparent single chain 4-α-helix transmembrane protein frame that binds multiple heme and light activatable porphyrins. Such man-made cofactor-binding oxidoreductases, designed from first principles with minimal reference to natural protein sequences, are known as maquettes. This design is an adaptable frame aiming to uncover core engineering principles governing bioenergetic transmembrane electron-transfer function and recapitulate protein archetypes proposed to represent the origins of photosynthesis. This article is part of a Special Issue entitled Biodesign for Bioenergetics — the design and engineering of electronic transfer cofactors, proteins and protein networks, edited by Ronald L. Koder and J.L. Ross Anderson.

  2. Systems biology definition of the core proteome of metabolism and expression is consistent with high-throughput data.

    Science.gov (United States)

    Yang, Laurence; Tan, Justin; O'Brien, Edward J; Monk, Jonathan M; Kim, Donghyuk; Li, Howard J; Charusanti, Pep; Ebrahim, Ali; Lloyd, Colton J; Yurkovich, James T; Du, Bin; Dräger, Andreas; Thomas, Alex; Sun, Yuekai; Saunders, Michael A; Palsson, Bernhard O

    2015-08-25

    Finding the minimal set of gene functions needed to sustain life is of both fundamental and practical importance. Minimal gene lists have been proposed by using comparative genomics-based core proteome definitions. A definition of a core proteome that is supported by empirical data, is understood at the systems-level, and provides a basis for computing essential cell functions is lacking. Here, we use a systems biology-based genome-scale model of metabolism and expression to define a functional core proteome consisting of 356 gene products, accounting for 44% of the Escherichia coli proteome by mass based on proteomics data. This systems biology core proteome includes 212 genes not found in previous comparative genomics-based core proteome definitions, accounts for 65% of known essential genes in E. coli, and has 78% gene function overlap with minimal genomes (Buchnera aphidicola and Mycoplasma genitalium). Based on transcriptomics data across environmental and genetic backgrounds, the systems biology core proteome is significantly enriched in nondifferentially expressed genes and depleted in differentially expressed genes. Compared with the noncore, core gene expression levels are also similar across genetic backgrounds (two times higher Spearman rank correlation) and exhibit significantly more complex transcriptional and posttranscriptional regulatory features (40% more transcription start sites per gene, 22% longer 5'UTR). Thus, genome-scale systems biology approaches rigorously identify a functional core proteome needed to support growth. This framework, validated by using high-throughput datasets, facilitates a mechanistic understanding of systems-level core proteome function through in silico models; it de facto defines a paleome.

  3. The origin of modern metabolic networks inferred from phylogenomic analysis of protein architecture.

    Science.gov (United States)

    Caetano-Anollés, Gustavo; Kim, Hee Shin; Mittenthal, Jay E

    2007-05-29

    Metabolism represents a complex collection of enzymatic reactions and transport processes that convert metabolites into molecules capable of supporting cellular life. Here we explore the origins and evolution of modern metabolism. Using phylogenomic information linked to the structure of metabolic enzymes, we sort out recruitment processes and discover that most enzymatic activities were associated with the nine most ancient and widely distributed protein fold architectures. An analysis of newly discovered functions showed enzymatic diversification occurred early, during the onset of the modern protein world. Most importantly, phylogenetic reconstruction exercises and other evidence suggest strongly that metabolism originated in enzymes with the P-loop hydrolase fold in nucleotide metabolism, probably in pathways linked to the purine metabolic subnetwork. Consequently, the first enzymatic takeover of an ancient biochemistry or prebiotic chemistry was related to the synthesis of nucleotides for the RNA world.

  4. Study of Stationary Phase Metabolism Via Isotopomer Analysis of Amino Acids from an Isolated Protein

    Energy Technology Data Exchange (ETDEWEB)

    Shaikh, AfshanS.; Tang, YinjieJ.; Mukhopadhyay, Aindrila; Martin, Hector Garcia; Gin, Jennifer; Benke, Peter; Keasling, Jay D.

    2009-09-14

    Microbial production of many commercially important secondary metabolites occurs during stationary phase, and methods to measure metabolic flux during this growth phase would be valuable. Metabolic flux analysis is often based on isotopomer information from proteinogenic amino acids. As such, flux analysis primarily reflects the metabolism pertinent to the growth phase during which most proteins are synthesized. To investigate central metabolism and amino acids synthesis activity during stationary phase, addition of fully 13C-labeled glucose followed by induction of green fluorescent protein (GFP) expression during stationary phase was used. Our results indicate that Escherichia coli was able to produce new proteins (i.e., GFP) in the stationary phase, and the amino acids in GFP were mostly from degraded proteins synthesized during the exponential growth phase. Among amino acid biosynthetic pathways, only those for serine, alanine, glutamate/glutamine, and aspartate/asparagine had significant activity during the stationary phase.

  5. Evaluation of the protein metabolism during hepatic coma evidenced by 15N tracer data

    International Nuclear Information System (INIS)

    Matkowitz, R.; Hartig, W.; Junghans, P.; Jung, K.; Hirschberg, K.; Bornhak, H.

    1983-01-01

    In patients in coma hepaticum as well as in pigs with experimental hepatic coma the protein metabolism was studied under conditions of parenteral application of an amino acid diet using 15 N-glycine as tracer

  6. Circulating adipocyte fatty acid-binding protein, juvenile obesity, and metabolic syndrome

    NARCIS (Netherlands)

    Krzystek-Korpacka, Malgorzata; Patryn, Eliza; Bednarz-Misa, Iwona; Mierzchala, Magdalena; Hotowy, Katarzyna; Czapinska, Elzbieta; Kustrzeba-Wojcicka, Irena; Gamian, Andrzej; Noczynska, Anna

    2011-01-01

    Adipocyte fatty acid-binding protein (A-FABP) links obesity and metabolic syndrome (MetS) and might be targeted in future therapies. Its utility as a MetS biomarker has been suggested in adults but has not been examined in children/adolescents. Our objectives were to identify metabolic parameters

  7. Effects of Quercetin Supplementation on Lipid and Protein Metabolism after Classic Boxing Training

    Science.gov (United States)

    Demirci, Nevzat

    2017-01-01

    The metabolic fitness (MF) is a component of athletes' physical conditioning. This study aims to investigate the effects of quercetin supplementation on Turkish Junior athletes' lipid and protein metabolism relating to MF after one month classic boxing training. Totally 20 voluntary junior male athletes were separated into two equal groups as the…

  8. Rift Valley fever phlebovirus NSs protein core domain structure suggests molecular basis for nuclear filaments.

    Science.gov (United States)

    Barski, Michal; Brennan, Benjamin; Miller, Ona K; Potter, Jane A; Vijayakrishnan, Swetha; Bhella, David; Naismith, James H; Elliott, Richard M; Schwarz-Linek, Ulrich

    2017-09-15

    Rift Valley fever phlebovirus (RVFV) is a clinically and economically important pathogen increasingly likely to cause widespread epidemics. RVFV virulence depends on the interferon antagonist non-structural protein (NSs), which remains poorly characterized. We identified a stable core domain of RVFV NSs (residues 83-248), and solved its crystal structure, a novel all-helical fold organized into highly ordered fibrils. A hallmark of RVFV pathology is NSs filament formation in infected cell nuclei. Recombinant virus encoding the NSs core domain induced intranuclear filaments, suggesting it contains all essential determinants for nuclear translocation and filament formation. Mutations of key crystal fibril interface residues in viruses encoding full-length NSs completely abrogated intranuclear filament formation in infected cells. We propose the fibrillar arrangement of the NSs core domain in crystals reveals the molecular basis of assembly of this key virulence factor in cell nuclei. Our findings have important implications for fundamental understanding of RVFV virulence.

  9. Functional analysis of thermostable proteins involved in carbohydrate metabolism

    NARCIS (Netherlands)

    Akerboom, A.P.

    2007-01-01

    Thermostable proteins can resist temperature stress whilst keeping their integrity and functionality. In many cases, thermostable proteins originate from hyperthermophilic microorganisms that thrive in extreme environments. These systems are generally located close to geothermal (volcanic) activity,

  10. Effect of protein provision via milk replacer or solid feed on protein metabolism in veal calves.

    Science.gov (United States)

    Berends, H; van den Borne, J J G C; Røjen, B A; Hendriks, W H; Gerrits, W J J

    2015-02-01

    The current study evaluated the effects of protein provision to calves fed a combination of solid feed (SF) and milk replacer (MR) at equal total N intake on urea recycling and N retention. Nitrogen balance traits and [(15)N2]urea kinetics were measured in 30 calves (23 wk of age, 180±3.7kg of body weight), after being exposed to the following experimental treatments for 11 wk: a low level of SF with a low N content (SF providing 12% of total N intake), a high level of SF with a low N content (SF providing 22% of total N intake), or a high level of SF with a high N content (SF providing 36% of total N intake). The SF mixture consisted of 50% concentrates, 25% corn silage, and 25% straw on a dry matter basis. Total N intake was equalized to 1.8g of N·kg of BW(-0.75)·d(-1) by adjusting N intake via MR. All calves were housed individually on metabolic cages to allow for quantification of a N balance of calves for 5 d, and for the assessment of urea recycling from [(15)N2]urea kinetics. Increasing low-N SF intake at equal total N intake resulted in a shift from urinary to fecal N excretion but did not affect protein retention (0.71g of N·kg of BW(-0.75)·d(-1)). Increasing low-N SF intake increased urea recycling but urea reused for anabolism remained unaffected. Total-tract neutral detergent fiber digestibility decreased (-9%) with increasing low-N SF intake, indicating reduced rumen fermentation. Increasing the N content of SF at equal total N intake resulted in decreased urea production, excretion, and return to ornithine cycle, and increased protein retention by 17%. This increase was likely related to an effect of energy availability on protein retention due to an increase in total-tract neutral detergent fiber digestion (>10%) and due to an increased energy supply via the MR. In conclusion, increasing low-N SF intake at the expense of N intake from MR, did not affect protein retention efficiency in calves. Increasing the N content of SF at equal total N

  11. A novel approach to preparing magnetic protein microspheres with core-shell structure

    International Nuclear Information System (INIS)

    Jiang Wei; Sun Zhendong; Li Fengsheng; Chen Kai; Liu Tianyu; Liu Jialing; Zhou Tianle; Guo Rui

    2011-01-01

    Magnetic protein microspheres with core-shell structure were prepared through a novel approach based on the sonochemical method and the emulsion solvent evaporation method. The microspheres are composed of the oleic acid and undecylenic acid modified Fe 3 O 4 cores and coated with globular bovine serum albumin (BSA). Under an optimized condition, up to 57.8 wt% of approximately 10 nm superparamagnetic Fe 3 O 4 nanoparticles could be uniformly encapsulated into the BSA microspheres with the diameter of approximately 160 nm and the high saturation magnetization of 38.5 emu/g, besides of the abundant functional groups. The possible formation mechanism of magnetic microspheres was discussed in detail. - Research Highlights: → Magnetic protein microspheres with core-shell structure were prepared through a novel approach based on the sonochemical method and the emulsion solvent evaporation method.→ The microspheres are composed of the oleic acid and undecylenic acid modified Fe 3 O 4 cores and coated with globular bovine serum albumin (BSA).→ 57.8 wt% of approximately 10 nm superparamagnetic Fe 3 O 4 nanoparticles could be uniformly encapsulated into the BSA microspheres with the diameter of approximately 160 nm and the high saturation magnetization of 38.5 emu/g, besides the abundant functional groups.

  12. A novel approach to preparing magnetic protein microspheres with core-shell structure

    Energy Technology Data Exchange (ETDEWEB)

    Jiang Wei, E-mail: climentjw@126.co [National Special Superfine Powder Engineering Research Center, Nanjing University of Science and Technology, Nanjing 210094 (China); Sun Zhendong; Li Fengsheng [National Special Superfine Powder Engineering Research Center, Nanjing University of Science and Technology, Nanjing 210094 (China); Chen Kai; Liu Tianyu; Liu Jialing [Department of Physics, Nanjing University of Science and Technology, Nanjing 210094 (China); Zhou Tianle [Department of Materials Science and Engineering, Nanjing University of Science and Technology, Nanjing 210094 (China); Guo Rui [Department of Mechanical Engineering, Nanjing University of Science and Technology, Nanjing 210094 (China)

    2011-03-15

    Magnetic protein microspheres with core-shell structure were prepared through a novel approach based on the sonochemical method and the emulsion solvent evaporation method. The microspheres are composed of the oleic acid and undecylenic acid modified Fe{sub 3}O{sub 4} cores and coated with globular bovine serum albumin (BSA). Under an optimized condition, up to 57.8 wt% of approximately 10 nm superparamagnetic Fe{sub 3}O{sub 4} nanoparticles could be uniformly encapsulated into the BSA microspheres with the diameter of approximately 160 nm and the high saturation magnetization of 38.5 emu/g, besides of the abundant functional groups. The possible formation mechanism of magnetic microspheres was discussed in detail. - Research Highlights: Magnetic protein microspheres with core-shell structure were prepared through a novel approach based on the sonochemical method and the emulsion solvent evaporation method. The microspheres are composed of the oleic acid and undecylenic acid modified Fe{sub 3}O{sub 4} cores and coated with globular bovine serum albumin (BSA). 57.8 wt% of approximately 10 nm superparamagnetic Fe{sub 3}O{sub 4} nanoparticles could be uniformly encapsulated into the BSA microspheres with the diameter of approximately 160 nm and the high saturation magnetization of 38.5 emu/g, besides the abundant functional groups.

  13. Metabolic clearance rate and urinary clearance of purified beta-core

    International Nuclear Information System (INIS)

    Wehmann, R.E.; Blithe, D.L.; Flack, M.R.; Nisula, B.C.

    1989-01-01

    We injected a highly purified preparation of the beta-core molecule, a fragment of hCG beta excreted in pregnancy urine, into five men and three women to determine its kinetic parameters, MCR, and urinary clearance. The beta-core molecule was distributed in an initial volume [1950 +/- 156 (mean +/- SEM) mL/m2 body surface area] approximately equal to the estimated plasma volume. Its disappearance was multiexponential on a semilogarithmic plot, with a rapid phase t1/2 of 3.5 +/- 0.7 min and a slow phase t1/2 of 22.4 +/- 4.2 min. The transit time (the mean time spent by a molecule of beta-core in transit) was 20.6 +/- 2.1 min. The MCR was 192.0 +/- 8.0 mL/min.m2 body surface area. About 5% of the injected dose of beta-core was excreted into the urine in the first 30 min after injection, and low levels of excretion persisted for up to 7 days. The urinary clearance rate of beta-core was 13.7 +/- 1.4 mL/min.m2, accounting for about 8% of the elimination of beta-core from the plasma. The beta-core immunoreactivity in serum and urine was characterized by gel filtration and three independent RIA systems to show that its properties were indistinguishable from those of the injected beta-core. Serum levels of beta-core in pregnant women were less than 0.2 ng/mL, while the amounts excreted in their urine were as much as 5 mg/day. Based on these clearance parameters of beta-core in normal subjects, less than 0.2% of the beta-core excreted in pregnancy urine is derived by urinary clearance of plasma beta-core. Therefore, more than 99% of the beta-core excreted in pregnancy urine is derived from beta-core in a compartment separate from plasma. In particular, these data indicate that there is relatively little placental secretion of beta-core into plasma and that placental secretion does not account for the vast majority of beta-core in pregnancy urine

  14. Vaccine delivery system for tuberculosis based on nano-sized hepatitis B virus core protein particles

    Directory of Open Access Journals (Sweden)

    Dhanasooraj D

    2013-02-01

    Full Text Available Dhananjayan Dhanasooraj, R Ajay Kumar, Sathish MundayoorMycobacterium Research Group, Rajiv Gandhi Centre for Biotechnology, Kerala, IndiaAbstract: Nano-sized hepatitis B virus core virus-like particles (HBc-VLP are suitable for uptake by antigen-presenting cells. Mycobacterium tuberculosis antigen culture filtrate protein 10 (CFP-10 is an important vaccine candidate against tuberculosis. The purified antigen shows low immune response without adjuvant and tends to have low protective efficacy. The present study is based on the assumption that expression of these proteins on HBc nanoparticles would provide higher protection when compared to the native antigen alone. The cfp-10 gene was expressed as a fusion on the major immunodominant region of HBc-VLP, and the immune response in Balb/c mice was studied and compared to pure proteins, a mixture of antigens, and fusion protein-VLP, all without using any adjuvant. The humoral, cytokine, and splenocyte cell proliferation responses suggested that the HBc-VLP bearing CFP-10 generated an antigen-specific immune response in a Th1-dependent manner. By virtue of its self-adjuvant nature and ability to form nano-sized particles, HBc-VLPs are an excellent vaccine delivery system for use with subunit protein antigens identified in the course of recent vaccine research.Keywords: Mycobacterium tuberculosis, VLP, hepatitis B virus core particle, CFP-10, self-adjuvant, vaccine delivery

  15. Effect of bacterial protein meal on protein and energy metabolism in growing chickens

    DEFF Research Database (Denmark)

    Hellwing, Anne Louise Frydendahl; Tauson, Anne-Helene; Skrede, Anders

    2006-01-01

    This experiment investigates the effect of increasing the dietary content of bacterial protein meal (BPM) on the protein and energy metabolism, and carcass chemical composition of growing chickens. Seventy-two Ross male chickens were allocated to four diets, each in three replicates with 0% (D0), 2...... (period 1), 5 chickens (period 2), and one chicken (periods 3-5). After each balance period, one chicken in each cage was killed and the carcass weight was recorded. Chemical Analyses were performed on the carcasses from periods, 1, 3, and 5. Weight gain, feed intake, and feed conversion rate were found...... to be similar for all diets. Chickens on D0 retained 1.59 g N·kg-°75·d-¹, respectively. This was probably caused by the higher nitrogen content of D0. Neither the HE (p=0.92) nor the retention of energy (P=0.88) were affected by diet. Carcass composition was similar between diets, in line with the values...

  16. Effect of altitude on protein metabolism in Bolivian children

    International Nuclear Information System (INIS)

    Beaufrere, B.; Gachon, P.; Boirie, Y.; San Miguel, J.L.; Maubois, J.L.; Coudert, J.

    1994-01-01

    Protein utilization during feeding is difficult to assess by classical tracer methodology, particularly under field conditions. We propose a new approach using the measurement of tracer recovery (expired 13 CO 2 ) after the ingestion of a single oral dose of a 13 C-leucine labelled milk protein. Protein will be obtained by infusing a cow with 13 C-leucine. The difference between the amounts of tracer given and recovered should be an index of protein utilization. Since altitude might influence protein absorption, this non-invasive method will be used in Bolivian children, living either at 3600 m (La Paz) or at sea level. (author). 14 refs

  17. Partial sequence determination of metabolically labeled radioactive proteins and peptides

    International Nuclear Information System (INIS)

    Anderson, C.W.

    1982-01-01

    The author has used the sequence analysis of radioactive proteins and peptides to approach several problems during the past few years. They, in collaboration with others, have mapped precisely several adenovirus proteins with respect to the nucleotide sequence of the adenovirus genome; identified hitherto missed proteins encoded by bacteriophage MS2 and by simian virus 40; analyzed the aminoterminal maturation of several virus proteins; determined the cleavage sites for processing of the poliovirus polyprotein; and analyzed the mechanism of frameshifting by excess normal tRNAs during cell-free protein synthesis. This chapter is designed to aid those without prior experience at protein sequence determinations. It is based primarily on the experience gained in the studies cited above, which made use of the Beckman 890 series automated protein sequencers

  18. Studies on the sulfur metabolism of cows on protein-free and low-protein feed

    Directory of Open Access Journals (Sweden)

    Eino Matikkala

    1977-09-01

    Full Text Available The influence of purified, protein-free feed with urea and ammonium salts as nitrogen sources (0-feed and of non-purified, urea-rich, low-protein feeds (ULP-feed on the sulfur metabolism of cows has been studied by determining the contents of sulfur fractions in faeces, urine, milk, blood and rumen fluid. The sulfur of 0-feed was composed entirely of inorganic sulfate. During balance trials the N:S ratio in the feed varied from 6.1 to 9.5, and the sulfur content from 0.22 to 0.31 % of the dry matter. In every trial (seven with 0-feed and two with ULP-feed, of five or seven days duration, the cows were in high-positive sulfur balance. The 0-cows excreted a greater proportion of their total sulfur output via urine than the ULP-cows. The excretion of inorganic sulfate sulfur, as a proportion of the urinary and faecal sulfur, was greater for 0-cows than for ULP- or NorP-cows (cows on normal, protein-rich feed; the opposite was the case with regard to the excretion of ester sulfate sulfur and neutral sulfur. The sulfur contents of milk and blood showed only minor inter-feed differences. The sulfate content in the rumen fluid of the 0-cow rose rapidly after the commencement of feeding and then fell quite rapidly. We conclude tentatively that in the rumen of the 0-cow hydrogen sulfide is generated so quickly that the whole of it cannot be used for the synthesis of sulfur-containing compounds, a considerable proportion of it being lost in eructations or excreted as inorganic sulfates in the urine.

  19. Thioacetamide effects on protein metabolism in the liver: lessons from isolated hepatocytes

    International Nuclear Information System (INIS)

    Cajone, F.; Bernelli-Zazzera, A.

    1984-01-01

    The effects of a short-term treatment with thioacetamide have been studied in isolated hepatocytes obtained from intoxicated rats. A technique has been developed which utilizes leucine alternatively labeled with either [ 14 C] or [ 3 H] and permits the simultaneous evaluation of protein synthesis, catabolism and secretion in the same cell during the same incubation period. The results indicate that short-term thioacetamide treatment causes an overall slowing-down of protein metabolism. Protein synthesis, however, decreases less than protein degradation and total protein secretion; albumin secretion, which is also less than normal, seems to be less compromised than total protein secretion

  20. Magnetic resonance metabolic profiling of breast cancer tissue obtained with core needle biopsy for predicting pathologic response to neoadjuvant chemotherapy.

    Directory of Open Access Journals (Sweden)

    Ji Soo Choi

    Full Text Available The purpose of this study was to determine whether metabolic profiling of core needle biopsy (CNB samples using high-resolution magic angle spinning (HR-MAS magnetic resonance spectroscopy (MRS could be used for predicting pathologic response to neoadjuvant chemotherapy (NAC in patients with locally advanced breast cancer. After institutional review board approval and informed consent were obtained, CNB tissue samples were collected from 37 malignant lesions in 37 patients before NAC treatment. The metabolic profiling of CNB samples were performed by HR-MAS MRS. Metabolic profiles were compared according to pathologic response to NAC using the Mann-Whitney test. Multivariate analysis was performed with orthogonal projections to latent structure-discriminant analysis (OPLS-DA. Various metabolites including choline-containing compounds were identified and quantified by HR-MAS MRS in all 37 breast cancer tissue samples obtained by CNB. In univariate analysis, the metabolite concentrations and metabolic ratios of CNB samples obtained with HR-MAS MRS were not significantly different between different pathologic response groups. However, there was a trend of lower levels of phosphocholine/creatine ratio and choline-containing metabolite concentrations in the pathologic complete response group compared to the non-pathologic complete response group. In multivariate analysis, the OPLS-DA models built with HR-MAS MR metabolic profiles showed visible discrimination between the pathologic response groups. This study showed OPLS-DA multivariate analysis using metabolic profiles of pretreatment CNB samples assessed by HR- MAS MRS may be used to predict pathologic response before NAC, although we did not identify the metabolite showing statistical significance in univariate analysis. Therefore, our preliminary results raise the necessity of further study on HR-MAS MR metabolic profiling of CNB samples for a large number of cancers.

  1. Integrating the protein and metabolic engineering toolkits for next-generation chemical biosynthesis.

    Science.gov (United States)

    Pirie, Christopher M; De Mey, Marjan; Jones Prather, Kristala L; Ajikumar, Parayil Kumaran

    2013-04-19

    Through microbial engineering, biosynthesis has the potential to produce thousands of chemicals used in everyday life. Metabolic engineering and synthetic biology are fields driven by the manipulation of genes, genetic regulatory systems, and enzymatic pathways for developing highly productive microbial strains. Fundamentally, it is the biochemical characteristics of the enzymes themselves that dictate flux through a biosynthetic pathway toward the product of interest. As metabolic engineers target sophisticated secondary metabolites, there has been little recognition of the reduced catalytic activity and increased substrate/product promiscuity of the corresponding enzymes compared to those of central metabolism. Thus, fine-tuning these enzymatic characteristics through protein engineering is paramount for developing high-productivity microbial strains for secondary metabolites. Here, we describe the importance of protein engineering for advancing metabolic engineering of secondary metabolism pathways. This pathway integrated enzyme optimization can enhance the collective toolkit of microbial engineering to shape the future of chemical manufacturing.

  2. Genome-scale metabolic model of Pichia pastoris with native and humanized glycosylation of recombinant proteins.

    Science.gov (United States)

    Irani, Zahra Azimzadeh; Kerkhoven, Eduard J; Shojaosadati, Seyed Abbas; Nielsen, Jens

    2016-05-01

    Pichia pastoris is used for commercial production of human therapeutic proteins, and genome-scale models of P. pastoris metabolism have been generated in the past to study the metabolism and associated protein production by this yeast. A major challenge with clinical usage of recombinant proteins produced by P. pastoris is the difference in N-glycosylation of proteins produced by humans and this yeast. However, through metabolic engineering, a P. pastoris strain capable of producing humanized N-glycosylated proteins was constructed. The current genome-scale models of P. pastoris do not address native nor humanized N-glycosylation, and we therefore developed ihGlycopastoris, an extension to the iLC915 model with both native and humanized N-glycosylation for recombinant protein production, but also an estimation of N-glycosylation of P. pastoris native proteins. This new model gives a better prediction of protein yield, demonstrates the effect of the different types of N-glycosylation of protein yield, and can be used to predict potential targets for strain improvement. The model represents a step towards a more complete description of protein production in P. pastoris, which is required for using these models to understand and optimize protein production processes. © 2015 Wiley Periodicals, Inc.

  3. Uncoupling of Metabolic Health from Longevity through Genetic Alteration of Adipose Tissue Lipid-Binding Proteins

    Directory of Open Access Journals (Sweden)

    Khanichi N. Charles

    2017-10-01

    Full Text Available Summary: Deterioration of metabolic health is a hallmark of aging and generally assumed to be detrimental to longevity. Exposure to a high-calorie diet impairs metabolism and accelerates aging; conversely, calorie restriction (CR prevents age-related metabolic diseases and extends lifespan. However, it is unclear whether preservation of metabolic health is sufficient to extend lifespan. We utilized a genetic mouse model lacking Fabp4/5 that confers protection against metabolic diseases and shares molecular and lipidomic features with CR to address this question. Fabp-deficient mice exhibit extended metabolic healthspan, with protection against insulin resistance and glucose intolerance, inflammation, deterioration of adipose tissue integrity, and fatty liver disease. Surprisingly, however, Fabp-deficient mice did not exhibit any extension of lifespan. These data indicate that extension of metabolic healthspan in the absence of CR can be uncoupled from lifespan, indicating the potential for independent drivers of these pathways, at least in laboratory mice. : Deterioration of metabolic health is a hallmark of aging and generally thought to be detrimental to longevity. Charles et al. utilize FABP-deficient mice as a model to demonstrate that the preservation of metabolic health in this model persists throughout life, even under metabolic stress, but does not increase longevity. Keywords: fatty acid binding protein, aging, calorie restriction, metabolic health, inflammation, metaflammation, diabetes, obesity, de novo lipogenesis

  4. Metabolic flux profiling of recombinant protein secreting Pichia pastoris growing on glucose:methanol mixtures

    Science.gov (United States)

    2012-01-01

    Background The methylotrophic yeast Pichia pastoris has emerged as one of the most promising yeast hosts for the production of heterologous proteins. Mixed feeds of methanol and a multicarbon source instead of methanol as sole carbon source have been shown to improve product productivities and alleviate metabolic burden derived from protein production. Nevertheless, systematic quantitative studies on the relationships between the central metabolism and recombinant protein production in P. pastoris are still rather limited, particularly when growing this yeast on mixed carbon sources, thus hampering future metabolic network engineering strategies for improved protein production. Results The metabolic flux distribution in the central metabolism of P. pastoris growing on a mixed feed of glucose and methanol was analyzed by Metabolic Flux Analysis (MFA) using 13C-NMR-derived constraints. For this purpose, we defined new flux ratios for methanol assimilation pathways in P. pastoris cells growing on glucose:methanol mixtures. By using this experimental approach, the metabolic burden caused by the overexpression and secretion of a Rhizopus oryzae lipase (Rol) in P. pastoris was further analyzed. This protein has been previously shown to trigger the unfolded protein response in P. pastoris. A series of 13C-tracer experiments were performed on aerobic chemostat cultivations with a control and two different Rol producing strains growing at a dilution rate of 0.09 h−1 using a glucose:methanol 80:20 (w/w) mix as carbon source. The MFA performed in this study reveals a significant redistristribution of carbon fluxes in the central carbon metabolism when comparing the two recombinant strains vs the control strain, reflected in increased glycolytic, TCA cycle and NADH regeneration fluxes, as well as higher methanol dissimilation rates. Conclusions Overall, a further 13C-based MFA development to characterise the central metabolism of methylotrophic yeasts when growing on mixed

  5. Metabolic flux profiling of recombinant protein secreting Pichia pastoris growing on glucose:methanol mixtures

    Directory of Open Access Journals (Sweden)

    Jordà Joel

    2012-05-01

    Full Text Available Abstract Background The methylotrophic yeast Pichia pastoris has emerged as one of the most promising yeast hosts for the production of heterologous proteins. Mixed feeds of methanol and a multicarbon source instead of methanol as sole carbon source have been shown to improve product productivities and alleviate metabolic burden derived from protein production. Nevertheless, systematic quantitative studies on the relationships between the central metabolism and recombinant protein production in P. pastoris are still rather limited, particularly when growing this yeast on mixed carbon sources, thus hampering future metabolic network engineering strategies for improved protein production. Results The metabolic flux distribution in the central metabolism of P. pastoris growing on a mixed feed of glucose and methanol was analyzed by Metabolic Flux Analysis (MFA using 13C-NMR-derived constraints. For this purpose, we defined new flux ratios for methanol assimilation pathways in P. pastoris cells growing on glucose:methanol mixtures. By using this experimental approach, the metabolic burden caused by the overexpression and secretion of a Rhizopus oryzae lipase (Rol in P. pastoris was further analyzed. This protein has been previously shown to trigger the unfolded protein response in P. pastoris. A series of 13C-tracer experiments were performed on aerobic chemostat cultivations with a control and two different Rol producing strains growing at a dilution rate of 0.09 h−1 using a glucose:methanol 80:20 (w/w mix as carbon source. The MFA performed in this study reveals a significant redistristribution of carbon fluxes in the central carbon metabolism when comparing the two recombinant strains vs the control strain, reflected in increased glycolytic, TCA cycle and NADH regeneration fluxes, as well as higher methanol dissimilation rates. Conclusions Overall, a further 13C-based MFA development to characterise the central metabolism of methylotrophic

  6. Protein metabolism of prolific ewes during late gestation and early lactation

    NARCIS (Netherlands)

    Sebek, L.B.J.

    2001-01-01

    Subject headings: protein / metabolism / ewes

    Introduction of prolific crossbred ewes and a new protein evaluation system for ruminants, the DVE/OEB system, necessitate a reconsideration of ewe feeding strategies. The objective of this thesis is to investigate the amount

  7. Deep proteomics of mouse skeletal muscle enables quantitation of protein isoforms, metabolic pathways and transcription factors

    DEFF Research Database (Denmark)

    Deshmukh, Atul S; Murgia, Marta; Nagaraja, Nagarjuna

    2015-01-01

    Skeletal muscle constitutes 40% of individual body mass and plays vital roles in locomotion and whole-body metabolism. Proteomics of skeletal muscle is challenging due to highly abundant contractile proteins that interfere with detection of regulatory proteins. Using a state-of-the art mass...

  8. Synergizing metabolic flux analysis and nucleotide sugar metabolism to understand the control of glycosylation of recombinant protein in CHO cells

    LENUS (Irish Health Repository)

    Burleigh, Susan C

    2011-10-18

    Abstract Background The glycosylation of recombinant proteins can be altered by a range of parameters including cellular metabolism, metabolic flux and the efficiency of the glycosylation process. We present an experimental set-up that allows determination of these key processes associated with the control of N-linked glycosylation of recombinant proteins. Results Chinese hamster ovary cells (CHO) were cultivated in shake flasks at 0 mM glutamine and displayed a reduced growth rate, glucose metabolism and a slower decrease in pH, when compared to other glutamine-supplemented cultures. The N-linked glycosylation of recombinant human chorionic gonadotrophin (HCG) was also altered under these conditions; the sialylation, fucosylation and antennarity decreased, while the proportion of neutral structures increased. A continuous culture set-up was subsequently used to understand the control of HCG glycosylation in the presence of varied glutamine concentrations; when glycolytic flux was reduced in the absence of glutamine, the glycosylation changes that were observed in shake flask culture were similarly detected. The intracellular content of UDP-GlcNAc was also reduced, which correlated with a decrease in sialylation and antennarity of the N-linked glycans attached to HCG. Conclusions The use of metabolic flux analysis illustrated a case of steady state multiplicity, where use of the same operating conditions at each steady state resulted in altered flux through glycolysis and the TCA cycle. This study clearly demonstrated that the control of glycoprotein microheterogeneity may be examined by use of a continuous culture system, metabolic flux analysis and assay of intracellular nucleotides. This system advances our knowledge of the relationship between metabolic flux and the glycosylation of biotherapeutics in CHO cells and will be of benefit to the bioprocessing industry.

  9. Fibroblast activation protein (FAP as a novel metabolic target

    Directory of Open Access Journals (Sweden)

    Miguel Angel Sánchez-Garrido

    2016-10-01

    Conclusions: We conclude that pharmacological inhibition of FAP enhances levels of FGF21 in obese mice to provide robust metabolic benefits not observed in lean animals, thus validating this enzyme as a novel drug target for the treatment of obesity and diabetes.

  10. Metabolic Models of Protein Allocation Call for the Kinetome

    DEFF Research Database (Denmark)

    Nilsson, Avlant; Nielsen, Jens; Palsson, Bernhard

    2017-01-01

    The flux of metabolites in the living cell depend on enzyme activities. Recently, many metabolic phenotypes have been explained by computer models that incorporate enzyme activity data. To move further, the scientific community needs to measure the kinetics of all enzymes in a systematic way....

  11. The influence of a steroid hormone and of physical exercise on protein metabolism in rats

    International Nuclear Information System (INIS)

    Menschikowski, M.; Jung, K.; Junghans, P.; Petzke, K.J.; Albrecht, V.; Akademie der Wissenschaften der DDR, Potsdam

    1989-01-01

    The influence of an anabolic steroid hormone preparation and of a physical exercise training program was studied on the nitrogen and protein metabolism in rats with the help of the 15 N tracer technique and the emission spectrometric 15 N isotope analysis. For the determination of the dynamic parameters of the protein metabolism graphic (stochastic) and computer-aided compartmental methods wer compared. Using the area method as a stochastic approach the animals showed significant differences in the protein turnover parameters under the influence of hormone treatment and (or) physical stress by swimming exercise in comparison to the controls. (author)

  12. Differential metabolic effects of casein and soy protein meals on skeletal muscle in healthy volunteers.

    Science.gov (United States)

    Luiking, Yvette C; Engelen, Mariëlle P K J; Soeters, Peter B; Boirie, Yves; Deutz, Nicolaas E P

    2011-02-01

    Dietary protein intake is known to affect whole body and interorgan protein turnover. We examined if moderate-nitrogen and carbohydrate casein and soy meals have a different effect on skeletal muscle protein and amino acid kinetics in healthy young subjects. Muscle protein and amino acid kinetics were measured in the postabsorptive state and during 4-h enteral intake of isonitrogenous [0.21 g protein/(kg body weight. 4 h)] protein-based test meals, which contained either casein (CAPM; n = 12) or soy protein (SOPM; n = 10) in 2 separate groups. Stable isotope and muscle biopsy techniques were used to study metabolic effects. The net uptake of glutamate, serine, histidine, and lysine across the leg was larger during CAPM than during SOPM intake. Muscle concentrations of glutamate, serine, histidine, glutamine, isoleucine and BCAA changed differently after CAPM and SOPM (P CAPM and SOPM, but differences in their (net) breakdown rates were not significant. Muscle protein synthesis was not different between CAPM and SOPM. Moderate-nitrogen casein and soy protein meals differently alter leg amino acid uptake without a significant difference in influencing acute muscle protein metabolism. Copyright © 2010 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

  13. Hepatitis C Virus core+1/ARF Protein Modulates the Cyclin D1/pRb Pathway and Promotes Carcinogenesis.

    Science.gov (United States)

    Moustafa, Savvina; Karakasiliotis, Ioannis; Mavromara, Penelope

    2018-05-01

    Viruses often encompass overlapping reading frames and unconventional translation mechanisms in order to maximize the output from a minimum genome and to orchestrate their timely gene expression. Hepatitis C virus (HCV) possesses such an unconventional open reading frame (ORF) within the core-coding region, encoding an additional protein, initially designated ARFP, F, or core+1. Two predominant isoforms of core+1/ARFP have been reported, core+1/L, initiating from codon 26, and core+1/S, initiating from codons 85/87 of the polyprotein coding region. The biological significance of core+1/ARFP expression remains elusive. The aim of the present study was to gain insight into the functional and pathological properties of core+1/ARFP through its interaction with the host cell, combining in vitro and in vivo approaches. Our data provide strong evidence that the core+1/ARFP of HCV-1a stimulates cell proliferation in Huh7-based cell lines expressing either core+1/S or core+1/L isoforms and in transgenic liver disease mouse models expressing core+1/S protein in a liver-specific manner. Both isoforms of core+1/ARFP increase the levels of cyclin D1 and phosphorylated Rb, thus promoting the cell cycle. In addition, core+1/S was found to enhance liver regeneration and oncogenesis in transgenic mice. The induction of the cell cycle together with increased mRNA levels of cell proliferation-related oncogenes in cells expressing the core+1/ARFP proteins argue for an oncogenic potential of these proteins and an important role in HCV-associated pathogenesis. IMPORTANCE This study sheds light on the biological importance of a unique HCV protein. We show here that core+1/ARFP of HCV-1a interacts with the host machinery, leading to acceleration of the cell cycle and enhancement of liver carcinogenesis. This pathological mechanism(s) may complement the action of other viral proteins with oncogenic properties, leading to the development of hepatocellular carcinoma. In addition, given that

  14. An α-Helical Core Encodes the Dual Functions of the Chlamydial Protein IncA*

    Science.gov (United States)

    Ronzone, Erik; Wesolowski, Jordan; Bauler, Laura D.; Bhardwaj, Anshul; Hackstadt, Ted; Paumet, Fabienne

    2014-01-01

    Chlamydia is an intracellular bacterium that establishes residence within parasitophorous compartments (inclusions) inside host cells. Chlamydial inclusions are uncoupled from the endolysosomal pathway and undergo fusion with cellular organelles and with each other. To do so, Chlamydia expresses proteins on the surface of the inclusion using a Type III secretion system. These proteins, termed Incs, are located at the interface between host and pathogen and carry out the functions necessary for Chlamydia survival. Among these Incs, IncA plays a critical role in both protecting the inclusion from lysosomal fusion and inducing the homotypic fusion of inclusions. Within IncA are two regions homologous to eukaryotic SNARE (soluble N-ethylmaleimide-sensitive factor attachment receptor) domains referred to as SNARE-like domain 1 (SLD1) and SNARE-like domain 2 (SLD2). Using a multidisciplinary approach, we have discovered the functional core of IncA that retains the ability to both inhibit SNARE-mediated fusion and promote the homotypic fusion of Chlamydia inclusions. Circular dichroism and analytical ultracentrifugation experiments show that this core region is composed almost entirely of α-helices and assembles into stable homodimers in solution. Altogether, we propose that both IncA functions are encoded in a structured core domain that encompasses SLD1 and part of SLD2. PMID:25324548

  15. An α-helical core encodes the dual functions of the chlamydial protein IncA.

    Science.gov (United States)

    Ronzone, Erik; Wesolowski, Jordan; Bauler, Laura D; Bhardwaj, Anshul; Hackstadt, Ted; Paumet, Fabienne

    2014-11-28

    Chlamydia is an intracellular bacterium that establishes residence within parasitophorous compartments (inclusions) inside host cells. Chlamydial inclusions are uncoupled from the endolysosomal pathway and undergo fusion with cellular organelles and with each other. To do so, Chlamydia expresses proteins on the surface of the inclusion using a Type III secretion system. These proteins, termed Incs, are located at the interface between host and pathogen and carry out the functions necessary for Chlamydia survival. Among these Incs, IncA plays a critical role in both protecting the inclusion from lysosomal fusion and inducing the homotypic fusion of inclusions. Within IncA are two regions homologous to eukaryotic SNARE (soluble N-ethylmaleimide-sensitive factor attachment receptor) domains referred to as SNARE-like domain 1 (SLD1) and SNARE-like domain 2 (SLD2). Using a multidisciplinary approach, we have discovered the functional core of IncA that retains the ability to both inhibit SNARE-mediated fusion and promote the homotypic fusion of Chlamydia inclusions. Circular dichroism and analytical ultracentrifugation experiments show that this core region is composed almost entirely of α-helices and assembles into stable homodimers in solution. Altogether, we propose that both IncA functions are encoded in a structured core domain that encompasses SLD1 and part of SLD2. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  16. Investigation of protein and lipid metabolism in thyroid pathology using whole-body radiometry

    International Nuclear Information System (INIS)

    Gorobets, V.F.; Matveenko, E.G.

    1987-01-01

    Radiometry of the whole body and its organs was employed to study certain aspects of protein-aminoacid and lipid metabolism in patients with thyroid diseases. Metabolism of human serum 131 I-albumin was studied in 12 patients with neurocirculatory dystonia, in 13 patients with diffuse toxic goiter (in 10 before and after drug therapy) and in 9 controls. 75 Se-methionine aminoacid metabolism was investigated in 9 patients with toxic thyroid adenoma and in 13 controls. The body cell mass was determined in 82 patients with thyrotoxicosis by a measurable amount of 40 K. These data were compared with those of 249 healthy persons. An increase in catabolism of labeled albumin, intensification of labeled methionine metabolism at the tissue level, signs of a decrease in the total amount of metabolic albumin in the body were revealed. Intensification of protein metabolism resulted in a decrease in the body cell mass of these patients. After adequate therapy the above indices of protein metabolism in patients with thyrotoxicosis returned to normal. The assimilation of fatty acids and neutral fat was disturbed both in thyrotoxicosis and hypothyroidism

  17. PanCoreGen - Profiling, detecting, annotating protein-coding genes in microbial genomes.

    Science.gov (United States)

    Paul, Sandip; Bhardwaj, Archana; Bag, Sumit K; Sokurenko, Evgeni V; Chattopadhyay, Sujay

    2015-12-01

    A large amount of genomic data, especially from multiple isolates of a single species, has opened new vistas for microbial genomics analysis. Analyzing the pan-genome (i.e. the sum of genetic repertoire) of microbial species is crucial in understanding the dynamics of molecular evolution, where virulence evolution is of major interest. Here we present PanCoreGen - a standalone application for pan- and core-genomic profiling of microbial protein-coding genes. PanCoreGen overcomes key limitations of the existing pan-genomic analysis tools, and develops an integrated annotation-structure for a species-specific pan-genomic profile. It provides important new features for annotating draft genomes/contigs and detecting unidentified genes in annotated genomes. It also generates user-defined group-specific datasets within the pan-genome. Interestingly, analyzing an example-set of Salmonella genomes, we detect potential footprints of adaptive convergence of horizontally transferred genes in two human-restricted pathogenic serovars - Typhi and Paratyphi A. Overall, PanCoreGen represents a state-of-the-art tool for microbial phylogenomics and pathogenomics study. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. PanCoreGen – profiling, detecting, annotating protein-coding genes in microbial genomes

    Science.gov (United States)

    Bhardwaj, Archana; Bag, Sumit K; Sokurenko, Evgeni V.

    2015-01-01

    A large amount of genomic data, especially from multiple isolates of a single species, has opened new vistas for microbial genomics analysis. Analyzing pan-genome (i.e. the sum of genetic repertoire) of microbial species is crucial in understanding the dynamics of molecular evolution, where virulence evolution is of major interest. Here we present PanCoreGen – a standalone application for pan- and core-genomic profiling of microbial protein-coding genes. PanCoreGen overcomes key limitations of the existing pan-genomic analysis tools, and develops an integrated annotation-structure for species-specific pan-genomic profile. It provides important new features for annotating draft genomes/contigs and detecting unidentified genes in annotated genomes. It also generates user-defined group-specific datasets within the pan-genome. Interestingly, analyzing an example-set of Salmonella genomes, we detect potential footprints of adaptive convergence of horizontally transferred genes in two human-restricted pathogenic serovars – Typhi and Paratyphi A. Overall, PanCoreGen represents a state-of-the-art tool for microbial phylogenomics and pathogenomics study. PMID:26456591

  19. Effect of long-term refeeding on protein metabolism in patients with cirrhosis of the liver

    DEFF Research Database (Denmark)

    Kondrup, J; Nielsen, K; Juul, A

    1997-01-01

    , protein requirement and protein utilization were investigated further by measuring protein synthesis and degradation. In two separate studies, five or six patients with cirrhosis of the liver were refed on a balanced diet for an average of 2 or 4 weeks. Protein and energy intakes were doubled in both...... studies. Initial and final whole-body protein metabolism was measured in the fed state by primed continuous [15N]glycine infusion. Refeeding caused a statistically significant increase of about 30% in protein synthesis in both studies while protein degradation was only slightly affected. The increase...... in protein synthesis was associated with significant increases in plasma concentrations of total amino acids (25%), leucine (58%), isoleucine (82%), valine (72%), proline (48%) and triiodothyronine (27%) while insulin, growth hormone, insulin-like growth factor (IGF)-I and IGF-binding protein-3 were...

  20. The Protein Cost of Metabolic Fluxes: Prediction from Enzymatic Rate Laws and Cost Minimization.

    Directory of Open Access Journals (Sweden)

    Elad Noor

    2016-11-01

    Full Text Available Bacterial growth depends crucially on metabolic fluxes, which are limited by the cell's capacity to maintain metabolic enzymes. The necessary enzyme amount per unit flux is a major determinant of metabolic strategies both in evolution and bioengineering. It depends on enzyme parameters (such as kcat and KM constants, but also on metabolite concentrations. Moreover, similar amounts of different enzymes might incur different costs for the cell, depending on enzyme-specific properties such as protein size and half-life. Here, we developed enzyme cost minimization (ECM, a scalable method for computing enzyme amounts that support a given metabolic flux at a minimal protein cost. The complex interplay of enzyme and metabolite concentrations, e.g. through thermodynamic driving forces and enzyme saturation, would make it hard to solve this optimization problem directly. By treating enzyme cost as a function of metabolite levels, we formulated ECM as a numerically tractable, convex optimization problem. Its tiered approach allows for building models at different levels of detail, depending on the amount of available data. Validating our method with measured metabolite and protein levels in E. coli central metabolism, we found typical prediction fold errors of 4.1 and 2.6, respectively, for the two kinds of data. This result from the cost-optimized metabolic state is significantly better than randomly sampled metabolite profiles, supporting the hypothesis that enzyme cost is important for the fitness of E. coli. ECM can be used to predict enzyme levels and protein cost in natural and engineered pathways, and could be a valuable computational tool to assist metabolic engineering projects. Furthermore, it establishes a direct connection between protein cost and thermodynamics, and provides a physically plausible and computationally tractable way to include enzyme kinetics into constraint-based metabolic models, where kinetics have usually been ignored or

  1. Early postnatal low-protein nutrition, metabolic programming and the autonomic nervous system in adult life

    Directory of Open Access Journals (Sweden)

    de Oliveira Júlio

    2012-09-01

    Full Text Available Abstract Protein restriction during lactation has been used as a rat model of metabolic programming to study the impact of perinatal malnutrition on adult metabolism. In contrast to protein restriction during fetal life, protein restriction during lactation did not appear to cause either obesity or the hallmarks of metabolic syndrome, such as hyperinsulinemia, when individuals reached adulthood. However, protein restriction provokes body underweight and hypoinsulinemia. This review is focused on the regulation of insulin secretion and the influence of the autonomic nervous system (ANS in adult rats that were protein-malnourished during lactation. The data available on the topic suggest that the perinatal phase of lactation, when insulted by protein deficit, imprints the adult metabolism and thereby alters the glycemic control. Although hypoinsulinemia programs adult rats to maintain normoglycemia, pancreatic β-cells are less sensitive to secretion stimuli, such as glucose and cholinergic agents. These pancreatic dysfunctions may be attributed to an imbalance of ANS activity recorded in adult rats that experienced maternal protein restriction.

  2. Fibroblast activation protein (FAP) as a novel metabolic target

    DEFF Research Database (Denmark)

    Sánchez-Garrido, Miguel Angel; Habegger, Kirk M; Clemmensen, Christoffer

    2016-01-01

    to block FAP enzymatic activity. RESULTS: TB administration to diet-induced obese (DIO) animals led to profound decreases in body weight, reduced food consumption and adiposity, increased energy expenditure, improved glucose tolerance and insulin sensitivity, and lowered cholesterol levels. Total...... (TB), we explored the impact of FAP inhibition on metabolic regulation in mice. METHODS: To address this question we evaluated the pharmacology of TB in various mouse models including those deficient in FGF21, GLP1 and GIP signaling. We also studied the ability of FAP to process FGF21 in vitro and TB...... and intact plasma FGF21 were observed to be elevated in TB-treated DIO mice but not lean animals where the metabolic impact of TB was significantly attenuated. Furthermore, and in stark contrast to naïve DIO mice, the administration of TB to obese FGF21 knockout animals demonstrated no appreciable effect...

  3. A novel approach to preparing magnetic protein microspheres with core-shell structure

    Science.gov (United States)

    Jiang, Wei; Sun, Zhendong; Li, Fengsheng; Chen, Kai; Liu, Tianyu; Liu, Jialing; Zhou, Tianle; Guo, Rui

    2011-03-01

    Magnetic protein microspheres with core-shell structure were prepared through a novel approach based on the sonochemical method and the emulsion solvent evaporation method. The microspheres are composed of the oleic acid and undecylenic acid modified Fe 3O 4 cores and coated with globular bovine serum albumin (BSA). Under an optimized condition, up to 57.8 wt% of approximately 10 nm superparamagnetic Fe 3O 4 nanoparticles could be uniformly encapsulated into the BSA microspheres with the diameter of approximately 160 nm and the high saturation magnetization of 38.5 emu/g, besides of the abundant functional groups. The possible formation mechanism of magnetic microspheres was discussed in detail.

  4. TRF2 Protein Interacts with Core Histones to Stabilize Chromosome Ends*

    Science.gov (United States)

    Izumi, Takashi; Shimizu, Shigeomi

    2016-01-01

    Mammalian chromosome ends are protected by a specialized nucleoprotein complex called telomeres. Both shelterin, a telomere-specific multi-protein complex, and higher order telomeric chromatin structures combine to stabilize the chromosome ends. Here, we showed that TRF2, a component of shelterin, binds to core histones to protect chromosome ends from inappropriate DNA damage response and loss of telomeric DNA. The N-terminal Gly/Arg-rich domain (GAR domain) of TRF2 directly binds to the globular domain of core histones. The conserved arginine residues in the GAR domain of TRF2 are required for this interaction. A TRF2 mutant with these arginine residues substituted by alanine lost the ability to protect telomeres and induced rapid telomere shortening caused by the cleavage of a loop structure of the telomeric chromatin. These findings showed a previously unnoticed interaction between the shelterin complex and nucleosomal histones to stabilize the chromosome ends. PMID:27514743

  5. Controlled-release and preserved bioactivity of proteins from (self-assembled core-shell double-walled microspheres

    Directory of Open Access Journals (Sweden)

    Yuan W

    2012-01-01

    Full Text Available Weien Yuan1,2, Zhenguo Liu11Department of Neurology, Xinhua Hospital, affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, 2School of Pharmacy, Shanghai Jiao Tong University, Shanghai, People’s Republic of ChinaAbstract: In order to address preserved protein bioactivities and protein sustained-release problems, a method for preparing double-walled microspheres with a core (protein-loaded nanoparticles with a polymer-suspended granule system-formed core and a second shell (a polymer-formed shell for controlled drug release and preserved protein bioactivities has been developed using (solid-in-oil phase-in-hydrophilic oil-in-water (S/O/Oh/W phases. The method, based on our previous microsphere preparation method (solid-in-oil phase-in-hydrophilic oil-in-water (S/O/Oh/W, employs different concentric poly(D,L-lactide-co-glycolide, poly(D,L-lactide, and protein-loaded nanoparticles to produce a suspended liquid which then self-assembles to form shell-core microspheres in the hydrophilic oil phase, which are then solidified in the water phase. Variations in the preparation parameters allowed complete encapsulation by the shell phase, including the efficient formation of a poly(D,L-lactide shell encapsulating a protein-loaded nanoparticle-based poly(D,L-lactide-co-glycolide core. This method produces core-shell double-walled microspheres that show controlled protein release and preserved protein bioactivities for 60 days. Based upon these results, we concluded that the core-shell double-walled microspheres might be applied for tissue engineering and therapy for chronic diseases, etc.Keywords: protein delivery, protein stability, core-shell microspheres, dextran nanoparticles

  6. YEAST A SINGLE CELL PROTEIN: CHARACTERISTICS and METABOLISM

    OpenAIRE

    AMATA, I.A

    2013-01-01

    Most of the developing countries of the world are facing a major problem of malnutrition. Due to rapid growth in the population, food and feed scarcity are prevalent leading to a deficiency of protein and essential nutrients amongst human beings and livestock. It is therefore important to take necessary measures to stem this trend by increasing protein production and making it available and more affordable to the population by utilizing methods available for the production of alternative sour...

  7. Engineered proteins with PUF scaffold to manipulate RNA metabolism

    Science.gov (United States)

    Wang, Yang; Wang, Zefeng; Tanaka Hall, Traci M.

    2013-01-01

    Pumilio/fem-3 mRNA binding factor (FBF) proteins are characterized by a sequence-specific RNA-binding domain. This unique single-stranded RNA recognition module, whose sequence specificity can be reprogrammed, has been fused with functional modules to engineer protein factors with various functions. Here we summarize the advancement in developing RNA regulatory tools and opportunities for the future. PMID:23731364

  8. Protein nutrition and metabolism during early development of the chick embryo

    International Nuclear Information System (INIS)

    Klein, N.W.

    1976-01-01

    Cultures of intact early chick embryos have been used as a model system in which to study the nutrition and metabolism of proteins during early embryonic development. Previous studies have shown that these embryos require nutrient proteins for growth and development. The protein requirement was found to be specific in that at least two proteins were essential; one a transferrin (either conalbumin or yolk transferrin) and the other either ovalbumin or lipovitellin. Variations in the quantity or type of protein provided in the medium altered the growth of embryo regions through regionally specific changes in protein breakdown. This was confirmed through protein synthetic studies with isolated polyribosomes. More recently such variations in protein nutrition have been shown also to affect the actual patterns of proteins synthesized by regions of the embryo. These observed responses to protein nutrition have been difficult to reconcile with our observation that proteins as such did not reach the embryo proper but were first degraded to amine acids within the yolk-sac membrane. Studies on the synthesis of serum proteins by the yolk-sac membrane have provided a possible explanation in that the relative synthesis of individual serum proteins was dramatically influenced by the protein composition of the culture medium. We are currently attempting to demonstrate that serum proteins are indeed the mediators of the response of embryos to protein nutrition. (author)

  9. Glucagon-Like Peptide 2 Stimulates Postresection Intestinal Adaptation in Preterm Pigs by Affecting Proteins Related to Protein, Carbohydrate, and Sulphur Metabolism

    DEFF Research Database (Denmark)

    Jiang, Pingping; Vegge, Andreas; Thymann, Thomas

    2017-01-01

    cellular structural proteins, while the added GLP-2 treatment affected proteins involved in protein processing and the metabolism of protein, carbohydrate, and sulphur. CONCLUSION: In the first days following resection, proteins affected by resection plus GLP-2 treatment differed markedly from those...

  10. Proteins involved in flor yeast carbon metabolism under biofilm formation conditions.

    Science.gov (United States)

    Moreno-García, Jaime; García-Martínez, Teresa; Moreno, Juan; Mauricio, Juan Carlos

    2015-04-01

    A lack of sugars during the production of biologically aged wines after fermentation of grape must causes flor yeasts to metabolize other carbon molecules formed during fermentation (ethanol and glycerol, mainly). In this work, a proteome analysis involving OFFGEL fractionation prior to LC/MS detection was used to elucidate the carbon metabolism of a flor yeast strain under biofilm formation conditions (BFC). The results were compared with those obtained under non-biofilm formation conditions (NBFC). Proteins associated to processes such as non-fermentable carbon uptake, the glyoxylate and TCA cycles, cellular respiration and inositol metabolism were detected at higher concentrations under BFC than under the reference conditions (NBFC). This study constitutes the first attempt at identifying the flor yeast proteins responsible for the peculiar sensory profile of biologically aged wines. A better metabolic knowledge of flor yeasts might facilitate the development of effective strategies for improved production of these special wines. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. Structural characterization of the fusion core in syncytin, envelope protein of human endogenous retrovirus family W

    International Nuclear Information System (INIS)

    Gong Rui; Peng Xiaoxue; Kang Shuli; Feng Huixing; Huang Jianying; Zhang Wentao; Lin Donghai; Tien Po; Xiao Gengfu

    2005-01-01

    Syncytin is a captive retroviral envelope protein, possibly involved in the formation of the placental syncytiotrophoblast layer generated by trophoblast cell fusion at the maternal-fetal interface. We found that syncytin and type I viral envelope proteins shared similar structural profiling, especially in the regions of N- and C-terminal heptad repeats (NHR and CHR). We expressed the predicted regions of NHR (41 aa) and CHR (34 aa) in syncytin as a native single chain (named 2-helix protein) to characterize it. 2-helix protein exists as a trimer and is highly α-helix, thermo-stable, and denatured by low pH. NHR and CHR could form a protease-resistant complex. The complex structure built by the molecular docking demonstrated that NHR and CHR associated in an antiparallel manner. Overall, the 2-helix protein could form a thermo-stable coiled coil trimer. The fusion core structure of syncytin was first demonstrated in endogenous retrovirus. These results support the explanation how syncytin mediates cytotrophoblast cell fusion involved in placental morphogenesis

  12. Effect of Prolonged Simulated Microgravity on Metabolic Proteins in Rat Hippocampus: Steps toward Safe Space Travel.

    Science.gov (United States)

    Wang, Yun; Javed, Iqbal; Liu, Yahui; Lu, Song; Peng, Guang; Zhang, Yongqian; Qing, Hong; Deng, Yulin

    2016-01-04

    Mitochondria are not only the main source of energy in cells but also produce reactive oxygen species (ROS), which result in oxidative stress when in space. This oxidative stress is responsible for energy imbalances and cellular damage. In this study, a rat tail suspension model was used in individual experiments for 7 and 21 days to explore the effect of simulated microgravity (SM) on metabolic proteins in the hippocampus, a vital brain region involved in learning, memory, and navigation. A comparative (18)O-labeled quantitative proteomic strategy was used to observe the differential expression of metabolic proteins. Forty-two and sixty-seven mitochondrial metabolic proteins were differentially expressed after 21 and 7 days of SM, respectively. Mitochondrial Complex I, III, and IV, isocitrate dehydrogenase and malate dehydrogenase were down-regulated. Moreover, DJ-1 and peroxiredoxin 6, which defend against oxidative damage, were up-regulated in the hippocampus. Western blot analysis of proteins DJ-1 and COX 5A confirmed the mass spectrometry results. Despite these changes in mitochondrial protein expression, no obvious cell apoptosis was observed after 21 days of SM. The results of this study indicate that the oxidative stress induced by SM has profound effects on metabolic proteins.

  13. HBV core protein allosteric modulators differentially alter cccDNA biosynthesis from de novo infection and intracellular amplification pathways

    Science.gov (United States)

    Guo, Fang; Zhao, Qiong; Cheng, Junjun; Qi, Yonghe; Su, Qing; Wei, Lai; Li, Wenhui; Chang, Jinhong

    2017-01-01

    Hepatitis B virus (HBV) core protein assembles viral pre-genomic (pg) RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs) and sulfamoylbenzamides (SBAs), have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or “empty” capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc) DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B. PMID:28945802

  14. HBV core protein allosteric modulators differentially alter cccDNA biosynthesis from de novo infection and intracellular amplification pathways.

    Science.gov (United States)

    Guo, Fang; Zhao, Qiong; Sheraz, Muhammad; Cheng, Junjun; Qi, Yonghe; Su, Qing; Cuconati, Andrea; Wei, Lai; Du, Yanming; Li, Wenhui; Chang, Jinhong; Guo, Ju-Tao

    2017-09-01

    Hepatitis B virus (HBV) core protein assembles viral pre-genomic (pg) RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs) and sulfamoylbenzamides (SBAs), have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or "empty" capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc) DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B.

  15. HBV core protein allosteric modulators differentially alter cccDNA biosynthesis from de novo infection and intracellular amplification pathways.

    Directory of Open Access Journals (Sweden)

    Fang Guo

    2017-09-01

    Full Text Available Hepatitis B virus (HBV core protein assembles viral pre-genomic (pg RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs and sulfamoylbenzamides (SBAs, have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or "empty" capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B.

  16. The TIM Barrel Architecture Facilitated the Early Evolution of Protein-Mediated Metabolism.

    Science.gov (United States)

    Goldman, Aaron David; Beatty, Joshua T; Landweber, Laura F

    2016-01-01

    The triosephosphate isomerase (TIM) barrel protein fold is a structurally repetitive architecture that is present in approximately 10% of all enzymes. It is generally assumed that this ubiquity in modern proteomes reflects an essential historical role in early protein-mediated metabolism. Here, we provide quantitative and comparative analyses to support several hypotheses about the early importance of the TIM barrel architecture. An information theoretical analysis of protein structures supports the hypothesis that the TIM barrel architecture could arise more easily by duplication and recombination compared to other mixed α/β structures. We show that TIM barrel enzymes corresponding to the most taxonomically broad superfamilies also have the broadest range of functions, often aided by metal and nucleotide-derived cofactors that are thought to reflect an earlier stage of metabolic evolution. By comparison to other putatively ancient protein architectures, we find that the functional diversity of TIM barrel proteins cannot be explained simply by their antiquity. Instead, the breadth of TIM barrel functions can be explained, in part, by the incorporation of a broad range of cofactors, a trend that does not appear to be shared by proteins in general. These results support the hypothesis that the simple and functionally general TIM barrel architecture may have arisen early in the evolution of protein biosynthesis and provided an ideal scaffold to facilitate the metabolic transition from ribozymes, peptides, and geochemical catalysts to modern protein enzymes.

  17. Role of Heme and Heme-Proteins in Trypanosomatid Essential Metabolic Pathways

    Directory of Open Access Journals (Sweden)

    Karina E. J. Tripodi

    2011-01-01

    Full Text Available Around the world, trypanosomatids are known for being etiological agents of several highly disabling and often fatal diseases like Chagas disease (Trypanosoma cruzi, leishmaniasis (Leishmania spp., and African trypanosomiasis (Trypanosoma brucei. Throughout their life cycle, they must cope with diverse environmental conditions, and the mechanisms involved in these processes are crucial for their survival. In this review, we describe the role of heme in several essential metabolic pathways of these protozoans. Notwithstanding trypanosomatids lack of the complete heme biosynthetic pathway, we focus our discussion in the metabolic role played for important heme-proteins, like cytochromes. Although several genes for different types of cytochromes, involved in mitochondrial respiration, polyunsaturated fatty acid metabolism, and sterol biosynthesis, are annotated at the Tritryp Genome Project, the encoded proteins have not yet been deeply studied. We pointed our attention into relevant aspects of these protein functions that are amenable to be considered for rational design of trypanocidal agents.

  18. Association between C-reactive protein and features of the metabolic syndrome

    DEFF Research Database (Denmark)

    Fröhlich, M; Imhof, A; Berg, Gabriele

    2000-01-01

    OBJECTIVE: To assess the association of circulating levels of C-reactive protein, a sensitive systemic marker of inflammation, with different components of the metabolic syndrome. RESEARCH DESIGN AND METHODS: Total cholesterol (TC), HDL cholesterol, triglycerides, uric acid, BMI , and prevalence...... concentrations in subjects grouped according to the presence of 0-1, 2-3, and > or =4 features of the metabolic syndrome were 1.11, 1.27, and 2.16 mg/l, respectively, with a statistically highly significant trend (P metabolic syndrome...

  19. Metabolic responses to high protein diet in Korean elite bodybuilders with high-intensity resistance exercise

    Directory of Open Access Journals (Sweden)

    Choue Ryowon

    2011-07-01

    Full Text Available Abstract Background High protein diet has been known to cause metabolic acidosis, which is manifested by increased urinary excretion of nitrogen and calcium. Bodybuilders habitually consumed excessive dietary protein over the amounts recommended for them to promote muscle mass accretion. This study investigated the metabolic response to high protein consumption in the elite bodybuilders. Methods Eight elite Korean bodybuilders within the age from 18 to 25, mean age 21.5 ± 2.6. For data collection, anthropometry, blood and urinary analysis, and dietary assessment were conducted. Results They consumed large amounts of protein (4.3 ± 1.2 g/kg BW/day and calories (5,621.7 ± 1,354.7 kcal/day, as well as more than the recommended amounts of vitamins and minerals, including potassium and calcium. Serum creatinine (1.3 ± 0.1 mg/dl and potassium (5.9 ± 0.8 mmol/L, and urinary urea nitrogen (24.7 ± 9.5 mg/dl and creatinine (2.3 ± 0.7 mg/dl were observed to be higher than the normal reference ranges. Urinary calcium (0.3 ± 0.1 mg/dl, and phosphorus (1.3 ± 0.4 mg/dl were on the border of upper limit of the reference range and the urine pH was in normal range. Conclusions Increased urinary excretion of urea nitrogen and creatinine might be due to the high rates of protein metabolism that follow high protein intake and muscle turnover. The obvious evidence of metabolic acidosis in response to high protein diet in the subjects with high potassium intake and intensive resistance exercise were not shown in this study results. However, this study implied that resistance exercise with adequate mineral supplementation, such as potassium and calcium, could reduce or offset the negative effects of protein-generated metabolic changes. This study provides preliminary information of metabolic response to high protein intake in bodybuilders who engaged in high-intensity resistance exercise. Further studies will be needed to determine the effects of the intensity

  20. Metabolism of homologous and heterologous serum proteins in garter snakes (Thamnophis ordinoides)

    International Nuclear Information System (INIS)

    Leong, D.; Coe, J.E.

    1978-01-01

    The half-life (Tsub(1/2) of serum immunoglobulin (Ig) and albumin from snakes and mammals were determined in both garter snakes (Thamnophis ordinoides) and mice (Mus musculus). Metabolism of serum proteins in snakes was similar to mammalian protein metabolism in that homologous serum albumin had shorter Tsub(1/2) (16 days) than IgG (38 days). Also, reptilian and mammalian serum proteins had a relatively longer Tsub(1/2) when injected into closely related species. Thus mammalian serum Ig (rabbit gamma globulin (RGG)) had a shorter Tsub(1/2) (6.3 days) in snake than did homologous snake IgG (38 days), whereas in mice, RGG had a longer Tsub(1/2) (3.8 days) than snake Ig (0.9 days). Differences between metabolism of homologous and heterologous albumins were apparent only in snakes in which the Tsub(1/2) of homologous albumin was approximately 8-fold greater than mammalian albumin. These results indicate that metabolism of both Ig and albumin in snakes is regulated by specific receptors whereas albumin receptors have been difficult to demonstrate in mammals. The results of this study suggest that one of the factors determining the metabolism of a protein is its foreignness to the host perhaps because of receptor cross reactions. (author)

  1. The Role of Maternal Dietary Proteins in Development of Metabolic Syndrome in Offspring

    Directory of Open Access Journals (Sweden)

    Alireza Jahan-Mihan

    2015-11-01

    Full Text Available The prevalence of metabolic syndrome and obesity has been increasing. Pre-natal environment has been suggested as a factor influencing the risk of metabolic syndrome in adulthood. Both observational and experimental studies showed that maternal diet is a major modifier of the development of regulatory systems in the offspring in utero and post-natally. Both protein content and source in maternal diet influence pre- and early post-natal development. High and low protein dams’ diets have detrimental effect on body weight, blood pressure191 and metabolic and intake regulatory systems in the offspring. Moreover, the role of the source of protein in a nutritionally adequate maternal diet in programming of food intake regulatory system, body weight, glucose metabolism and blood pressure in offspring is studied. However, underlying mechanisms are still elusive. The purpose of this review is to examine the current literature related to the role of proteins in maternal diets in development of characteristics of the metabolic syndrome in offspring.

  2. Data set of interactomes and metabolic pathways of proteins differentially expressed in brains with Alzheimer׳s disease

    Directory of Open Access Journals (Sweden)

    Benito Minjarez

    2016-06-01

    Full Text Available Alzheimer׳s disease is one of the main causes of dementia in the elderly and its frequency is on the rise worldwide. It is considered the result of complex interactions between genetic and environmental factors, being many of them unknown. Therefore, there is a dire necessity for the identification of novel molecular players for the understanding of this disease. In this data article we determined the protein expression profiles of whole protein extracts from cortex regions of brains from patients with Alzheimer׳s disease in comparison to a normal brain. We identified 721 iTRAQ-labeled polypeptides with more than 95% in confidence. We analyzed all proteins that changed in their expression level and located them in the KEGG metabolic pathways, as well as in the mitochondrial complexes of the electron transport chain and ATP synthase. In addition, we analyzed the over- and sub-expressed polypeptides through IPA software, specifically Core I and Biomarkers I modules. Data in this article is related to the research article “Identification of proteins that are differentially expressed in brains with Alzheimer’s disease using iTRAQ labeling and tandem mass spectrometry” (Minjarez et al., 2016 [1].

  3. Female hormones: do they influence muscle and tendon protein metabolism?

    DEFF Research Database (Denmark)

    Hansen, Mette

    2018-01-01

    (or lack of female hormones) on skeletal muscle protein turnover at rest and in response to exercise. This review is primarily based on data from human trials. Many elderly post-menopausal women experience physical disabilities and loss of independence related to sarcopenia, which reduces life quality...

  4. Effects of atorvastatin on human c reactive protein metabolism

    Science.gov (United States)

    Statins are known to reduce plasma C-reactive protein (CRP) concentrations. Our goals were to define the mechanisms by which CRP was reduced by maximal dose atorvastatin. Eight subjects with combined hyperlipidemia (5 men and 3 postmenopausal women) were enrolled in a randomized, placebo-controlled...

  5. Regulation of triglyceride metabolism by angiopoietin-like proteins

    NARCIS (Netherlands)

    Mattijssen, F.B.J.; Kersten, A.H.

    2012-01-01

    asma triglyceride concentrations are determined by the balance between production of the triglyceride-rich lipoproteins VLDL and chylomicrons in liver and intestine, and their lipoprotein lipase-mediated clearance in peripheral tissues. In the last decade, the group of Angiopoietin-like proteins has

  6. Deep proteomics of mouse skeletal muscle enables quantitation of protein isoforms, metabolic pathways, and transcription factors.

    Science.gov (United States)

    Deshmukh, Atul S; Murgia, Marta; Nagaraj, Nagarjuna; Treebak, Jonas T; Cox, Jürgen; Mann, Matthias

    2015-04-01

    Skeletal muscle constitutes 40% of individual body mass and plays vital roles in locomotion and whole-body metabolism. Proteomics of skeletal muscle is challenging because of highly abundant contractile proteins that interfere with detection of regulatory proteins. Using a state-of-the art MS workflow and a strategy to map identifications from the C2C12 cell line model to tissues, we identified a total of 10,218 proteins, including skeletal muscle specific transcription factors like myod1 and myogenin and circadian clock proteins. We obtain absolute abundances for proteins expressed in a muscle cell line and skeletal muscle, which should serve as a valuable resource. Quantitation of protein isoforms of glucose uptake signaling pathways and in glucose and lipid metabolic pathways provides a detailed metabolic map of the cell line compared with tissue. This revealed unexpectedly complex regulation of AMP-activated protein kinase and insulin signaling in muscle tissue at the level of enzyme isoforms. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Motile hepatocellular carcinoma cells preferentially secret sugar metabolism regulatory proteins via exosomes.

    Science.gov (United States)

    Zhang, Jing; Lu, Shaohua; Zhou, Ye; Meng, Kun; Chen, Zhipeng; Cui, Yizhi; Shi, Yunfeng; Wang, Tong; He, Qing-Yu

    2017-07-01

    Exosomes are deliverers of critically functional proteins, capable of transforming target cells in numerous cancers, including hepatocellular carcinoma (HCC). We hypothesize that the motility of HCC cells can be featured by comparative proteome of exosomes. Hence, we performed the super-SILAC-based MS analysis on the exosomes secreted by three human HCC cell lines, including the non-motile Hep3B cell, and the motile 97H and LM3 cells. More than 1400 exosomal proteins were confidently quantified in each MS analysis with highly biological reproducibility. We justified that 469 and 443 exosomal proteins represented differentially expressed proteins (DEPs) in the 97H/Hep3B and LM3/Hep3B comparisons, respectively. These DEPs focused on sugar metabolism-centric canonical pathways per ingenuity pathway analysis, which was consistent with the gene ontology analysis on biological process enrichment. These pathways included glycolysis I, gluconeogenesis I and pentose phosphate pathways; and the DEPs enriched in these pathways could form a tightly connected network. By analyzing the relative abundance of proteins and translating mRNAs, we found significantly positive correlation between exosomes and cells. The involved exosomal proteins were again focusing on sugar metabolism. In conclusion, motile HCC cells tend to preferentially export more sugar metabolism-associated proteins via exosomes that differentiate them from non-motile HCC cells. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Myocardial Oxidative Metabolism and Protein Synthesis during Mechanical Circulatory Support by Extracorporeal Membrane Oxygenation

    Energy Technology Data Exchange (ETDEWEB)

    Priddy, MD, Colleen M.; Kajimoto, Masaki; Ledee, Dolena; Bouchard, Bertrand; Isern, Nancy G.; Olson, Aaron; Des Rosiers, Christine; Portman, Michael A.

    2013-02-01

    Extracorporeal membrane oxygenation (ECMO) provides mechanical circulatory support essential for survival in infants and children with acute cardiac decompensation. However, ECMO also causes metabolic disturbances, which contribute to total body wasting and protein loss. Cardiac stunning can also occur which prevents ECMO weaning, and contributes to high mortality. The heart may specifically undergo metabolic impairments, which influence functional recovery. We tested the hypothesis that ECMO alters oxidative. We focused on the amino acid leucine, and integration with myocardial protein synthesis. We used a translational immature swine model in which we assessed in heart (i) the fractional contribution of leucine (FcLeucine) and pyruvate (FCpyruvate) to mitochondrial acetyl-CoA formation by nuclear magnetic resonance and (ii) global protein fractional synthesis (FSR) by gas chromatography-mass spectrometry. Immature mixed breed Yorkshire male piglets (n = 22) were divided into four groups based on loading status (8 hours of normal circulation or ECMO) and intracoronary infusion [13C6,15N]-L-leucine (3.7 mM) alone or with [2-13C]-pyruvate (7.4 mM). ECMO decreased pulse pressure and correspondingly lowered myocardial oxygen consumption (~ 40%, n = 5), indicating decreased overall mitochondrial oxidative metabolism. However, FcLeucine was maintained and myocardial protein FSR was marginally increased. Pyruvate addition decreased tissue leucine enrichment, FcLeucine, and Fc for endogenous substrates as well as protein FSR. Conclusion: The heart under ECMO shows reduced oxidative metabolism of substrates, including amino acids, while maintaining (i) metabolic flexibility indicated by ability to respond to pyruvate, and (ii) a normal or increased capacity for global protein synthesis, suggesting an improved protein balance.

  9. Structure of the protein core of the glypican Dally-like and localization of a region important for hedgehog signaling

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Min-Sung; Saunders, Adam M.; Hamaoka, Brent Y.; Beachy, Philip A.; Leahy, Daniel J. (Stanford-MED); (JHU)

    2011-09-20

    Glypicans are heparan sulfate proteoglycans that modulate the signaling of multiple growth factors active during animal development, and loss of glypican function is associated with widespread developmental abnormalities. Glypicans consist of a conserved, approximately 45-kDa N-terminal protein core region followed by a stalk region that is tethered to the cell membrane by a glycosyl-phosphatidylinositol anchor. The stalk regions are predicted to be random coil but contain a variable number of attachment sites for heparan sulfate chains. Both the N-terminal protein core and the heparan sulfate attachments are important for glypican function. We report here the 2.4-{angstrom} crystal structure of the N-terminal protein core region of the Drosophila glypican Dally-like (Dlp). This structure reveals an elongated, {alpha}-helical fold for glypican core regions that does not appear homologous to any known structure. The Dlp core protein is required for normal responsiveness to Hedgehog (Hh) signals, and we identify a localized region on the Dlp surface important for mediating its function in Hh signaling. Purified Dlp protein core does not, however, interact appreciably with either Hh or an Hh:Ihog complex.

  10. An evolutionarily conserved glycine-tyrosine motif forms a folding core in outer membrane proteins.

    Directory of Open Access Journals (Sweden)

    Marcin Michalik

    Full Text Available An intimate interaction between a pair of amino acids, a tyrosine and glycine on neighboring β-strands, has been previously reported to be important for the structural stability of autotransporters. Here, we show that the conservation of this interacting pair extends to nearly all major families of outer membrane β-barrel proteins, which are thought to have originated through duplication events involving an ancestral ββ hairpin. We analyzed the function of this motif using the prototypical outer membrane protein OmpX. Stopped-flow fluorescence shows that two folding processes occur in the millisecond time regime, the rates of which are reduced in the tyrosine mutant. Folding assays further demonstrate a reduction in the yield of folded protein for the mutant compared to the wild-type, as well as a reduction in thermal stability. Taken together, our data support the idea of an evolutionarily conserved 'folding core' that affects the folding, membrane insertion, and thermal stability of outer membrane protein β-barrels.

  11. Human Adenovirus Core Protein V Is Targeted by the Host SUMOylation Machinery To Limit Essential Viral Functions.

    Science.gov (United States)

    Freudenberger, Nora; Meyer, Tina; Groitl, Peter; Dobner, Thomas; Schreiner, Sabrina

    2018-02-15

    Human adenoviruses (HAdV) are nonenveloped viruses containing a linear, double-stranded DNA genome surrounded by an icosahedral capsid. To allow proper viral replication, the genome is imported through the nuclear pore complex associated with viral core proteins. Until now, the role of these incoming virion proteins during the early phase of infection was poorly understood. The core protein V is speculated to bridge the core and the surrounding capsid. It binds the genome in a sequence-independent manner and localizes in the nucleus of infected cells, accumulating at nucleoli. Here, we show that protein V contains conserved SUMO conjugation motifs (SCMs). Mutation of these consensus motifs resulted in reduced SUMOylation of the protein; thus, protein V represents a novel target of the host SUMOylation machinery. To understand the role of protein V SUMO posttranslational modification during productive HAdV infection, we generated a replication-competent HAdV with SCM mutations within the protein V coding sequence. Phenotypic analyses revealed that these SCM mutations are beneficial for adenoviral replication. Blocking protein V SUMOylation at specific sites shifts the onset of viral DNA replication to earlier time points during infection and promotes viral gene expression. Simultaneously, the altered kinetics within the viral life cycle are accompanied by more efficient proteasomal degradation of host determinants and increased virus progeny production than that observed during wild-type infection. Taken together, our studies show that protein V SUMOylation reduces virus growth; hence, protein V SUMOylation represents an important novel aspect of the host antiviral strategy to limit virus replication and thereby points to potential intervention strategies. IMPORTANCE Many decades of research have revealed that HAdV structural proteins promote viral entry and mainly physical stability of the viral genome in the capsid. Our work over the last years showed that this

  12. Methodical investigation of the protein metabolism and of the bioenergetics of the protein retention in growing animals. 2

    International Nuclear Information System (INIS)

    Voelker, T.; Krawielitzki, K.; Klein, M.; Keller, J.

    1983-01-01

    The amino acid composition of the proteins in selected body fractions of chickens and the 15 N -excess of amino acids isolated from them resulting from a feeding experiment with long-term 15 NH 4 -acetate labelling were determined. The amino acid spectra of feathers, breast and leg muscles are characterized by differences in the content of individual amino acids specific for the organs, the composition of the proteins, however, is independent of the protein content of the ration and the age of the animals. The sarcoplasmatic and myofibrillar proteins also have typical amino acid patterns, which-with the exception of the histidine content-are neither influenced by the extraction of the proteins from the breast or leg muscles nor by the energy level of the feeding or the age of the animals. There are no significant differences in the metabolization of the main protein fraction of the breast and leg muscles. The oral supply of 15 N-ammonium acetate to broilers predominantly labels the non-essential amino acids so that the derived kinetic data chiefly represent the metabolism of the non-essential amino acids. (author)

  13. Maternal protein intake in pregnancy and offspring metabolic health at age 9-16 y

    DEFF Research Database (Denmark)

    Maslova, Ekaterina; Hansen, Susanne; Grunnet, Louise Groth

    2017-01-01

    in free-living populations remains limited. Objective: We examined the association of protein intake in pregnancy with offspring metabolic health at age 9-16 y in a longitudinal cohort that oversampled pregnancies with gestational diabetes mellitus (GDM). Design: Six hundred eight women with an index...... provide little support for an association of maternal protein intake in pregnancy with measures of offspring metabolic health. Further studies in larger cohorts are needed to determine whether low maternal protein intake in pregnancy may improve glucose homeostasis in GDM-exposed and male offspring....... pregnancy affected by gestational diabetes mellitus and 626 controls enrolled in the Danish National Birth Cohort were used for the analysis. Protein (total, animal, vegetable) intake was assessed by using a foodfrequency questionnaire in gestational week 25. The offspring underwent a clinical examination...

  14. Identification of Proteins Involved in Carbohydrate Metabolism and Energy Metabolism Pathways and Their Regulation of Cytoplasmic Male Sterility in Wheat

    Directory of Open Access Journals (Sweden)

    Xingxia Geng

    2018-01-01

    Full Text Available Cytoplasmic male sterility (CMS where no functional pollen is produced has important roles in wheat breeding. The anther is a unique organ for male gametogenesis and its abnormal development can cause male sterility. However, the mechanisms and regulatory networks related to plant male sterility are poorly understood. In this study, we conducted comparative analyses using isobaric tags for relative and absolute quantification (iTRAQ of the pollen proteins in a CMS line and its wheat maintainer. Differentially abundant proteins (DAPs were analyzed based on Gene Ontology classifications, metabolic pathways and transcriptional regulation networks using Blast2GO. We identified 5570 proteins based on 23,277 peptides, which matched with 73,688 spectra, including proteins in key pathways such as glyceraldehyde-3-phosphate dehydrogenase, pyruvate kinase and 6-phosphofructokinase 1 in the glycolysis pathway, isocitrate dehydrogenase and citrate synthase in the tricarboxylic acid cycle and nicotinamide adenine dinucleotide (NADH-dehydrogenase and adenosine-triphosphate (ATP synthases in the oxidative phosphorylation pathway. These proteins may comprise a network that regulates male sterility in wheat. Quantitative real time polymerase chain reaction (qRT-PCR analysis, ATP assays and total sugar assays validated the iTRAQ results. These DAPs could be associated with abnormal pollen grain formation and male sterility. Our findings provide insights into the molecular mechanism related to male sterility in wheat.

  15. Identification of Proteins Involved in Carbohydrate Metabolism and Energy Metabolism Pathways and Their Regulation of Cytoplasmic Male Sterility in Wheat.

    Science.gov (United States)

    Geng, Xingxia; Ye, Jiali; Yang, Xuetong; Li, Sha; Zhang, Lingli; Song, Xiyue

    2018-01-23

    Cytoplasmic male sterility (CMS) where no functional pollen is produced has important roles in wheat breeding. The anther is a unique organ for male gametogenesis and its abnormal development can cause male sterility. However, the mechanisms and regulatory networks related to plant male sterility are poorly understood. In this study, we conducted comparative analyses using isobaric tags for relative and absolute quantification (iTRAQ) of the pollen proteins in a CMS line and its wheat maintainer. Differentially abundant proteins (DAPs) were analyzed based on Gene Ontology classifications, metabolic pathways and transcriptional regulation networks using Blast2GO. We identified 5570 proteins based on 23,277 peptides, which matched with 73,688 spectra, including proteins in key pathways such as glyceraldehyde-3-phosphate dehydrogenase, pyruvate kinase and 6-phosphofructokinase 1 in the glycolysis pathway, isocitrate dehydrogenase and citrate synthase in the tricarboxylic acid cycle and nicotinamide adenine dinucleotide (NADH)-dehydrogenase and adenosine-triphosphate (ATP) synthases in the oxidative phosphorylation pathway. These proteins may comprise a network that regulates male sterility in wheat. Quantitative real time polymerase chain reaction (qRT-PCR) analysis, ATP assays and total sugar assays validated the iTRAQ results. These DAPs could be associated with abnormal pollen grain formation and male sterility. Our findings provide insights into the molecular mechanism related to male sterility in wheat.

  16. Influence of culture conditions on growth and protein metabolism in chlorella pyranosides

    International Nuclear Information System (INIS)

    Mazon Matanzo, M. P.; Fernandez Gonzalez, J.; Batuecas Suarez, B.

    1981-01-01

    Growth and protein metabolism of Chlorella pyranoside under different conditions of temperature, photo period and CO 2 concentration was studied. The optimum of biomass production was observed at 25 degree centigree, 40.000 ppm of CO 2 in air and a 20 h. light period, followed of 4 h. of darkness. Some variations in free aminoacids content was observed under different conditions but no change did occur in protein. (Author) 68 refs

  17. Influence of culture conditions on growth and protein metabolism in chlorella pyrenoidosa

    International Nuclear Information System (INIS)

    Fernandez Gonzalez, J.; Mazon, M.P.; Batuecas, B.

    1981-01-01

    Growth and protein metabolism of chlorella pyrenoidosa under differents conditions of temperature, photoperiod and CO 2 concentration was studied. The optimum of biomas production was observed at 25 deg C, 40.000 ppm of CO 2 in air and a 20 h. light period, followed of 4 h. of darkness. Some variations in free aminoacids content was observed under differents conditions but no change did occur in protein. (author)

  18. Association between C-reactive protein and features of the metabolic syndrome

    DEFF Research Database (Denmark)

    Fröhlich, M; Imhof, A; Berg, Gabriele

    2000-01-01

    OBJECTIVE: To assess the association of circulating levels of C-reactive protein, a sensitive systemic marker of inflammation, with different components of the metabolic syndrome. RESEARCH DESIGN AND METHODS: Total cholesterol (TC), HDL cholesterol, triglycerides, uric acid, BMI , and prevalence...... C-reactive protein and TC (R = 0.19), TG (R = 0.29), BMI (R = 0.32), glucose (R = 0.11), and uric acid (R = 0.14) (all P

  19. Effects of Dairy Protein and Fat on the Metabolic Syndrome and Type 2 Diabetes

    OpenAIRE

    Bjørnshave, Ann; Hermansen, Kjeld

    2014-01-01

    The incidence of the metabolic syndrome (MetS) and type 2 diabetes (T2D) is increasing worldwide. Evidence supports a negative relationship between the consumption of dairy products and risk of MetS and T2D. Dairy proteins are known to have a directly beneficial effect on hypertension, dyslipidemia, and hyperglycemia, but a detailed understanding of the underlying mechanisms is missing. It has been confirmed by observations that the insulinotropic effect of dairy proteins is associated with t...

  20. Molecular events in matrix protein metabolism in the aging kidney

    Science.gov (United States)

    Sataranatarajan, Kavithalakshmi; Feliers, Denis; Mariappan, Meenalakshmi M.; Lee, Hak Joo; Lee, Myung Ja; Day, Robert T.; Yalamanchili, Hima Bindu; Choudhury, Goutam G.; Barnes, Jeffrey L.; Van Remmen, Holly; Richardson, Arlan; Kasinath, Balakuntalam S.

    2018-01-01

    Summary We explored molecular events associated with aging-induced matrix changes in the kidney. C57BL6 mice were studied in youth, middle age, and old age. Albuminuria and serum cystatin C level (an index of glomerular filtration) increased with aging. Renal hypertrophy was evident in middle-aged and old mice and was associated with glomerulomegaly and increase in mesangial fraction occupied by extracellular matrix. Content of collagen types I and III and fibronectin was increased with aging; increment in their mRNA varied with the phase of aging. The content of ZEB1 and ZEB2, collagen type I transcription inhibitors, and their binding to the collagen type Iα2 promoter by ChIP assay also showed age-phase-specific changes. Lack of increase in mRNA and data from polysome assay suggested decreased degradation as a potential mechanism for kidney collagen type I accumulation in the middle-aged mice. These changes occurred with increment in TGFβ mRNA and protein and activation of its SMAD3 pathway; SMAD3 binding to the collagen type Iα2 promoter was also increased. TGFβ-regulated microRNAs (miRs) exhibited selective regulation. The renal cortical content of miR-21 and miR-200c, but not miR-192, miR-200a, or miR-200b, was increased with aging. Increased miR-21 and miR-200c contents were associated with reduced expression of their targets, Sprouty-1 and ZEB2, respectively. These data show that aging is associated with complex molecular events in the kidney that are already evident in the middle age and progress to old age. Agephase-specific regulation of matrix protein synthesis occurs and involves matrix protein-specific transcriptional and post-transcriptional mechanisms. PMID:23020145

  1. Insulin Stimulates S100B Secretion and These Proteins Antagonistically Modulate Brain Glucose Metabolism.

    Science.gov (United States)

    Wartchow, Krista Minéia; Tramontina, Ana Carolina; de Souza, Daniela F; Biasibetti, Regina; Bobermin, Larissa D; Gonçalves, Carlos-Alberto

    2016-06-01

    Brain metabolism is highly dependent on glucose, which is derived from the blood circulation and metabolized by the astrocytes and other neural cells via several pathways. Glucose uptake in the brain does not involve insulin-dependent glucose transporters; however, this hormone affects the glucose influx to the brain. Changes in cerebrospinal fluid levels of S100B (an astrocyte-derived protein) have been associated with alterations in glucose metabolism; however, there is no evidence whether insulin modulates glucose metabolism and S100B secretion. Herein, we investigated the effect of S100B on glucose metabolism, measuring D-(3)H-glucose incorporation in two preparations, C6 glioma cells and acute hippocampal slices, and we also investigated the effect of insulin on S100B secretion. Our results showed that: (a) S100B at physiological levels decreases glucose uptake, through the multiligand receptor RAGE and mitogen-activated protein kinase/ERK signaling, and (b) insulin stimulated S100B secretion via PI3K signaling. Our findings indicate the existence of insulin-S100B modulation of glucose utilization in the brain tissue, and may improve our understanding of glucose metabolism in several conditions such as ketosis, streptozotocin-induced dementia and pharmacological exposure to antipsychotics, situations that lead to changes in insulin signaling and extracellular levels of S100B.

  2. The effect of milk and milk proteins on risk factors of metabolic syndrome in overweight adolecents

    DEFF Research Database (Denmark)

    Arnberg, Karina

    This PhD is based on data from an intervention study with milk and milk proteins conducted in Danish adolescents with overweight. There is a high prevalence of overweight in Danish adolescents. Metabolic syndrome is a cluster of risk factors related to overweight and believed to increase the risk...... of type-2 diabetes and atherosclerotic cardiovascular diseases. Overweight children have higher concentrations of the metabolic syndrome risk factors than normal weight children and the pathological condition underlying cardiovascular diseases, called atherosclerosis, seems to start in childhood. A well...... skimmed milk, whey, casein or water for three months. The background for the intervention is that milk is an important source of protein in the Western diet and epidemiological studies in children have shown that children drinking low amounts of milk have higher concentrations of the metabolic risk...

  3. Influence on bone metabolism of dietary trace elements, protein, fat, carbohydrates, and vitamins.

    Science.gov (United States)

    Sarazin, M; Alexandre, C; Thomas, T

    2000-01-01

    Osteoporosis is a multifactorial disease driven primarily by the genetic factors that control bone metabolism. Among environmental factors, diet may play a key role, affording a target for low-cost intervention. Calcium and vitamin D are well known to affect bone metabolism. Other nutrients may influence bone mass changes; for instance, a number of trace elements and vitamins other than vitamin D are essential to many of the steps of bone metabolism. A wide variety of foods provide these nutrients, and in industrialized countries deficiencies are more often due to idiosyncratic eating habits than to cultural influences. Both culture and vogue influence the amount of carbohydrate, fat, and protein in the typical diet. In children, the current trend is to reduce protein and to increase carbohydrate and fat. Data from epidemiological and animal studies suggest that this may adversely affect bone mass and the fracture risk.

  4. Role of the Mixed-Lineage Protein Kinase Pathway in the Metabolic Stress Response to Obesity

    Directory of Open Access Journals (Sweden)

    Shashi Kant

    2013-08-01

    Full Text Available Saturated free fatty acid (FFA is implicated in the metabolic response to obesity. In vitro studies indicate that FFA signaling may be mediated by the mixed-lineage protein kinase (MLK pathway that activates cJun NH2-terminal kinase (JNK. Here, we examined the role of the MLK pathway in vivo using a mouse model of diet-induced obesity. The ubiquitously expressed MLK2 and MLK3 protein kinases have partially redundant functions. We therefore compared wild-type and compound mutant mice that lack expression of MLK2 and MLK3. MLK deficiency protected mice against high-fat-diet-induced insulin resistance and obesity. Reduced JNK activation and increased energy expenditure contribute to the metabolic effects of MLK deficiency. These data confirm that the MLK pathway plays a critical role in the metabolic response to obesity.

  5. Metabolic Regulation of CaMKII Protein and Caspases in Xenopus laevis Egg Extracts*

    Science.gov (United States)

    McCoy, Francis; Darbandi, Rashid; Chen, Si-Ing; Eckard, Laura; Dodd, Keela; Jones, Kelly; Baucum, Anthony J.; Gibbons, Jennifer A.; Lin, Sue-Hwa; Colbran, Roger J.; Nutt, Leta K.

    2013-01-01

    The metabolism of the Xenopus laevis egg provides a cell survival signal. We found previously that increased carbon flux from glucose-6-phosphate (G6P) through the pentose phosphate pathway in egg extracts maintains NADPH levels and calcium/calmodulin regulated protein kinase II (CaMKII) activity to phosphorylate caspase 2 and suppress cell death pathways. Here we show that the addition of G6P to oocyte extracts inhibits the dephosphorylation/inactivation of CaMKII bound to caspase 2 by protein phosphatase 1. Thus, G6P sustains the phosphorylation of caspase 2 by CaMKII at Ser-135, preventing the induction of caspase 2-mediated apoptotic pathways. These findings expand our understanding of oocyte biology and clarify mechanisms underlying the metabolic regulation of CaMKII and apoptosis. Furthermore, these findings suggest novel approaches to disrupt the suppressive effects of the abnormal metabolism on cell death pathways. PMID:23400775

  6. Effects of antenatal steroids on protein metabolism in preterm infants on the first day of life

    NARCIS (Netherlands)

    de Pipaon, M.S.; Vanbeek, R.H.T.; Zimmermann, L.J.J.; Wattimena, D.J.L.; Quero, J.; Sauer, P. J. J.

    Objective To analyze, in an existing cohort of infants, whether antenatal administered corticosteroids influence protein metabolism in preterm infants on the first day of life. Study design Three groups of infants were studied. The mothers of 25 infants had received 2 or more doses of

  7. Heme metabolism in stress regulation and protein production: from Cinderella to a key player

    DEFF Research Database (Denmark)

    Martinez Ruiz, José Luis; Petranovic, D.; Nielsen, Jens

    2016-01-01

    Heme biosynthesis is a highly conserved pathway which is present in all kingdoms, from Archaea to higher organisms such as plants and mammals. The heme molecule acts as a prosthetic group for different proteins and enzymes involved in energy metabolism and reactions involved in electron transfer....

  8. Effect of supplemental protein source during the winter on pre- and postpartum glucose metabolism

    Science.gov (United States)

    Circulating serum glucose concentrations as well as glucose utilization have been shown to be affected by forage quality. Supplemental protein provided to grazing range cows while consuming low quality forage may improve glucose metabolism. The objective of our study was to determine the effects of ...

  9. Riboflavin carrier protein-targeted fluorescent USPIO for the assessment of vascular metabolism in tumors

    NARCIS (Netherlands)

    Jayapaul, J.; Arns, S.; Lederle, W.; Lammers, Twan Gerardus Gertudis Maria; Comba, P.; Gätjens, J.; Kiessling, F.

    2012-01-01

    Abstract Riboflavin (Rf) and its metabolic analogs flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD) are essential for normal cellular growth and function. Their intracellular transport is regulated by the riboflavin carrier protein (RCP), which has been shown to be over-expressed by

  10. Myostatin promotes distinct responses on protein metabolism of skeletal and cardiac muscle fibers of rodents.

    Science.gov (United States)

    Manfredi, L H; Paula-Gomes, S; Zanon, N M; Kettelhut, I C

    2017-10-19

    Myostatin is a novel negative regulator of skeletal muscle mass. Myostatin expression is also found in heart in a much less extent, but it can be upregulated in pathological conditions, such as heart failure. Myostatin may be involved in inhibiting protein synthesis and/or increasing protein degradation in skeletal and cardiac muscles. Herein, we used cell cultures and isolated muscles from rats to determine protein degradation and synthesis. Muscles incubated with myostatin exhibited an increase in proteolysis with an increase of Atrogin-1, MuRF1 and LC3 genes. Extensor digitorum longus muscles and C2C12 myotubes exhibited a reduction in protein turnover. Cardiomyocytes showed an increase in proteolysis by activating autophagy and the ubiquitin proteasome system, and a decrease in protein synthesis by decreasing P70S6K. The effect of myostatin on protein metabolism is related to fiber type composition, which may be associated to the extent of atrophy mediated effect of myostatin on muscle.

  11. Protein metabolism in Turner syndrome and the impact of hormone replacement therapy.

    Science.gov (United States)

    Gravholt, Claus Højbjerg; Riis, Anne Lene; Møller, Niels; Christiansen, Jens Sandahl

    2007-09-01

    Studies have documented an altered body composition in Turner syndrome (TS). Body fat is increased and muscle mass is decreased. Ovarian failure necessitates substitution with female hormone replacement therapy (HRT), and HRT induces favourable changes in body composition. It is unknown how HRT affects protein metabolism. To test whether alterations in body composition before and after HRT in TS are a result of altered protein metabolism. We performed a randomized crossover study with active treatment (HRT in TS and oral contraceptives in controls) or no treatment. We studied eight women (age 29.7 +/- 5.6 (mean +/- SD) years) with TS, verified by karyotype, and eight age-matched controls (age 27.3 +/- 4.9 years). All subjects underwent a 3-h study in the postabsorptive state. Protein dynamics of the whole body and of the forearm muscles were measured by an amino acid tracer dilution technique using [(15)N]phenylalanine and [(2)H(4)]tyrosine. Substrate metabolism was examined by indirect calorimetry. Energy expenditure was comparable among TS and controls, and did not change during active treatment. Whole-body phenylalanine and tyrosine fluxes were similar in the untreated situations, and did not change during active treatment. Amino acid degradation and protein synthesis were similar in all situations. Muscle protein breakdown was similar among groups, and was not affected by treatment. Muscle protein synthesis rate and forearm blood flow did not differ among groups or due to treatment. Protein metabolism in TS is comparable to controls, and is not affected by HRT.

  12. Regulation of lifespan, metabolism, and stress responses by the Drosophila SH2B protein, Lnk.

    Directory of Open Access Journals (Sweden)

    Cathy Slack

    2010-03-01

    Full Text Available Drosophila Lnk is the single ancestral orthologue of a highly conserved family of structurally-related intracellular adaptor proteins, the SH2B proteins. As adaptors, they lack catalytic activity but contain several protein-protein interaction domains, thus playing a critical role in signal transduction from receptor tyrosine kinases to form protein networks. Physiological studies of SH2B function in mammals have produced conflicting data. However, a recent study in Drosophila has shown that Lnk is an important regulator of the insulin/insulin-like growth factor (IGF-1 signaling (IIS pathway during growth, functioning in parallel to the insulin receptor substrate, Chico. As this pathway also has an evolutionary conserved role in the determination of organism lifespan, we investigated whether Lnk is required for normal lifespan in Drosophila. Phenotypic analysis of mutants for Lnk revealed that loss of Lnk function results in increased lifespan and improved survival under conditions of oxidative stress and starvation. Starvation resistance was found to be associated with increased metabolic stores of carbohydrates and lipids indicative of impaired metabolism. Biochemical and genetic data suggest that Lnk functions in both the IIS and Ras/Mitogen activated protein Kinase (MapK signaling pathways. Microarray studies support this model, showing transcriptional feedback onto genes in both pathways as well as indicating global changes in both lipid and carbohydrate metabolism. Finally, our data also suggest that Lnk itself may be a direct target of the IIS responsive transcription factor, dFoxo, and that dFoxo may repress Lnk expression. We therefore describe novel functions for a member of the SH2B protein family and provide the first evidence for potential mechanisms of SH2B regulation. Our findings suggest that IIS signaling in Drosophila may require the activity of a second intracellular adaptor, thereby yielding fundamental new insights into the

  13. Impact of weight loss and maintenance with ad libitum diets varying in protein and glycemic index content on metabolic syndrome

    DEFF Research Database (Denmark)

    Papadaki, Angeliki; Linardakis, Manolis; Plada, Maria

    2014-01-01

    We investigated the effects of weight loss and maintenance with diets that varied with regard to protein content and glycemic index (GI) on metabolic syndrome (MetSyn) status.......We investigated the effects of weight loss and maintenance with diets that varied with regard to protein content and glycemic index (GI) on metabolic syndrome (MetSyn) status....

  14. New method for the quality check of food proteins of the maintenance metabolism. 4

    International Nuclear Information System (INIS)

    Simon, O.; Hernandez, M.; Bergner, H.

    1981-01-01

    Male adult rats (370 g body weight) were fed on maintenance level (460 kJ ME/kgsup(0,75). In a 10 days preliminary period they received a casein/methionine (95/5) diet supplemented with 10 mg 15 N excess per 0.178 kg metabolic body weight in form of ammonium acetate. Thereafter the animals were put on 8 isonitrogenous diets containing as protein sources casein, soya protein, gelatine, whole-egg, fish meal, pea, wheat and yeast. The 15 N excretion via urine and feces was used to evaluate the dietary proteins for maintenance. 15 N in urine was lowest in animals fed on wheat diet and highest after feeding whole-egg diet. From these data a so called ' 15 N excretion biological valence (BV)' was calculated, which indicated the highest quality for wheat and soy protein in meeting the needs of the intermediary maintenance metabolism. On the other hand, dietary protein sources influence the loss of endogenous nitrogen as metabolic fecal nitrogen (MFN). It was found to be lowest in animals fed on diets containing isolated proteins (6 mg MFN/100 g body weight) and highest after feeding protein sources of plant origin with a high content in crude fibre (10 mg MFN/100 g). Both, losses of 15 N via urine and via feces were combined in a parameter called 'total BV'. According to this parameter the differences in quality for maintenance were only little between the protein sources tested (casein 100, soy protein 100, pea 99, wheat 99, whole egg 92, fish meal 89, gelatin 89). It was concluded that in the state of maintenance the supply with essential amino acids is not critical and that the supply with dispensable amino acids (or nonspecific nitrogen) is of great importance. (author)

  15. Effect of protein malnutrition on the metabolism of bone collagen in albino rats

    Energy Technology Data Exchange (ETDEWEB)

    Rao, J S; Rao, V H [Central Leather Research Inst., Madras (India)

    1981-01-01

    The effect of protein malnutrition on the metabolism of collagen in bone was studied in young female albino rats after a single injection of /sup 3/H-proline. Both specific and total radioactivities of hydroxyproline in the total collagen of the bone were found to decrease in the protein-deficient animals, indicating decreased rate of collagen synthesis. In the urine the amount of hydroxyproline excreted and total radioactivity of /sup 3/H-hydroxyproline were greatly decreased. The results of the present investigation therefore clearly indicate decreased synthesis and catabolism of collagen in bones of protein deficient animals compared to controls.

  16. Hepatitis B virus core protein allosteric modulators can distort and disrupt intact capsids.

    Science.gov (United States)

    Schlicksup, Christopher John; Wang, Joseph Che-Yen; Francis, Samson; Venkatakrishnan, Balasubramanian; Turner, William W; VanNieuwenhze, Michael; Zlotnick, Adam

    2018-01-29

    Defining mechanisms of direct-acting antivirals facilitates drug development and our understanding of virus function. Heteroaryldihydropyrimidines (HAPs) inappropriately activate assembly of hepatitis B virus (HBV) core protein (Cp), suppressing formation of virions. We examined a fluorophore-labeled HAP, HAP-TAMRA. HAP-TAMRA induced Cp assembly and also bound pre-assembled capsids. Kinetic and spectroscopic studies imply that HAP-binding sites are usually not available but are bound cooperatively. Using cryo-EM, we observed that HAP-TAMRA asymmetrically deformed capsids, creating a heterogeneous array of sharp angles, flat regions, and outright breaks. To achieve high resolution reconstruction (HAP-TAMRA caused quasi-sixfold vertices to become flatter and fivefold more angular. This transition led to asymmetric faceting. That a disordered crosslink could rescue symmetry implies that capsids have tensegrity properties. Capsid distortion and disruption is a new mechanism by which molecules like the HAPs can block HBV infection. © 2017, Schlicksup et al.

  17. Hepatitis B virus core protein allosteric modulators can distort and disrupt intact capsids

    Science.gov (United States)

    Schlicksup, Christopher John; Wang, Joseph Che-Yen; Francis, Samson; Venkatakrishnan, Balasubramanian; Turner, William W; VanNieuwenhze, Michael

    2018-01-01

    Defining mechanisms of direct-acting antivirals facilitates drug development and our understanding of virus function. Heteroaryldihydropyrimidines (HAPs) inappropriately activate assembly of hepatitis B virus (HBV) core protein (Cp), suppressing formation of virions. We examined a fluorophore-labeled HAP, HAP-TAMRA. HAP-TAMRA induced Cp assembly and also bound pre-assembled capsids. Kinetic and spectroscopic studies imply that HAP-binding sites are usually not available but are bound cooperatively. Using cryo-EM, we observed that HAP-TAMRA asymmetrically deformed capsids, creating a heterogeneous array of sharp angles, flat regions, and outright breaks. To achieve high resolution reconstruction (particle symmetry. We deduced that HAP-TAMRA caused quasi-sixfold vertices to become flatter and fivefold more angular. This transition led to asymmetric faceting. That a disordered crosslink could rescue symmetry implies that capsids have tensegrity properties. Capsid distortion and disruption is a new mechanism by which molecules like the HAPs can block HBV infection. PMID:29377794

  18. The H1 linker histones: multifunctional proteins beyond the nucleosomal core particle.

    Science.gov (United States)

    Hergeth, Sonja P; Schneider, Robert

    2015-11-01

    The linker histone H1 family members are a key component of chromatin and bind to the nucleosomal core particle around the DNA entry and exit sites. H1 can stabilize both nucleosome structure and higher-order chromatin architecture. In general, H1 molecules consist of a central globular domain with more flexible tail regions at both their N- and C-terminal ends. The existence of multiple H1 subtypes and a large variety of posttranslational modifications brings about a considerable degree of complexity and makes studying this protein family challenging. Here, we review recent progress in understanding the function of linker histones and their subtypes beyond their role as merely structural chromatin components. We summarize current findings on the role of H1 in heterochromatin formation, transcriptional regulation and embryogenesis with a focus on H1 subtypes and their specific modifications. © 2015 The Authors.

  19. TRF2 Protein Interacts with Core Histones to Stabilize Chromosome Ends.

    Science.gov (United States)

    Konishi, Akimitsu; Izumi, Takashi; Shimizu, Shigeomi

    2016-09-23

    Mammalian chromosome ends are protected by a specialized nucleoprotein complex called telomeres. Both shelterin, a telomere-specific multi-protein complex, and higher order telomeric chromatin structures combine to stabilize the chromosome ends. Here, we showed that TRF2, a component of shelterin, binds to core histones to protect chromosome ends from inappropriate DNA damage response and loss of telomeric DNA. The N-terminal Gly/Arg-rich domain (GAR domain) of TRF2 directly binds to the globular domain of core histones. The conserved arginine residues in the GAR domain of TRF2 are required for this interaction. A TRF2 mutant with these arginine residues substituted by alanine lost the ability to protect telomeres and induced rapid telomere shortening caused by the cleavage of a loop structure of the telomeric chromatin. These findings showed a previously unnoticed interaction between the shelterin complex and nucleosomal histones to stabilize the chromosome ends. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Insulin resistance and protein energy metabolism in patients with advanced chronic kidney disease.

    Science.gov (United States)

    Siew, Edward D; Ikizler, Talat Alp

    2010-01-01

    Insulin resistance (IR), the reciprocal of insulin sensitivity is a known complication of advanced chronic kidney disease (CKD) and is associated with a number of metabolic derangements. The complex metabolic abnormalities observed in CKD such as vitamin D deficiency, obesity, metabolic acidosis, inflammation, and accumulation of "uremic toxins" are believed to contribute to the etiology of IR and acquired defects in the insulin-receptor signaling pathway in this patient population. Only a few investigations have explored the validity of commonly used assessment methods in comparison to gold standard hyperinsulinemic hyperglycemic clamp technique in CKD patients. An important consequence of insulin resistance is its role in the pathogenesis of protein energy wasting, a state of metabolic derangement characterized by loss of somatic and visceral protein stores not entirely accounted for by inadequate nutrient intake. In the general population, insulin resistance has been associated with accelerated protein catabolism. Among end-stage renal disease (ESRD) patients, enhanced muscle protein breakdown has been observed in patients with Type II diabetes compared to ESRD patients without diabetes. In the absence of diabetes mellitus (DM) or severe obesity, insulin resistance is detectable in dialysis patients and strongly associated with increased muscle protein breakdown, primarily mediated by the ubiquitin-proteasome pathway. Recent epidemiological data indicate a survival advantage and better nutritional status in insulin-free Type II DM patients treated with insulin sensitizer thiazolidinediones. Given the high prevalence of protein energy wasting in ESRD and its unequivocal association with adverse clinical outcomes, insulin resistance may represent an important modifiable target for intervention in the ESRD population.

  1. Metabolism

    Science.gov (United States)

    ... Are More Common in People With Type 1 Diabetes Metabolic Syndrome Your Child's Weight Healthy Eating Endocrine System Blood Test: Basic Metabolic Panel (BMP) Activity: Endocrine System Growth Disorders Diabetes Center Thyroid Disorders Your Endocrine System Movie: Endocrine ...

  2. Prioritizing Popular Proteins in Liver Cancer: Remodelling One-Carbon Metabolism.

    Science.gov (United States)

    Mora, María Isabel; Molina, Manuela; Odriozola, Leticia; Elortza, Félix; Mato, José María; Sitek, Barbara; Zhang, Pumin; He, Fuchu; Latasa, María Uxue; Ávila, Matías Antonio; Corrales, Fernando José

    2017-12-01

    Primary liver cancer (HCC) is recognized as the fifth most common neoplasm and the second leading cause of cancer death worldwide. Most risk factors are known, and the molecular pathogenesis has been widely studied in the past decade; however, the underlying molecular mechanisms remain to be unveiled, as they will facilitate the definition of novel biomarkers and clinical targets for more effective patient management. We utilize the B/D-HPP popular protein strategy. We report a list of popular proteins that have been highly cocited with the expression "liver cancer". Several enzymes highlight the known metabolic remodeling of liver cancer cells, four of which participate in one-carbon metabolism. This pathway is central to the maintenance of differentiated hepatocytes, as it is considered the connection between intermediate metabolism and epigenetic regulation. We designed a targeted selective reaction monitoring (SRM) method to follow up one-carbon metabolism adaptation in mouse HCC and in regenerating liver following exposure to CCl 4 . This method allows systematic monitoring of one-carbon metabolism and could prove useful in the follow-up of HCC and of chronically liver-diseased patients (cirrhosis) at risk of HCC. The SRM data are available via ProteomeXchange in PASSEL (PASS01060).

  3. Protein metabolism in obese patients during very low-calorie mixed diets containing different amounts of proteins and carbohydrates.

    Science.gov (United States)

    Pasquali, R; Casimirri, F; Melchionda, N

    1987-12-01

    To assess long-term nitrogen sparing capacity of very low-calorie mixed diets, we administered two isoenergetic (2092KJ) liquid formula regimens of different composition for 8 weeks to two matched groups of massively obese patients (group 1: proteins 60 g, carbohydrate 54 g; group 2: proteins 41 g, carbohydrates 81 g). Weight loss was similar in both groups. Daily nitrogen balance (g) during the second month resulted more a negative in group 2 with respect to group 1. However, within the groups individual nitrogen sparing capacity varied markedly; only a few in group 1 and one in group 2 were able to attain nitrogen equilibrium throughout the study. Daily urine excretion of 3-methylhistidine fell significantly in group 1 but did not change in group 2. Unlike total proteins, albumins, and transferrin, serum levels of retinol-binding protein, thyroxin-binding globulin, and complement-C3 fell significantly in both groups but per cent variations of complement-C3 were more pronounced in the first group. Prealbumin levels fell persistently in group 1 and transiently in group 2. The results indicate that even with this type of diet an adequate amount of dietary protein represents the most important factor in minimizing whole body protein catabolism during long-term semistarvation in massively obese patients. Moreover, they confirm the possible role of dietary carbohydrates in the regulation of some visceral protein metabolism.

  4. [The effect of copper on the metabolism of iodine, carbohydrates and proteins in rats].

    Science.gov (United States)

    Esipenko, B E; Marsakova, N V

    1990-01-01

    Experiments on 156 rats maintained at ration with copper deficiency have demonstrated a decrease in the values of iodine metabolism in organs and tissues excluding the liver where a sharp increase in the concentration and content of inorganic iodine was observed. A disturbance in indices of carbohydrate and proteins metabolism in the organism of animals is marked. A direct relationship with a correlation coefficient equaling 0.87-1.00 is determined between changes in the concentration of protein-bound iodine in blood and concentration of glycogen in the liver, skeletal muscles, albumins, alpha 1-, alpha 2-globulins, urea concentration; an inverse relationship with glucose, activity of blood lipo-dehydrogenase and liver mitochondria, aldolase, concentration of pyruvic and lactic acids is established as well. It is concluded that copper deficiency can exert both a direct effect on metabolic processes (as data from literature testify) and an indirect one disturbing iodine metabolism, i. e. sharply decreasing protein-bound iodine production by the thyroid gland.

  5. Unravelling core microbial metabolisms in the hypersaline microbial mats of Shark Bay using high-throughput metagenomics

    Energy Technology Data Exchange (ETDEWEB)

    Ruvindy, Rendy; White III, Richard Allen; Neilan, Brett Anthony; Burns, Brendan Paul

    2015-05-29

    Modern microbial mats are potential analogues of some of Earth’s earliest ecosystems. Excellent examples can be found in Shark Bay, Australia, with mats of various morphologies. To further our understanding of the functional genetic potential of these complex microbial ecosystems, we conducted for the first time shotgun metagenomic analyses. We assembled metagenomic nextgeneration sequencing data to classify the taxonomic and metabolic potential across diverse morphologies of marine mats in Shark Bay. The microbial community across taxonomic classifications using protein-coding and small subunit rRNA genes directly extracted from the metagenomes suggests that three phyla Proteobacteria, Cyanobacteria and Bacteriodetes dominate all marine mats. However, the microbial community structure between Shark Bay and Highbourne Cay (Bahamas) marine systems appears to be distinct from each other. The metabolic potential (based on SEED subsystem classifications) of the Shark Bay and Highbourne Cay microbial communities were also distinct. Shark Bay metagenomes have a metabolic pathway profile consisting of both heterotrophic and photosynthetic pathways, whereas Highbourne Cay appears to be dominated almost exclusively by photosynthetic pathways. Alternative non-rubisco-based carbon metabolism including reductive TCA cycle and 3-hydroxypropionate/4-hydroxybutyrate pathways is highly represented in Shark Bay metagenomes while not represented in Highbourne Cay microbial mats or any other mat forming ecosystems investigated to date. Potentially novel aspects of nitrogen cycling were also observed, as well as putative heavy metal cycling (arsenic, mercury, copper and cadmium). Finally, archaea are highly represented in Shark Bay and may have critical roles in overall ecosystem function in these modern microbial mats.

  6. Hypolipidemic effect of dietary pea proteins: Impact on genes regulating hepatic lipid metabolism.

    Science.gov (United States)

    Rigamonti, Elena; Parolini, Cinzia; Marchesi, Marta; Diani, Erika; Brambilla, Stefano; Sirtori, Cesare R; Chiesa, Giulia

    2010-05-01

    Controversial data on the lipid-lowering effect of dietary pea proteins have been provided and the mechanisms behind this effect are not completely understood. The aim of the study was to evaluate a possible hypolipidemic activity of a pea protein isolate and to determine whether pea proteins could affect the hepatic lipid metabolism through regulation of genes involved in cholesterol and fatty acid homeostasis. Rats were fed Nath's hypercholesterolemic diets for 28 days, the protein sources being casein or a pea protein isolate from Pisum sativum. After 14 and 28 days of dietary treatment, rats fed pea proteins had markedly lower plasma cholesterol and triglyceride levels than rats fed casein (pPea protein-fed rats displayed higher hepatic mRNA levels of LDL receptor versus those fed casein (ppea protein-fed rats than in rats fed casein (ppea proteins in rats. Moreover, pea proteins appear to affect cellular lipid homeostasis by upregulating genes involved in hepatic cholesterol uptake and by downregulating fatty acid synthesis genes.

  7. Whey Protein Supplementation Enhances Whole Body Protein Metabolism and Performance Recovery after Resistance Exercise: A Double-Blind Crossover Study.

    Science.gov (United States)

    West, Daniel W D; Abou Sawan, Sidney; Mazzulla, Michael; Williamson, Eric; Moore, Daniel R

    2017-07-11

    No study has concurrently measured changes in free-living whole body protein metabolism and exercise performance during recovery from an acute bout of resistance exercise. We aimed to determine if whey protein ingestion enhances whole body net protein balance and recovery of exercise performance during overnight (10 h) and 24 h recovery after whole body resistance exercise in trained men. In a double-blind crossover design, 12 trained men (76 ± 8 kg, 24 ± 4 years old, 14% ± 5% body fat; means ± standard deviation (SD)) performed resistance exercise in the evening prior to consuming either 25 g of whey protein (PRO; MuscleTech 100% Whey) or an energy-matched placebo (CHO) immediately post-exercise (0 h), and again the following morning (~10 h of recovery). A third randomized trial, completed by the same participants, involving no exercise and no supplement served as a rested control trial (Rest). Participants ingested [ 15 N]glycine to determine whole body protein kinetics and net protein balance over 10 and 24 h of recovery. Performance was assessed pre-exercise and at 0, 10, and 24 h of recovery using a battery of tests. Net protein balance tended to improve in PRO ( P = 0.064; effect size (ES) = 0.61, PRO vs. CHO) during overnight recovery. Over 24 h, net balance was enhanced in PRO ( P = 0.036) but not in CHO ( P = 0.84; ES = 0.69, PRO vs. CHO), which was mediated primarily by a reduction in protein breakdown (PRO protein supplementation improved MVC (ES = 0.76), REP (ES = 0.44), and peak power (ES = 0.55). In conclusion, whey protein supplementation enhances whole body anabolism, and may improve acute recovery of exercise performance after a strenuous bout of resistance exercise.

  8. Effects of Human C-Reactive Protein on Pathogenesis of Features of the Metabolic Syndrome

    Czech Academy of Sciences Publication Activity Database

    Pravenec, Michal; Kajiya, T.; Zídek, Václav; Landa, Vladimír; Mlejnek, Petr; Šimáková, Miroslava; Šilhavý, Jan; Malínská, H.; Oliyarnyk, O.; Kazdová, L.; Fan, J.; Wang, J.; Kurtz, T. W.

    2011-01-01

    Roč. 57, č. 4 (2011), s. 731-737 ISSN 0194-911X R&D Projects: GA MZd(CZ) NS9759; GA MŠk(CZ) ME08006; GA MŠk(CZ) 1M0520; GA ČR(CZ) GAP301/10/0290; GA ČR GAP303/10/0505; GA AV ČR(CZ) IAA500110805 Grant - others:EC(XE) HEALTH-F4-2010-241504 Institutional research plan: CEZ:AV0Z50110509 Keywords : C-reactive protein * metabolic syndrome * transgenic rat Subject RIV: FB - Endocrinology, Diabetology, Metabolism, Nutrition Impact factor: 6.207, year: 2011

  9. Putative drug and vaccine target protein identification using comparative genomic analysis of KEGG annotated metabolic pathways of Mycoplasma hyopneumoniae.

    Science.gov (United States)

    Damte, Dereje; Suh, Joo-Won; Lee, Seung-Jin; Yohannes, Sileshi Belew; Hossain, Md Akil; Park, Seung-Chun

    2013-07-01

    In the present study, a computational comparative and subtractive genomic/proteomic analysis aimed at the identification of putative therapeutic target and vaccine candidate proteins from Kyoto Encyclopedia of Genes and Genomes (KEGG) annotated metabolic pathways of Mycoplasma hyopneumoniae was performed for drug design and vaccine production pipelines against M.hyopneumoniae. The employed comparative genomic and metabolic pathway analysis with a predefined computational systemic workflow extracted a total of 41 annotated metabolic pathways from KEGG among which five were unique to M. hyopneumoniae. A total of 234 proteins were identified to be involved in these metabolic pathways. Although 125 non homologous and predicted essential proteins were found from the total that could serve as potential drug targets and vaccine candidates, additional prioritizing parameters characterize 21 proteins as vaccine candidate while druggability of each of the identified proteins evaluated by the DrugBank database prioritized 42 proteins suitable for drug targets. Copyright © 2013 Elsevier Inc. All rights reserved.

  10. Expression of Lipid Metabolism-Related Proteins Differs between Invasive Lobular Carcinoma and Invasive Ductal Carcinoma.

    Science.gov (United States)

    Cha, Yoon Jin; Kim, Hye Min; Koo, Ja Seung

    2017-01-23

    We comparatively investigated the expression and clinical implications of lipid metabolism-related proteins in invasive lobular carcinoma (ILC) and invasive ductal carcinoma (IDC) of the breast. A total of 584 breast cancers (108 ILC and 476 IDC) were subjected to tissue microarray and immunohistochemical analysis for lipid metabolism-related proteins including hormone-sensitive lipase (HSL), perilipin A, fatty acid binding protein (FABP)4, carnitine palmitoyltransferase (CPT)-1, acyl-CoA oxidase 1, and fatty acid synthetase (FASN). HSL, perilipin A, and FABP4 expression (all p invasive cancers, HSL and FABP4 were highly expressed in luminal A-type ILC ( p cancers, HSL and FABP4 were more highly expressed in ILC ( p < 0.001). Univariate analysis found associations of shorter disease-free survival with CPT-1 positivity ( p = 0.004) and acyl-CoA oxidase 1 positivity ( p = 0.032) and of shorter overall survival with acyl-CoA oxidase 1 positivity ( p = 0.027). In conclusion, ILC and IDC exhibited different immunohistochemical lipid metabolism-related protein expression profiles. Notably, ILC exhibited high HSL and FABP4 and low perilipin A expression.

  11. Expression of Lipid Metabolism-Related Proteins Differs between Invasive Lobular Carcinoma and Invasive Ductal Carcinoma

    Directory of Open Access Journals (Sweden)

    Yoon Jin Cha

    2017-01-01

    Full Text Available We comparatively investigated the expression and clinical implications of lipid metabolism-related proteins in invasive lobular carcinoma (ILC and invasive ductal carcinoma (IDC of the breast. A total of 584 breast cancers (108 ILC and 476 IDC were subjected to tissue microarray and immunohistochemical analysis for lipid metabolism-related proteins including hormone-sensitive lipase (HSL, perilipin A, fatty acid binding protein (FABP4, carnitine palmitoyltransferase (CPT-1, acyl-CoA oxidase 1, and fatty acid synthetase (FASN. HSL, perilipin A, and FABP4 expression (all p < 0.001 differed significantly: HSL and FABP4 were more frequently present in ILC, whereas perilipin A was more frequently detected in IDC. Among all invasive cancers, HSL and FABP4 were highly expressed in luminal A-type ILC (p < 0.001 and perilipin A in luminal A-type IDC (p = 0.007. Among luminal B-type cancers, HSL and FABP4 were more highly expressed in ILC (p < 0.001. Univariate analysis found associations of shorter disease-free survival with CPT-1 positivity (p = 0.004 and acyl-CoA oxidase 1 positivity (p = 0.032 and of shorter overall survival with acyl-CoA oxidase 1 positivity (p = 0.027. In conclusion, ILC and IDC exhibited different immunohistochemical lipid metabolism-related protein expression profiles. Notably, ILC exhibited high HSL and FABP4 and low perilipin A expression.

  12. [L-arginine metabolism enzyme activities in rat liver subcellular fractions under condition of protein deprivation].

    Science.gov (United States)

    Kopyl'chuk, G P; Buchkovskaia, I M

    2014-01-01

    The features of arginase and NO-synthase pathways of arginine's metabolism have been studied in rat liver subcellular fractions under condition of protein deprivation. During the experimental period (28 days) albino male rats were kept on semi synthetic casein diet AIN-93. The protein deprivation conditions were designed as total absence of protein in the diet and consumption of the diet partially deprived with 1/2 of the casein amount compared to in the regular diet. Daily diet consumption was regulated according to the pair feeding approach. It has been shown that the changes of enzyme activities, involved in L-arginine metabolism, were characterized by 1.4-1.7 fold decrease in arginase activity, accompanied with unchanged NO-synthase activity in cytosol. In mitochondrial fraction the unchanged arginase activity was accompanied by 3-5 fold increase of NO-synthase activity. At the terminal stages of the experiment the monodirectional dynamics in the studied activities have been observed in the mitochondrial and cytosolfractions in both experimental groups. In the studied subcellular fractions arginase activity decreased (2.4-2.7 fold with no protein in the diet and 1.5 fold with partly supplied protein) and was accompanied by NO-synthase activity increase by 3.8 fold in cytosole fraction, by 7.2 fold in mitochondrial fraction in the group with no protein in the diet and by 2.2 and 3.5 fold in the group partialy supplied with protein respectively. The observed tendency is presumably caused by the switch of L-arginine metabolism from arginase into oxidizing NO-synthase parthway.

  13. "Hot cores" in proteins: Comparative analysis of the apolar contact area in structures from hyper/thermophilic and mesophilic organisms

    Directory of Open Access Journals (Sweden)

    Bossa Francesco

    2008-02-01

    Full Text Available Abstract Background A wide variety of stabilizing factors have been invoked so far to elucidate the structural basis of protein thermostability. These include, amongst the others, a higher number of ion-pairs interactions and hydrogen bonds, together with a better packing of hydrophobic residues. It has been frequently observed that packing of hydrophobic side chains is improved in hyperthermophilic proteins, when compared to their mesophilic counterparts. In this work, protein crystal structures from hyper/thermophilic organisms and their mesophilic homologs have been compared, in order to quantify the difference of apolar contact area and to assess the role played by the hydrophobic contacts in the stabilization of the protein core, at high temperatures. Results The construction of two datasets was carried out so as to satisfy several restrictive criteria, such as minimum redundancy, resolution and R-value thresholds and lack of any structural defect in the collected structures. This approach allowed to quantify with relatively high precision the apolar contact area between interacting residues, reducing the uncertainty due to the position of atoms in the crystal structures, the redundancy of data and the size of the dataset. To identify the common core regions of these proteins, the study was focused on segments that conserve a similar main chain conformation in the structures analyzed, excluding the intervening regions whose structure differs markedly. The results indicated that hyperthermophilic proteins underwent a significant increase of the hydrophobic contact area contributed by those residues composing the alpha-helices of the structurally conserved regions. Conclusion This study indicates the decreased flexibility of alpha-helices in proteins core as a major factor contributing to the enhanced termostability of a number of hyperthermophilic proteins. This effect, in turn, may be due to an increased number of buried methyl groups in

  14. Mechanisms and Effects on HBV Replication of the Interaction between HBV Core Protein and Cellular Filamin B.

    Science.gov (United States)

    Li, Yilin; Sun, Yishuang; Sun, Fuyun; Hua, Rong; Li, Chenlin; Chen, Lang; Guo, Deyin; Mu, Jingfang

    2018-03-28

    Hepatitis B virus (HBV) infection is one of the major problems that threatens global health. There have been many studies on HBV, but the relationship between HBV and host factors is largely unexplored and more studies are needed to clarify these interactions. Filamin B is an actin-binding protein that acts as a cytoskeleton protein, and it is involved in cell development and several signaling pathways. In this study, we showed that filamin B interacted with HBV core protein, and the interaction promoted HBV replication. The interaction between filamin B and core protein was observed in HEK 293T, Huh7 and HepG2 cell lines by co-immunoprecipitation and co-localization immnofluoresence. Overexpression of filamin B increased the levels of HBV total RNAs and pre-genome RNA (pgRNA), and improved the secretion level of hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg). In contrast, filamin B knockdown inhibited HBV replication, decreased the level of HBV total RNAs and pgRNA, and reduced the secretion level of HBsAg and HBeAg. In addition, we found that filamin B and core protein may interact with each other via four blocks of argentine residues at the C-terminus of core protein. In conclusion, we identify filamin B as a novel host factor that can interact with core protein to promote HBV replication in hepatocytes. Our study provides new insights into the relationship between HBV and host factors and may provide new strategies for the treatment of HBV infection.

  15. Cloning and sequencing of the cDNA encoding a core protein of the paired helical filament of Alzheimer's disease: Identification as the microtubule-associated protein tau

    International Nuclear Information System (INIS)

    Goedert, M.; Wischik, C.M.; Crowther, R.A.; Walker, J.E.; Klug, A.

    1988-01-01

    Screening of cDNA libraries prepared from the frontal cortex of an Alzheimer's disease patient and from fetal human brain has led to isolation of the cDNA for a core protein of the paired helical filament of Alzheimer's disease. The partial amino acid sequence of this core protein was used to design synthetic oligonucleotide probes. The cDNA encodes a protein of 352 amino acids that contains a characteristic amino acid repeat in its carboxyl-terminal half. This protein is highly homologous to the sequence of the mouse microtubule-associated protein tau and thus constitutes the human equivalent of mouse tau. RNA blot analysis indicates the presence of two major transcripts, 6 and 2 kilobases long, with a wide distribution in normal human brain. Tau protein mRNAs were found in normal amounts in the frontal cortex from patients with Alzheimer's disease. The proof that at least part of tau protein forms a component of the paired helical filament core opens the way to understanding the mode of formation of paired helical filaments and thus, ultimately, the pathogenesis of Alzheimer's disease

  16. Protein carbonylation associated to high-fat, high-sucrose diet and its metabolic effects.

    Science.gov (United States)

    Méndez, Lucía; Pazos, Manuel; Molinar-Toribio, Eunice; Sánchez-Martos, Vanesa; Gallardo, José M; Rosa Nogués, M; Torres, Josep L; Medina, Isabel

    2014-12-01

    The present research draws a map of the characteristic carbonylation of proteins in rats fed high-caloric diets with the aim of providing a new insight of the pathogenesis of metabolic diseases derived from the high consumption of fat and refined carbohydrates. Protein carbonylation was analyzed in plasma, liver and skeletal muscle of Sprague-Dawley rats fed a high-fat, high-sucrose (HFHS) diet by a proteomics approach based on carbonyl-specific fluorescence-labeling, gel electrophoresis and mass spectrometry. Oxidized proteins along with specific sites of oxidative damage were identified and discussed to illustrate the consequences of protein oxidation. The results indicated that long-term HFHS consumption increased protein oxidation in plasma and liver; meanwhile, protein carbonyls from skeletal muscle did not change. The increment of carbonylation by HFHS diet was singularly selective on specific target proteins: albumin from plasma and liver, and hepatic proteins such as mitochondrial carbamoyl-phosphate synthase (ammonia), mitochondrial aldehyde dehydrogenase, argininosuccinate synthetase, regucalcin, mitochondrial adenosine triphosphate synthase subunit beta, actin cytoplasmic 1 and mitochondrial glutamate dehydrogenase 1. The possible consequences that these specific protein carbonylations have on the excessive weight gain, insulin resistance and nonalcoholic fatty liver disease resulting from HFHS diet consumption are discussed. Copyright © 2014 Elsevier Inc. All rights reserved.

  17. The Copper Metabolism MURR1 Domain protein 1 (COMMD1) modulates the aggregation of misfolded protein species in a client-specific manner

    NARCIS (Netherlands)

    W.I.M. Vonk (Willianne I.); V. Kakkar (Vaishali); P. Bartuzi (Paulina); D. Jaarsma (Dick); R. Berger (Ruud); M.A. Hofker (Marten); L.W.J. Klomp (Leo W.); C. Wijmenga (Cisca); H. Kampinga (Harm); B. van de Sluis (Bart)

    2014-01-01

    textabstractThe Copper Metabolism MURR1 domain protein 1 (COMMD1) is a protein involved in multiple cellular pathways, including copper homeostasis, NF-κB and hypoxia signalling. Acting as a scaffold protein, COMMD1 mediates the levels, stability and proteolysis of its substrates (e.g. the

  18. Short communication: Proteins from circulating exosomes represent metabolic state in transition dairy cows.

    Science.gov (United States)

    Crookenden, M A; Walker, C G; Peiris, H; Koh, Y; Heiser, A; Loor, J J; Moyes, K M; Murray, A; Dukkipati, V S R; Kay, J K; Meier, S; Roche, J R; Mitchell, M D

    2016-09-01

    Biomarkers that identify prepathological disease could enhance preventive management, improve animal health and productivity, and reduce costs. Circulating extracellular vesicles, particularly exosomes, are considered to be long-distance, intercellular communication systems in human medicine. Exosomes provide tissue-specific messages of functional state and can alter the cellular activity of recipient tissues through their protein and microRNA content. We hypothesized that exosomes circulating in the blood of cows during early lactation would contain proteins representative of the metabolic state of important tissues, such as liver, which play integral roles in regulating the physiology of cows postpartum. From a total of 150 cows of known metabolic phenotype, 10 cows were selected with high (n=5; high risk) and low (n=5; low risk) concentrations of nonesterified fatty acids, β-hydroxybutyrate, and liver triacylglycerol during wk 1 and 2 after calving. Exosomes were extracted from blood on the day of calving (d 0) and postcalving at wk 1 and wk 4, and their protein composition was determined by mass spectroscopy. Extracellular vesicle protein concentration and the number of exosome vesicles were not affected by risk category; however, the exosome protein cargo differed between the groups, with proteins at each time point identified as being unique to the high- and low-risk groups. The proteins α-2 macroglobulin, fibrinogen, and oncoprotein-induced transcript 3 were unique to the high-risk cows on d 0 and have been associated with metabolic syndrome and liver function in humans. Their presence may indicate a more severe inflammatory state and a greater degree of liver dysfunction in the high-risk cows than in the low-risk cows, consistent with the high-risk cows' greater plasma β-hydroxybutyrate and liver triacylglycerol concentrations. The commonly shared proteins and those unique to the low-risk category indicate a role for exosomes in immune function. The data

  19. Reduced endothelial thioredoxin-interacting protein protects arteries from damage induced by metabolic stress in vivo.

    Science.gov (United States)

    Bedarida, Tatiana; Domingues, Alison; Baron, Stephanie; Ferreira, Chrystophe; Vibert, Francoise; Cottart, Charles-Henry; Paul, Jean-Louis; Escriou, Virginie; Bigey, Pascal; Gaussem, Pascale; Leguillier, Teddy; Nivet-Antoine, Valerie

    2018-06-01

    Although thioredoxin-interacting protein (TXNIP) is involved in a variety of biologic functions, the contribution of endothelial TXNIP has not been well defined. To investigate the endothelial function of TXNIP, we generated a TXNIP knockout mouse on the Cdh5-cre background (TXNIP fl/fl cdh5 cre ). Control (TXNIP fl/fl ) and TXNIP fl/fl cdh5 cre mice were fed a high protein-low carbohydrate (HP-LC) diet for 3 mo to induce metabolic stress. We found that TXNIP fl/fl and TXNIP fl/fl cdh5 cre mice on an HP-LC diet displayed impaired glucose tolerance and dyslipidemia concretizing the metabolic stress induced. We evaluated the impact of this metabolic stress on mice with reduced endothelial TXNIP expression with regard to arterial structure and function. TXNIP fl/fl cdh5 cre mice on an HP-LC diet exhibited less endothelial dysfunction than littermate mice on an HP-LC diet. These mice were protected from decreased aortic medial cell content, impaired aortic distensibility, and increased plasminogen activator inhibitor 1 secretion. This protective effect came with lower oxidative stress and lower inflammation, with a reduced NLRP3 inflammasome expression, leading to a decrease in cleaved IL-1β. We also show the major role of TXNIP in inflammation with a knockdown model, using a TXNIP-specific, small interfering RNA included in a lipoplex. These findings demonstrate a key role for endothelial TXNIP in arterial impairments induced by metabolic stress, making endothelial TXNIP a potential therapeutic target.-Bedarida, T., Domingues, A., Baron, S., Ferreira, C., Vibert, F., Cottart, C.-H., Paul, J.-L., Escriou, V., Bigey, P., Gaussem, P., Leguillier, T., Nivet-Antoine, V. Reduced endothelial thioredoxin-interacting protein protects arteries from damage induced by metabolic stress in vivo.

  20. Matrix rigidity regulates cancer cell growth by modulating cellular metabolism and protein synthesis.

    Directory of Open Access Journals (Sweden)

    Robert W Tilghman

    Full Text Available Tumor cells in vivo encounter diverse types of microenvironments both at the site of the primary tumor and at sites of distant metastases. Understanding how the various mechanical properties of these microenvironments affect the biology of tumor cells during disease progression is critical in identifying molecular targets for cancer therapy.This study uses flexible polyacrylamide gels as substrates for cell growth in conjunction with a novel proteomic approach to identify the properties of rigidity-dependent cancer cell lines that contribute to their differential growth on soft and rigid substrates. Compared to cells growing on more rigid/stiff substrates (>10,000 Pa, cells on soft substrates (150-300 Pa exhibited a longer cell cycle, due predominantly to an extension of the G1 phase of the cell cycle, and were metabolically less active, showing decreased levels of intracellular ATP and a marked reduction in protein synthesis. Using stable isotope labeling of amino acids in culture (SILAC and mass spectrometry, we measured the rates of protein synthesis of over 1200 cellular proteins under growth conditions on soft and rigid/stiff substrates. We identified cellular proteins whose syntheses were either preferentially inhibited or preserved on soft matrices. The former category included proteins that regulate cytoskeletal structures (e.g., tubulins and glycolysis (e.g., phosphofructokinase-1, whereas the latter category included proteins that regulate key metabolic pathways required for survival, e.g., nicotinamide phosphoribosyltransferase, a regulator of the NAD salvage pathway.The cellular properties of rigidity-dependent cancer cells growing on soft matrices are reminiscent of the properties of dormant cancer cells, e.g., slow growth rate and reduced metabolism. We suggest that the use of relatively soft gels as cell culture substrates would allow molecular pathways to be studied under conditions that reflect the different mechanical

  1. The Rab-GTPase-activating protein TBC1D1 regulates skeletal muscle glucose metabolism

    DEFF Research Database (Denmark)

    Szekeres, Ferenc; Chadt, Alexandra; Tom, Robby Z

    2012-01-01

    The Rab-GTPase-activating protein TBC1D1 has emerged as a novel candidate involved in metabolic regulation. Our aim was to determine whether TBC1D1 is involved in insulin as well as energy-sensing signals controlling skeletal muscle metabolism. TBC1D1-deficient congenic B6.SJL-Nob1.10 (Nob1.10(SJL...... be explained partly by a 50% reduction in GLUT4 protein, since proximal signaling at the level of Akt, AMPK, and acetyl-CoA carboxylase (ACC) was unaltered. Paradoxically, in vivo insulin-stimulated 2-deoxyglucose uptake was increased in EDL and tibialis anterior muscle from TBC1D1-deficient mice......)) and wild-type littermates were studied. Glucose and insulin tolerance, glucose utilization, hepatic glucose production, and tissue-specific insulin-mediated glucose uptake were determined. The effect of insulin, AICAR, or contraction on glucose transport was studied in isolated skeletal muscle. Glucose...

  2. A liver stress-endocrine nexus promotes metabolic integrity during dietary protein dilution

    DEFF Research Database (Denmark)

    Maida, Adriano; Zota, Annika; Sjøberg, Kim Anker

    2016-01-01

    of impaired glucose homeostasis independently of obesity and food intake. DPD-mediated metabolic inefficiency and improvement of glucose homeostasis were independent of uncoupling protein 1 (UCP1), but required expression of liver-derived fibroblast growth factor 21 (FGF21) in both lean and obese mice. FGF21...... expression and secretion as well as the associated metabolic remodeling induced by DPD also required induction of liver-integrated stress response-driven nuclear protein 1 (NUPR1). Insufficiency of select nonessential amino acids (NEAAs) was necessary and adequate for NUPR1 and subsequent FGF21 induction...... and secretion in hepatocytes in vitro and in vivo. Taken together, these data indicate that DPD promotes improved glucose homeostasis through an NEAA insufficiency-induced liver NUPR1/FGF21 axis....

  3. [New theory of holistic integrative physiology and medicine. III: New insight of neurohumoral mechanism and pattern of control and regulation for core axe of respiration, circulation and metabolism].

    Science.gov (United States)

    Sun, Xing-guo

    2015-07-01

    Systemic mechanism of neurohumoral control and regulation for human is limited. We used the new theory of holistic integrative physiology and medicine to approach the mechanism and pattern of neurohumoral control and regulation for life. As the core of human life, there are two core axes of functions. The first one is the common goal of respiration and circulation to transport oxygen and carbon dioxide for cells, and the second one is the goal of gastrointestinal tract and circulation to transport energy material and metabolic product for cells. These two core axes maintain the metabolism. The neurohumoral regulation is holistically integrated and unified for all functions in human body. We simplified explain the mechanism of neurohumoral control and regulation life (respiration and circulation) as the example pattern of sound system. Based upon integrated regulation of life, we described the neurohumoral pattern to control respiration and circulation.

  4. Muscle protein degradation and amino acid metabolism during prolonged knee-extensor exercise in humans

    DEFF Research Database (Denmark)

    Van Hall, Gerrit; Saltin, B; Wagenmakers, A J

    1999-01-01

    to a substantial increase in net muscle protein degradation, and that a lowering of the starting muscle glycogen content leads to a further increase. The carbon atoms of the branched-chain amino acids (BCAA), glutamate, aspartate and asparagine, liberated by protein degradation, and the BCAA and glutamate......The aim of this study was to investigate whether prolonged one-leg knee-extensor exercise enhances net protein degradation in muscle with a normal or low glycogen content. Net amino acid production, as a measure of net protein degradation, was estimated from leg exchange and from changes...... in the concentrations of amino acids that are not metabolized in skeletal muscle. Experiments were performed at rest and during one-leg knee-extensor exercise in six subjects having one leg with a normal glycogen content and the other with a low glycogen content. Exercise was performed for 90 min at a workload of 60...

  5. Role of the mixed-lineage protein kinase pathway in the metabolic stress response to obesity

    OpenAIRE

    Kant, Shashi; Barrett, Tamera; Vertii, Anastassiia; Noh, Yun Hee; Jung, Dae Young; Kim, Jason K.; Davis, Roger J.

    2013-01-01

    Saturated free fatty acid (FFA) is implicated in the metabolic response to obesity. In vitro studies indicate that FFA signaling may be mediated by the mixed-lineage protein kinase (MLK) pathway that activates cJun NH2-terminal kinase (JNK). Here, we examined the role of the MLK pathway in vivo using a mouse model of diet-induced obesity. The ubiquitously expressed MLK2 and MLK3 protein kinases have partially redundant functions. We therefore compared wild-type and compound mutant mice that l...

  6. Altered Mitochondria, Protein Synthesis Machinery, and Purine Metabolism Are Molecular Contributors to the Pathogenesis of Creutzfeldt-Jakob Disease.

    Science.gov (United States)

    Ansoleaga, Belén; Garcia-Esparcia, Paula; Llorens, Franc; Hernández-Ortega, Karina; Carmona Tech, Margarita; Antonio Del Rio, José; Zerr, Inga; Ferrer, Isidro

    2016-06-12

    Neuron loss, synaptic decline, and spongiform change are the hallmarks of sporadic Creutzfeldt-Jakob disease (sCJD), and may be related to deficiencies in mitochondria, energy metabolism, and protein synthesis. To investigate these relationships, we determined the expression levels of genes encoding subunits of the 5 protein complexes of the electron transport chain, proteins involved in energy metabolism, nucleolar and ribosomal proteins, and enzymes of purine metabolism in frontal cortex samples from 15 cases of sCJD MM1 and age-matched controls. We also assessed the protein expression levels of subunits of the respiratory chain, initiation and elongation translation factors of protein synthesis, and localization of selected mitochondrial components. We identified marked, generalized alterations of mRNA and protein expression of most subunits of all 5 mitochondrial respiratory chain complexes in sCJD cases. Expression of molecules involved in protein synthesis and purine metabolism were also altered in sCJD. These findings point to altered mRNA and protein expression of components of mitochondria, protein synthesis machinery, and purine metabolism as components of the pathogenesis of CJD. © 2016 American Association of Neuropathologists, Inc. All rights reserved.

  7. Metabolic and mitochondrial dysfunction in early mouse embryos following maternal dietary protein intervention.

    Science.gov (United States)

    Mitchell, Megan; Schulz, Samantha L; Armstrong, David T; Lane, Michelle

    2009-04-01

    Dietary supply of nutrients, both periconception and during pregnancy, influence the growth and development of the fetus and offspring and their health into adult life. Despite the importance of research efforts surrounding the developmental origins of health and disease hypothesis, the biological mechanisms involved remain elusive. Mitochondria are of major importance in the oocyte and early embryo, particularly as a source of ATP generation, and perturbations in their function have been related to reduced embryo quality. The present study examined embryo development following periconception exposure of females to a high-protein diet (HPD) or a low-protein diet (LPD) relative to a medium-protein diet (MPD; control), and we hypothesized that perturbed mitochondrial metabolism in the mouse embryo may be responsible for the impaired embryo and fetal development reported by others. Although the rate of development to the blastocyst stage did not differ between diets, both the HPD and LPD reduced the number of inner cell mass cells in the blastocyst-stage embryo. Furthermore, mitochondrial membrane potential was reduced and mitochondrial calcium levels increased in the 2-cell embryo. Embryos from HPD females had elevated levels of reactive oxygen species and ADP concentrations, indicative of metabolic stress and, potentially, the uncoupling of oxidative phosphorylation, whereas embryos from LPD females had reduced mitochondrial clustering around the nucleus, suggestive of an overall quietening of metabolism. Thus, although periconception dietary supply of different levels of protein is permissive of development, mitochondrial metabolism is altered in the early embryo, and the nature of the perturbation differs between HPD and LPD exposure.

  8. Importance of the cyanobacterial Gun4 protein for chlorophyll metabolism and assembly of photosynthetic complexes

    Czech Academy of Sciences Publication Activity Database

    Sobotka, Roman; Dühring, U.; Komenda, Josef; Peter, E.; Gardian, Zdenko; Tichý, Martin; Grimm, D.; Wilde, A.

    2008-01-01

    Roč. 283, č. 38 (2008), s. 25794-25802 ISSN 0021-9258 R&D Projects: GA AV ČR IAA500200713 Grant - others:DE(DE) SFB429; DE(DE) TPA8 Institutional research plan: CEZ:AV0Z50200510; CEZ:AV0Z50510513 Keywords : gun4 protein * chlorophyll metabolism * photosystem II Subject RIV: EE - Microbiology, Virology Impact factor: 5.520, year: 2008

  9. Effect of hyperbaric oxygenation on carbohydrate metabolism protein synthesis in the myocardium during sustained hypodynamia

    Science.gov (United States)

    Makarov, G. A.

    1980-01-01

    Glycolysis and the intensity of protein synthesis were studied in 140 white male rats in subcellular fractions of the myocardium during 45 day hypodynamia and hyperbaric oxygenation. Hypodynamia increased: (1) the amount of lactic acids; (2) the amount of pyruvic acid; (3) the lactate/pyruvate coefficient; and (4) the activities of aldolase and lactate dehydrogenase. Hyperbaric oxygenation was found to have a favorable metabolic effect on the animals with hypodynamia.

  10. Metabolism of serine in growing rats and chicks at various dietary protein levels

    International Nuclear Information System (INIS)

    Tanaka, Hideyuki; Yamaguchi, Michio; Kametaka, Masao

    1976-01-01

    The metabolic fate of the carbon skeleton of L-serine-U- 14 C has been investigated, in vivo and in vitro, in growing rats and chicks fed the diets with various protein calories percents (C %) at 410 kcal of metabolizable energy. The incorporation of 14 C into body protein at 12 hr after the injection of serine- 14 C was about 49% of the injected dose in rats fed the 10 or 15 PC% diet, though the value was reduced in rats fed lower and higher protein diets. The 14 CO 2 production was smaller in rats fed the 10 and 15 PC% diet, and it showed an inverse pattern to that of the 14 C incorporation into body protein. Urinary excretion of 14 C was higher in rats fed 10 and higher PC% diets, whose growth rate and net body protein retention were maximum. In contrast to the case of rats, the incorporation of 14 C into body protein of chicks at 6 hr after the injection was rather reduced in the 15 PC% group. The proportion of 14 C excreted as uric acid was remarkably increased above the 10 PC% group, and about 19% of the injected dose was recovered in the 50 PC% group. The catabolic rate of serine in the liver slices of rats and chicks was increased by high protein diets. These results support the concept that the nutritional significance of metabolism of the carbon skeleton of serine in growing rats and chicks is different from each other, especially at high protein diets. (auth.)

  11. Effects of Dietary Protein Source and Quantity during Weight Loss on Appetite, Energy Expenditure, and Cardio-Metabolic Responses

    Directory of Open Access Journals (Sweden)

    Jia Li

    2016-01-01

    Full Text Available Higher protein meals increase satiety and the thermic effect of feeding (TEF in acute settings, but it is unclear whether these effects remain after a person becomes acclimated to energy restriction or a given protein intake. This study assessed the effects of predominant protein source (omnivorous, beef/pork vs. lacto-ovo vegetarian, soy/legume and quantity (10%, 20%, or 30% of energy from protein on appetite, energy expenditure, and cardio-metabolic indices during energy restriction (ER in overweight and obese adults. Subjects were randomly assigned to one protein source and then consumed diets with different quantities of protein (4 weeks each in a randomized crossover manner. Perceived appetite ratings (free-living and in-lab, TEF, and fasting cardio-metabolic indices were assessed at the end of each 4-week period. Protein source and quantity did not affect TEF, hunger, or desire to eat, other than a modestly higher daily composite fullness rating with 30% vs. 10% protein diet (p = 0.03. While the 20% and 30% protein diets reduced cholesterol, triacylglycerol, and APO-B vs. 10% protein (p < 0.05, protein source did not affect cardio-metabolic indices. In conclusion, diets varying in protein quantity with either beef/pork or soy/legume as the predominant source have minimal effects on appetite control, energy expenditure and cardio-metabolic risk factors during ER-induced weight loss.

  12. The LDL Receptor-Related Protein 1: At the Crossroads of Lipoprotein Metabolism and Insulin Signaling

    Directory of Open Access Journals (Sweden)

    Dianaly T. Au

    2017-01-01

    Full Text Available The metabolic syndrome is an escalating worldwide public health concern. Defined by a combination of physiological, metabolic, and biochemical factors, the metabolic syndrome is used as a clinical guideline to identify individuals with a higher risk for type 2 diabetes and cardiovascular disease. Although risk factors for type 2 diabetes and cardiovascular disease have been known for decades, the molecular mechanisms involved in the pathophysiology of these diseases and their interrelationship remain unclear. The LDL receptor-related protein 1 (LRP1 is a large endocytic and signaling receptor that is widely expressed in several tissues. As a member of the LDL receptor family, LRP1 is involved in the clearance of chylomicron remnants from the circulation and has been demonstrated to be atheroprotective. Recently, studies have shown that LRP1 is involved in insulin receptor trafficking and regulation and glucose metabolism. This review summarizes the role of tissue-specific LRP1 in insulin signaling and its potential role as a link between lipoprotein and glucose metabolism in diabetes.

  13. The Roles of Vitamin A in the Regulation of Carbohydrate, Lipid, and Protein Metabolism

    Directory of Open Access Journals (Sweden)

    Wei Chen

    2014-05-01

    Full Text Available Currently, two-thirds of American adults are overweight or obese. This high prevalence of overweight/obesity negatively affects the health of the population, as obese individuals tend to develop several chronic diseases, such as type 2 diabetes and cardiovascular diseases. Due to obesity’s impact on health, medical costs, and longevity, the rise in the number of obese people has become a public health concern. Both genetic and environmental/dietary factors play a role in the development of metabolic diseases. Intuitively, it seems to be obvious to link over-nutrition to the development of obesity and other metabolic diseases. However, the underlying mechanisms are still unclear. Dietary nutrients not only provide energy derived from macronutrients, but also factors such as micronutrients with regulatory roles. How micronutrients, such as vitamin A (VA; retinol, regulate macronutrient homeostasis is still an ongoing research topic. As an essential micronutrient, VA plays a key role in the general health of an individual. This review summarizes recent research progress regarding VA’s role in carbohydrate, lipid, and protein metabolism. Due to the large amount of information regarding VA functions, this review focusses on metabolism in metabolic active organs and tissues. Additionally, some perspectives for future studies will be provided.

  14. Relationship of C-reactive protein, metabolic syndrome and diabetes mellitus: potential role of statins.

    Science.gov (United States)

    Nash, David T

    2005-12-01

    Atherosclerosis and the metabolic derangements of insulin resistance, metabolic syndrome and diabetes mellitus are all associated with underlying inflammatory processes. C-reactive protein (CRP), a marker of inflammation, has been shown to be a strong independent predictor of vascular events. It adds to cardiovascular disease risk at all levels of low-density-lipoprotein cholesterol and Framingham risk scores, and elevated levels are also associated with increasing severity of the metabolic syndrome. The development of a simple, stable, noninvasive test to measure high-sensitivity CRP has provided a clinical tool that may have an important role in the identification and assessment of individuals likely to develop cardiovascular or metabolic disease. The role of CRP in predicting cardiovascular risk is less clear in African Americans, however, than in white populations. Statins and thiazolidinediones are being investigated for their potential role in the prevention and treatment of the inflammatory processes involved in the metabolic syndrome and cardiovascular disease. In the future, assessment of CRP levels may contribute importantly to clinical decision-making in reducing cardiovascular risk.

  15. The Role of Carbohydrate Response Element Binding Protein in Intestinal and Hepatic Fructose Metabolism

    Directory of Open Access Journals (Sweden)

    Katsumi Iizuka

    2017-02-01

    Full Text Available Many articles have discussed the relationship between fructose consumption and the incidence of obesity and related diseases. Fructose is absorbed in the intestine and metabolized in the liver to glucose, lactate, glycogen, and, to a lesser extent, lipids. Unabsorbed fructose causes bacterial fermentation, resulting in irritable bowl syndrome. Therefore, understanding the mechanisms underlying intestinal and hepatic fructose metabolism is important for the treatment of metabolic syndrome and fructose malabsorption. Carbohydrate response element binding protein (ChREBP is a glucose-activated transcription factor that controls approximately 50% of de novo lipogenesis in the liver. ChREBP target genes are involved in glycolysis (Glut2, liver pyruvate kinase, fructolysis (Glut5, ketohexokinase, and lipogenesis (acetyl CoA carboxylase, fatty acid synthase. ChREBP gene deletion protects against high sucrose diet-induced and leptin-deficient obesity, because Chrebp−/− mice cannot consume fructose or sucrose. Moreover, ChREBP contributes to some of the physiological effects of fructose on sweet taste preference and glucose production through regulation of ChREBP target genes, such as fibroblast growth factor-21 and glucose-6-phosphatase catalytic subunits. Thus, ChREBP might play roles in fructose metabolism. Restriction of excess fructose intake will be beneficial for preventing not only metabolic syndrome but also irritable bowl syndrome.

  16. DISTRIBUTION OF GBM HEPARAN-SULFATE PROTEOGLYCAN CORE PROTEIN AND SIDE-CHAINS IN HUMAN GLOMERULAR-DISEASES

    NARCIS (Netherlands)

    VANDENBORN, J; VANDENHEUVEL, LPWJ; BAKKER, MAH; VEERKAMP, JH; ASSMANN, KJM; WEENING, JJ; BERDEN, JHM

    Using monoclonal antibodies (mAbs) recognizing either the core protein or the heparan sulfate (HS) side chain of human GBM heparan sulfate proteoglycan (HSPG), we investigated their glomerular distribution on cryostat sections of human kidney tissues. The study involved 95 biopsies comprising twelve

  17. Aberrant expression of mucin core proteins and o-linked glycans associated with progression of pancreatic cancer

    DEFF Research Database (Denmark)

    Remmers, Neeley; Anderson, Judy M; Linde, Erin M

    2013-01-01

    Mucin expression is a common feature of most adenocarcinomas and features prominently in current attempts to improve diagnosis and therapy for pancreatic cancer and other adenocarcinomas. We investigated the expression of a number of mucin core proteins and associated O-linked glycans expressed i...

  18. Bioinformatic analysis suggests that the Cypovirus 1 major core protein cistron harbours an overlapping gene

    Directory of Open Access Journals (Sweden)

    Atkins John F

    2008-05-01

    Full Text Available Abstract Members of the genus Cypovirus (family Reoviridae are common pathogens of insects. These viruses have linear dsRNA genomes divided into 10–11 segments, which have generally been assumed to be monocistronic. Here, bioinformatic evidence is presented for a short overlapping coding sequence (CDS in the cypovirus genome segment encoding the major core capsid protein VP1, overlapping the 5'-terminal region of the VP1 ORF in the +1 reading frame. In Cypovirus type 1 (CPV-1, a 62-codon AUG-initiated open reading frame (hereafter ORFX is present in all four available segment 1 sequences. The pattern of base variations across the sequence alignment indicates that ORFX is subject to functional constraints at the amino acid level (even when the constraints due to coding in the overlapping VP1 reading frame are taken into account; MLOGD software. In fact the translated ORFX shows greater amino acid conservation than the overlapping region of VP1. The genomic location of ORFX is consistent with translation via leaky scanning. A 62–64 codon AUG-initiated ORF is present in a corresponding location and reading frame in other available cypovirus sequences (2 CPV-14, 1 CPV-15 and an 87-codon ORFX homologue may also be present in Aedes pseudoscutellaris reovirus. The ORFX amino acid sequences are hydrophilic and basic, with between 12 and 16 Arg/Lys residues in each though, at 7.5–10.2 kDa, the putative ORFX product is too small to appear on typical published protein gels.

  19. Metabolism

    Science.gov (United States)

    ... lin), which signals cells to increase their anabolic activities. Metabolism is a complicated chemical process, so it's not ... how those enzymes or hormones work. When the metabolism of body chemicals is ... Hyperthyroidism (pronounced: hi-per-THIGH-roy-dih-zum). Hyperthyroidism ...

  20. Identification of protein W, the elusive sixth subunit of the Rhodopseudomonas palustris reaction center-light harvesting 1 core complex.

    Science.gov (United States)

    Jackson, Philip J; Hitchcock, Andrew; Swainsbury, David J K; Qian, Pu; Martin, Elizabeth C; Farmer, David A; Dickman, Mark J; Canniffe, Daniel P; Hunter, C Neil

    2018-02-01

    The X-ray crystal structure of the Rhodopseudomonas (Rps.) palustris reaction center-light harvesting 1 (RC-LH1) core complex revealed the presence of a sixth protein component, variably referred to in the literature as helix W, subunit W or protein W. The position of this protein prevents closure of the LH1 ring, possibly to allow diffusion of ubiquinone/ubiquinol between the RC and the cytochrome bc 1 complex in analogous fashion to the well-studied PufX protein from Rhodobacter sphaeroides. The identity and function of helix W have remained unknown for over 13years; here we use a combination of biochemistry, mass spectrometry, molecular genetics and electron microscopy to identify this protein as RPA4402 in Rps. palustris CGA009. Protein W shares key conserved sequence features with PufX homologs, and although a deletion mutant was able to grow under photosynthetic conditions with no discernible phenotype, we show that a tagged version of protein W pulls down the RC-LH1 complex. Protein W is not encoded in the photosynthesis gene cluster and our data indicate that only approximately 10% of wild-type Rps. palustris core complexes contain this non-essential subunit; functional and evolutionary consequences of this observation are discussed. The ability to purify uniform RC-LH1 and RC-LH1-protein W preparations will also be beneficial for future structural studies of these bacterial core complexes. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  1. Human Adenovirus Infection Causes Cellular E3 Ubiquitin Ligase MKRN1 Degradation Involving the Viral Core Protein pVII.

    Science.gov (United States)

    Inturi, Raviteja; Mun, Kwangchol; Singethan, Katrin; Schreiner, Sabrina; Punga, Tanel

    2018-02-01

    Human adenoviruses (HAdVs) are common human pathogens encoding a highly abundant histone-like core protein, VII, which is involved in nuclear delivery and protection of viral DNA as well as in sequestering immune danger signals in infected cells. The molecular details of how protein VII acts as a multifunctional protein have remained to a large extent enigmatic. Here we report the identification of several cellular proteins interacting with the precursor pVII protein. We show that the cellular E3 ubiquitin ligase MKRN1 is a novel precursor pVII-interacting protein in HAdV-C5-infected cells. Surprisingly, the endogenous MKRN1 protein underwent proteasomal degradation during the late phase of HAdV-C5 infection in various human cell lines. MKRN1 protein degradation occurred independently of the HAdV E1B55K and E4orf6 proteins. We provide experimental evidence that the precursor pVII protein binding enhances MKRN1 self-ubiquitination, whereas the processed mature VII protein is deficient in this function. Based on these data, we propose that the pVII protein binding promotes MKRN1 self-ubiquitination, followed by proteasomal degradation of the MKRN1 protein, in HAdV-C5-infected cells. In addition, we show that measles virus and vesicular stomatitis virus infections reduce the MKRN1 protein accumulation in the recipient cells. Taken together, our results expand the functional repertoire of the HAdV-C5 precursor pVII protein in lytic virus infection and highlight MKRN1 as a potential common target during different virus infections. IMPORTANCE Human adenoviruses (HAdVs) are common pathogens causing a wide range of diseases. To achieve pathogenicity, HAdVs have to counteract a variety of host cell antiviral defense systems, which would otherwise hamper virus replication. In this study, we show that the HAdV-C5 histone-like core protein pVII binds to and promotes self-ubiquitination of a cellular E3 ubiquitin ligase named MKRN1. This mutual interaction between the pVII and

  2. Metabolism of histones and nonhistone proteins of the nuclei and chromatin of liver cells in rats of different ages

    International Nuclear Information System (INIS)

    Klimenko, A.I.; Malyshev, A.B.; Kulachenko, B.V.

    1986-01-01

    The metabolism of various classes of histones and nonhistone proteins in whole nuclei and liver chromatin of albino Wistar rats 1, 3, 12, and 24 months of age was studied. It was shown that in the course of postnatal ontogenesis, the metabolism of nonhistone proteins, extractable by a 0.14 M solution of NaCl, is increased in the animals. The incorporation of labeled precursors into the HMG 14 and HMG 17 proteins decreases with age of the animals; a higher level of specific radioactivity was established for the HMG 1+2 proteins in the 3- and 24-month old animals. The intensity of the metabolism of nonhistone proteins and histones is higher in the chromatin complex than in the whole nucleus at all stages of postnatal development of the animals. Among the histone proteins, H1 histones possess a higher level of specific radioactivity in animals of all age groups

  3. Effect of protein provision via milk replacer or solid feed on protein metabolism in veal calves

    NARCIS (Netherlands)

    Berends, H.; Borne, van den J.J.G.C.; Røjen, B.A.; Hendriks, W.H.; Gerrits, W.J.J.

    2015-01-01

    The current study evaluated the effects of protein provision to calves fed a combination of solid feed (SF) and milk replacer (MR) at equal total N intake on urea recycling and N retention. Nitrogen balance traits and [15N2]urea kinetics were measured in 30 calves (23 wk of age, 180 ± 3.7 kg of body

  4. The Effect of Oral Leucine on Protein Metabolism in Adolescents with Type 1 Diabetes Mellitus

    Directory of Open Access Journals (Sweden)

    Wilson ThomasA

    2010-11-01

    Full Text Available Lack of insulin results in a catabolic state in subjects with insulin-dependent diabetes mellitus which is reversed by insulin treatment. Amino acid supply, especially branched chain amino acids such as leucine, enhances protein synthesis in both animal and human studies. This small study was undertaken to assess the acute effect of supplemental leucine on protein metabolism in adolescents with type 1 diabetes. L-[1-13C] Leucine was used to assess whole-body protein metabolism in six adolescent females (16–18 yrs with type 1 diabetes during consumption of a basal diet (containing 58 μmoles leucine/kg/h and the basal diet with supplemental leucine (232 μmoles leucine/kg/h. Net leucine balance was significantly higher with supplemental leucine ( μmoles leucine/kg body weight/hr than with the basal diet (, due to reduced protein degradation ( μmoles leucine/kg body weight/hr compared to the basal diet (, .

  5. Dietary live yeast alters metabolic profiles, protein biosynthesis and thermal stress tolerance of Drosophila melanogaster.

    Science.gov (United States)

    Colinet, Hervé; Renault, David

    2014-04-01

    The impact of nutritional factors on insect's life-history traits such as reproduction and lifespan has been excessively examined; however, nutritional determinant of insect's thermal tolerance has not received a lot of attention. Dietary live yeast represents a prominent source of proteins and amino acids for laboratory-reared drosophilids. In this study, Drosophila melanogaster adults were fed on diets supplemented or not with live yeast. We hypothesized that manipulating nutritional conditions through live yeast supplementation would translate into altered physiology and stress tolerance. We verified how live yeast supplementation affected body mass characteristics, total lipids and proteins, metabolic profiles and cold tolerance (acute and chronic stress). Females fed with live yeast had increased body mass and contained more lipids and proteins. Using GC/MS profiling, we found distinct metabolic fingerprints according to nutritional conditions. Metabolite pathway enrichment analysis corroborated that live yeast supplementation was associated with amino acid and protein biosyntheses. The cold assays revealed that the presence of dietary live yeast greatly promoted cold tolerance. Hence, this study conclusively demonstrates a significant interaction between nutritional conditions and thermal tolerance. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Does altered protein metabolism interfere with postmortem degradation analysis for PMI estimation?

    Science.gov (United States)

    Zissler, A; Ehrenfellner, B; Foditsch, E E; Monticelli, F C; Pittner, S

    2018-03-02

    An accurate estimation of the postmortem interval (PMI) is a central aspect in forensic routine. Recently, a novel approach based on the analysis of postmortem muscle protein degradation has been proposed. However, a number of questions remain to be answered until sensible application of this method to a broad variety of forensic cases is possible. To evaluate whether altered in vivo protein metabolism interferes with postmortem degradation patterns, we conducted a comparative study. We developed a standardized animal degradation model in rats, and collected additional muscle samples from animals recovering from muscle injury and from rats with developed disuse muscle atrophy after induced spinal cord injury. All samples were analyzed by SDS-PAGE and Western blot, labeling well-characterized muscle proteins. Tropomyosin was found to be stable throughout the investigated PMI and no alterations were detected in regenerating and atrophic muscles. In contrast, significant predictable postmortem changes occurred in desmin and vinculin protein band patterns. While no significant deviations from native patterns were detected in at-death samples of disuse muscle atrophy, interestingly, samples of rats recovering from muscle injury revealed additional desmin and vinculin degradation bands that did not occur in this form in any of the examined postmortem samples regardless of PMI. It remains to be investigated whether in vivo-altered metabolism influences postmortem degradation kinetics or if such muscle samples undergo postmortem degradation in a regular fashion.

  7. The Effect of Protein Restriction in the In Vitro Metabolism of Albendazole in Rats.

    Science.gov (United States)

    Belaz, Kátia Roberta A; de O Cardoso, Josiane; da Silva, Carlos Alberto; Oliveira, Regina V

    2015-01-01

    This work presents an in vitro investigation of the effect of protein restriction on the metabolism of albendazole (ABZ). This study was conducted using liver microsomal fractions obtained from Wistar rats. For the quantitative analysis, a multidimensional High Performance Liquid Chromatography (2D HPLC) method was fully validated for the determination of the ABZ metabolites: albendazole sulfoxide, albendazole sulfone and albendazole 2-aminesulfone. The target compounds were directly extracted using a C8-RAM-BSA column (5.0x0.46 cm i.d.) and analyzed on a chromatographic chiral column containing amylose tris(3,5-dimethylphenylcarbamate) (150x4.6 mm i.d.). The in vitro biotransformation results showed that the protein restriction influenced the oxidative metabolism of ABZ. The production of R-(+)-ABZ-SO (1309 nmol/L) and S-(-)-ABZ-SO (1456 nmol/L) was higher in the control animals than in the animals fed with a diet containing 6% protein, which produced 778.7 nmol/L and 709.5 nmol/L for R-(+) and S-(-)-ABZ-SO enantiomers, respectively. These results were statistically inspected by Student´s t test and the results showed a significant difference between the two means (p0.05). Furthermore, animal nutritional condition could affect the pattern of ABZ sulphoxidation indicating that the protein nutrition affect primarily the formation of R-(+)-ABZSO and S-(-)-ABZ-SO enantiomers.

  8. Whey Protein Supplementation Enhances Whole Body Protein Metabolism and Performance Recovery after Resistance Exercise: A Double-Blind Crossover Study

    Directory of Open Access Journals (Sweden)

    Daniel W. D. West

    2017-07-01

    Full Text Available No study has concurrently measured changes in free-living whole body protein metabolism and exercise performance during recovery from an acute bout of resistance exercise. We aimed to determine if whey protein ingestion enhances whole body net protein balance and recovery of exercise performance during overnight (10 h and 24 h recovery after whole body resistance exercise in trained men. In a double-blind crossover design, 12 trained men (76 ± 8 kg, 24 ± 4 years old, 14% ± 5% body fat; means ± standard deviation (SD performed resistance exercise in the evening prior to consuming either 25 g of whey protein (PRO; MuscleTech 100% Whey or an energy-matched placebo (CHO immediately post-exercise (0 h, and again the following morning (~10 h of recovery. A third randomized trial, completed by the same participants, involving no exercise and no supplement served as a rested control trial (Rest. Participants ingested [15N]glycine to determine whole body protein kinetics and net protein balance over 10 and 24 h of recovery. Performance was assessed pre-exercise and at 0, 10, and 24 h of recovery using a battery of tests. Net protein balance tended to improve in PRO (P = 0.064; effect size (ES = 0.61, PRO vs. CHO during overnight recovery. Over 24 h, net balance was enhanced in PRO (P = 0.036 but not in CHO (P = 0.84; ES = 0.69, PRO vs. CHO, which was mediated primarily by a reduction in protein breakdown (PRO < CHO; P < 0.01. Exercise decreased repetitions to failure (REP, maximal strength (MVC, peak and mean power, and countermovement jump performance (CMJ at 0 h (all P < 0.05 vs. Pre. At 10 h, there were small-to-moderate effects for enhanced recovery of the MVC (ES = 0.56, mean power (ES = 0.49, and CMJ variables (ES: 0.27–0.49 in PRO. At 24 h, protein supplementation improved MVC (ES = 0.76, REP (ES = 0.44, and peak power (ES = 0.55. In conclusion, whey protein supplementation enhances whole body anabolism, and may improve acute recovery of

  9. High-protein-low-carbohydrate diet: deleterious metabolic and cardiovascular effects depend on age.

    Science.gov (United States)

    Bedarida, Tatiana; Baron, Stephanie; Vessieres, Emilie; Vibert, Francoise; Ayer, Audrey; Marchiol-Fournigault, Carmen; Henrion, Daniel; Paul, Jean-Louis; Noble, Florence; Golmard, Jean-Louis; Beaudeux, Jean-Louis; Cottart, Charles-Henry; Nivet-Antoine, Valerie

    2014-09-01

    High-protein-low-carbohydrate (HP-LC) diets have become widespread. Yet their deleterious consequences, especially on glucose metabolism and arteries, have already been underlined. Our previous study (2) has already shown glucose intolerance with major arterial dysfunction in very old mice subjected to an HP-LC diet. The hypothesis of this work was that this diet had an age-dependent deleterious metabolic and cardiovascular outcome. Two groups of mice, young and adult (3 and 6 mo old), were subjected for 12 wk to a standard or to an HP-LC diet. Glucose and lipid metabolism was studied. The cardiovascular system was explored from the functional stage with Doppler-echography to the molecular stage (arterial reactivity, mRNA, immunohistochemistry). Young mice did not exhibit any significant metabolic modification, whereas adult mice presented marked glucose intolerance associated with an increase in resistin and triglyceride levels. These metabolic disturbances were responsible for cardiovascular damages only in adult mice, with decreased aortic distensibility and left ventricle dysfunction. These seemed to be the consequence of arterial dysfunctions. Mesenteric arteries were the worst affected with a major oxidative stress, whereas aorta function seemed to be maintained with an appreciable role of cyclooxygenase-2 to preserve endothelial function. This study highlights for the first time the age-dependent deleterious effects of an HP-LC diet on metabolism, with glucose intolerance and lipid disorders and vascular (especially microvessels) and cardiac functions. This work shows that HP-LC lead to equivalent cardiovascular alterations, as observed in very old age, and underlines the danger of such diet. Copyright © 2014 the American Physiological Society.

  10. Nonstructural protein 5A is incorporated into hepatitis C virus low-density particle through interaction with core protein and microtubules during intracellular transport.

    Directory of Open Access Journals (Sweden)

    Chao-Kuen Lai

    Full Text Available Nonstructural protein 5A (NS5A of hepatitis C virus (HCV serves dual functions in viral RNA replication and virus assembly. Here, we demonstrate that HCV replication complex along with NS5A and Core protein was transported to the lipid droplet (LD through microtubules, and NS5A-Core complexes were then transported from LD through early-to-late endosomes to the plasma membrane via microtubules. Further studies by cofractionation analysis and immunoelectron microscopy of the released particles showed that NS5A-Core complexes, but not NS4B, were present in the low-density fractions, but not in the high-density fractions, of the HCV RNA-containing virions and associated with the internal virion core. Furthermore, exosomal markers CD63 and CD81 were also detected in the low-density fractions, but not in the high-density fractions. Overall, our results suggest that HCV NS5A is associated with the core of the low-density virus particles which exit the cell through a preexisting endosome/exosome pathway and may contribute to HCV natural infection.

  11. Liver carbohydrates metabolism: A new islet-neogenesis associated protein peptide (INGAP-PP) target.

    Science.gov (United States)

    Villagarcía, Hernán Gonzalo; Román, Carolina Lisi; Castro, María Cecilia; González, Luisa Arbeláez; Ronco, María Teresa; Francés, Daniel Eleazar; Massa, María Laura; Maiztegui, Bárbara; Flores, Luis Emilio; Gagliardino, Juan José; Francini, Flavio

    2018-03-01

    Islet-Neogenesis Associated Protein-Pentadecapeptide (INGAP-PP) increases β-cell mass and enhances glucose and amino acids-induced insulin secretion. Our aim was to demonstrate its effect on liver metabolism. For that purpose, adult male Wistar rats were injected twice-daily (10 days) with saline solution or INGAP-PP (250 μg). Thereafter, serum glucose, triglyceride and insulin levels were measured and homeostasis model assessment (HOMA-IR) and hepatic insulin sensitivity (HIS) were determined. Liver glucokinase and glucose-6-phosphatase (G-6-Pase) expression and activity, phosphoenolpyruvate carboxykinase (PEPCK) expression, phosphofructokinase-2 (PFK-2) protein content, P-Akt/Akt and glycogen synthase kinase-3β (P-GSK3/GSK3) protein ratios and glycogen deposit were also determined. Additionally, glucokinase activity and G-6-Pase and PEPCK gene expression were also determined in isolated hepatocytes from normal rats incubated with INGAP-PP (5 μg/ml). INGAP-PP administration did not modify any of the serum parameters tested but significantly increased activity of liver glucokinase and the protein level of its cytosolic activator, PFK-2. Conversely, INGAP-PP treated rats decreased gene expression and enzyme activity of gluconeogenic enzymes, G-6-Pase and PEPCK. They also showed a higher glycogen deposit and P-GSK3/GSK3 and P-Akt/Akt ratio. In isolated hepatocytes, INGAP-PP increased GK activity and decreased G-6-Pase and PEPCK expression. These results demonstrate a direct effect of INGAP-PP on the liver acting through P-Akt signaling pathway. INGAP-PP enhances liver glucose metabolism and deposit and reduces its production/output, thereby contributing to maintain normal glucose homeostasis. These results reinforce the concept that INGAP-PP might become a useful tool to treat people with impaired islet/liver glucose metabolism as it occurs in T2D. Copyright © 2018 Elsevier Inc. All rights reserved.

  12. Historical and contemporary stable isotope tracer approaches to studying mammalian protein metabolism

    Science.gov (United States)

    2016-01-01

    Over a century ago, Frederick Soddy provided the first evidence for the existence of isotopes; elements that occupy the same position in the periodic table are essentially chemically identical but differ in mass due to a different number of neutrons within the atomic nucleus. Allied to the discovery of isotopes was the development of some of the first forms of mass spectrometers, driven forward by the Nobel laureates JJ Thomson and FW Aston, enabling the accurate separation, identification, and quantification of the relative abundance of these isotopes. As a result, within a few years, the number of known isotopes both stable and radioactive had greatly increased and there are now over 300 stable or radioisotopes presently known. Unknown at the time, however, was the potential utility of these isotopes within biological disciplines, it was soon discovered that these stable isotopes, particularly those of carbon (13C), nitrogen (15N), oxygen (18O), and hydrogen (2H) could be chemically introduced into organic compounds, such as fatty acids, amino acids, and sugars, and used to “trace” the metabolic fate of these compounds within biological systems. From this important breakthrough, the age of the isotope tracer was born. Over the following 80 yrs, stable isotopes would become a vital tool in not only the biological sciences, but also areas as diverse as forensics, geology, and art. This progress has been almost exclusively driven through the development of new and innovative mass spectrometry equipment from IRMS to GC‐MS to LC‐MS, which has allowed for the accurate quantitation of isotopic abundance within samples of complex matrices. This historical review details the development of stable isotope tracers as metabolic tools, with particular reference to their use in monitoring protein metabolism, highlighting the unique array of tools that are now available for the investigation of protein metabolism in vivo at a whole body down to a single protein level

  13. Rearrangement of a polar core provides a conserved mechanism for constitutive activation of class B G protein-coupled receptors

    Science.gov (United States)

    Yin, Yanting; de Waal, Parker W.; He, Yuanzheng; Zhao, Li-Hua; Yang, Dehua; Cai, Xiaoqing; Jiang, Yi; Melcher, Karsten; Wang, Ming-Wei; Xu, H. Eric

    2017-01-01

    The glucagon receptor (GCGR) belongs to the secretin-like (class B) family of G protein-coupled receptors (GPCRs) and is activated by the peptide hormone glucagon. The structures of an activated class B GPCR have remained unsolved, preventing a mechanistic understanding of how these receptors are activated. Using a combination of structural modeling and mutagenesis studies, we present here two modes of ligand-independent activation of GCGR. First, we identified a GCGR-specific hydrophobic lock comprising Met-338 and Phe-345 within the IC3 loop and transmembrane helix 6 (TM6) and found that this lock stabilizes the TM6 helix in the inactive conformation. Disruption of this hydrophobic lock led to constitutive G protein and arrestin signaling. Second, we discovered a polar core comprising conserved residues in TM2, TM3, TM6, and TM7, and mutations that disrupt this polar core led to constitutive GCGR activity. On the basis of these results, we propose a mechanistic model of GCGR activation in which TM6 is held in an inactive conformation by the conserved polar core and the hydrophobic lock. Mutations that disrupt these inhibitory elements allow TM6 to swing outward to adopt an active TM6 conformation similar to that of the canonical β2-adrenergic receptor complexed with G protein and to that of rhodopsin complexed with arrestin. Importantly, mutations in the corresponding polar core of several other members of class B GPCRs, including PTH1R, PAC1R, VIP1R, and CRFR1, also induce constitutive G protein signaling, suggesting that the rearrangement of the polar core is a conserved mechanism for class B GPCR activation. PMID:28356352

  14. Rearrangement of a polar core provides a conserved mechanism for constitutive activation of class B G protein-coupled receptors.

    Science.gov (United States)

    Yin, Yanting; de Waal, Parker W; He, Yuanzheng; Zhao, Li-Hua; Yang, Dehua; Cai, Xiaoqing; Jiang, Yi; Melcher, Karsten; Wang, Ming-Wei; Xu, H Eric

    2017-06-16

    The glucagon receptor (GCGR) belongs to the secretin-like (class B) family of G protein-coupled receptors (GPCRs) and is activated by the peptide hormone glucagon. The structures of an activated class B GPCR have remained unsolved, preventing a mechanistic understanding of how these receptors are activated. Using a combination of structural modeling and mutagenesis studies, we present here two modes of ligand-independent activation of GCGR. First, we identified a GCGR-specific hydrophobic lock comprising Met-338 and Phe-345 within the IC3 loop and transmembrane helix 6 (TM6) and found that this lock stabilizes the TM6 helix in the inactive conformation. Disruption of this hydrophobic lock led to constitutive G protein and arrestin signaling. Second, we discovered a polar core comprising conserved residues in TM2, TM3, TM6, and TM7, and mutations that disrupt this polar core led to constitutive GCGR activity. On the basis of these results, we propose a mechanistic model of GCGR activation in which TM6 is held in an inactive conformation by the conserved polar core and the hydrophobic lock. Mutations that disrupt these inhibitory elements allow TM6 to swing outward to adopt an active TM6 conformation similar to that of the canonical β 2 -adrenergic receptor complexed with G protein and to that of rhodopsin complexed with arrestin. Importantly, mutations in the corresponding polar core of several other members of class B GPCRs, including PTH1R, PAC1R, VIP1R, and CRFR1, also induce constitutive G protein signaling, suggesting that the rearrangement of the polar core is a conserved mechanism for class B GPCR activation. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. The Ablation of Mitochondrial Protein Phosphatase Pgam5 Confers Resistance Against Metabolic Stress.

    Science.gov (United States)

    Sekine, Shiori; Yao, Akari; Hattori, Kazuki; Sugawara, Sho; Naguro, Isao; Koike, Masato; Uchiyama, Yasuo; Takeda, Kohsuke; Ichijo, Hidenori

    2016-03-01

    Phosphoglycerate mutase family member 5 (PGAM5) is a mitochondrial protein phosphatase that has been reported to be involved in various stress responses from mitochondrial quality control to cell death. However, its roles in vivo are largely unknown. Here, we show that Pgam5-deficient mice are resistant to several metabolic insults. Under cold stress combined with fasting, Pgam5-deficient mice better maintained body temperature than wild-type mice and showed an extended survival rate. Serum triglycerides and lipid content in brown adipose tissue (BAT), a center of adaptive thermogenesis, were severely reduced in Pgam5-deficient mice. Moreover, although Pgam5 deficiency failed to maintain proper mitochondrial integrity in BAT, it reciprocally resulted in the dramatic induction of fibroblast growth factor 21 (FGF21) that activates various functions of BAT including thermogenesis. Thus, the enhancement of lipid metabolism and FGF21 may contribute to the cold resistance of Pgam5-deficient mice under fasting condition. Finally, we also found that Pgam5-deficient mice are resistant to high-fat-diet-induced obesity. Our study uncovered that PGAM5 is involved in the whole-body metabolism in response to stresses that impose metabolic challenges on mitochondria.

  16. The Ablation of Mitochondrial Protein Phosphatase Pgam5 Confers Resistance Against Metabolic Stress

    Directory of Open Access Journals (Sweden)

    Shiori Sekine

    2016-03-01

    Full Text Available Phosphoglycerate mutase family member 5 (PGAM5 is a mitochondrial protein phosphatase that has been reported to be involved in various stress responses from mitochondrial quality control to cell death. However, its roles in vivo are largely unknown. Here, we show that Pgam5-deficient mice are resistant to several metabolic insults. Under cold stress combined with fasting, Pgam5-deficient mice better maintained body temperature than wild-type mice and showed an extended survival rate. Serum triglycerides and lipid content in brown adipose tissue (BAT, a center of adaptive thermogenesis, were severely reduced in Pgam5-deficient mice. Moreover, although Pgam5 deficiency failed to maintain proper mitochondrial integrity in BAT, it reciprocally resulted in the dramatic induction of fibroblast growth factor 21 (FGF21 that activates various functions of BAT including thermogenesis. Thus, the enhancement of lipid metabolism and FGF21 may contribute to the cold resistance of Pgam5-deficient mice under fasting condition. Finally, we also found that Pgam5-deficient mice are resistant to high-fat-diet-induced obesity. Our study uncovered that PGAM5 is involved in the whole-body metabolism in response to stresses that impose metabolic challenges on mitochondria.

  17. Myostatin promotes distinct responses on protein metabolism of skeletal and cardiac muscle fibers of rodents

    Directory of Open Access Journals (Sweden)

    L.H. Manfredi

    2017-10-01

    Full Text Available Myostatin is a novel negative regulator of skeletal muscle mass. Myostatin expression is also found in heart in a much less extent, but it can be upregulated in pathological conditions, such as heart failure. Myostatin may be involved in inhibiting protein synthesis and/or increasing protein degradation in skeletal and cardiac muscles. Herein, we used cell cultures and isolated muscles from rats to determine protein degradation and synthesis. Muscles incubated with myostatin exhibited an increase in proteolysis with an increase of Atrogin-1, MuRF1 and LC3 genes. Extensor digitorum longus muscles and C2C12 myotubes exhibited a reduction in protein turnover. Cardiomyocytes showed an increase in proteolysis by activating autophagy and the ubiquitin proteasome system, and a decrease in protein synthesis by decreasing P70S6K. The effect of myostatin on protein metabolism is related to fiber type composition, which may be associated to the extent of atrophy mediated effect of myostatin on muscle.

  18. Effects of a high protein diet on cognition and brain metabolism in cirrhotic rats.

    Science.gov (United States)

    Méndez-López, M; Méndez, M; Arias, J; Arias, J L

    2015-10-01

    Hepatic encephalopathy (HE) is a neurological complication observed in patients with liver disease. Patients who suffer from HE present neuropsychiatric, neuromuscular and behavioral symptoms. Animal models proposed to study HE resulting from cirrhosis mimic the clinical characteristics of cirrhosis and portal hypertension, and require the administration of hepatotoxins such as thioacetamide (TAA). The aim of this study was to assess the effects of a high protein diet on motor function, anxiety and memory processes in a model of cirrhosis induced by TAA administration. In addition, we used cytochrome c-oxidase (COx) histochemistry to assess the metabolic activity of the limbic system regions. Male rats were distributed into groups: control, animals with cirrhosis, Control rats receiving a high protein diet, and animals with cirrhosis receiving a high protein diet. Results showed preserved motor function and normal anxiety levels in all the groups. The animals with cirrhosis showed an impairment in active avoidance behavior and spatial memory, regardless of the diet they received. However, the animals with cirrhosis and a high protein diet showed longer escape latencies on the spatial memory task. The model of cirrhosis presented an under-activation of the dentate gyrus and CA3 hippocampal subfields and the medial part of the medial mammillary nucleus. The results suggest that a high protein intake worsens spatial memory deficits shown by the TAA-induced model of cirrhosis. However, high protein ingestion has no influence on the COx hypoactivity associated with the model. Copyright © 2015 Elsevier Inc. All rights reserved.

  19. Suitability of magnetic single- and multi-core nanoparticles to detect protein binding with dynamic magnetic measurement techniques

    International Nuclear Information System (INIS)

    Remmer, Hilke; Dieckhoff, Jan; Schilling, Meinhard; Ludwig, Frank

    2015-01-01

    We investigated the binding of biotinylated proteins to various streptavidin functionalized magnetic nanoparticles with different dynamic magnetic measurement techniques to examine their potential for homogeneous bioassays. As particle systems, single-core nanoparticles with a nominal core diameter of 30 nm as well as multi-core nanoparticles with hydrodynamic sizes varying between nominally 60 nm and 100 nm were chosen. As experimental techniques, fluxgate magnetorelaxometry (MRX), complex ac susceptibility (ACS) and measurements of the phase lag between rotating field and sample magnetization are applied. MRX measurements are only suited for the detection of small analytes if the multivalency of functionalized nanoparticles and analytes causes cross-linking, thus forming larger aggregates. ACS measurements showed for all nanoparticle systems a shift of the imaginary part's maximum towards small frequencies. In rotating field measurements only the single-core nanoparticle systems with dominating Brownian mechanism exhibit an increase of the phase lag upon binding in the investigated frequency range. The coexistence of Brownian and Néel relaxation processes can cause a more complex phase lag change behavior, as demonstrated for multi-core nanoparticle systems. - Highlights: • Cealization of homogeneous magnetic bioassays using different magnetic techniques. • Comparison of single- and multi-core nanoparticle systems. • ac Susceptibility favorable for detection of small analytes. • Magnetorelaxometry favorable for detection of large analytes or cross-linking assays

  20. [Indicators of protein metabolism in infants with intrauterine dystrophy red various dietary mixtures].

    Science.gov (United States)

    Krukowa, A; Symonowicz, H; Wachnik, Z; Koziej, M

    1979-01-01

    In the previous work published in No 7 of "Development Period Medicine" ( Problemy Medycyny Wieku Rozwojowego ) the results of nitrogen balance studies in S-f-D infants fed different milk formulas were described. The present study concerns other protein metabolism indices in the same infants. The infants were divided into four groups according to the formula they were fed. The composition of formulas is shown in table I. In the infants besides the balance study, serum urea nitrogen, protein and albumin level, were estimated once a month. Also urea, creatine and creatinine, and hydroxyproline in 24-hours urine collections were examined. Excretion of creatine, creatinine and hydroxyproline was summarized in 5 boys from the group of 38 investigated infants in the first five months of life when meat-free diet was fed. The above mentioned indices permit for better assessment of the effect of the diet on protein metabolism and the requirement of protein for S-f-D infants. The results of protein metabolism indices were compared with the indices obtained in F.S. infants similarly fed. Group S of S-f-D infants was compared with group A of F.S. infants and the other groups of S-f-D infants were compared with each other. In S-f-D infants fed formula S, a lower level of serum urea nitrogen was observed in comparison with F.S. infants of group A in spite of greater protein intake in S-f-D infants. This should prove a greater protein requirement in S-f-D infants. Decreased protein content and cow's milk fat modification also had profitable influence on protein utilization because serum urea nitrogen and nitrogen in urine were low in S-f-D infants fed this formula. Urine urea nitrogen as a part of total urine nitrogen is bigger in group S and C infants, and the lowest in group G infants (formula with lower fat and total protein content). Serum protein and albumin level was generally higher in S-f-D infants than in FS ones. Particularly high level of these parameters was observed

  1. Decorin core protein (decoron) shape complements collagen fibril surface structure and mediates its binding.

    Science.gov (United States)

    Orgel, Joseph P R O; Eid, Aya; Antipova, Olga; Bella, Jordi; Scott, John E

    2009-09-15

    Decorin is the archetypal small leucine rich repeat proteoglycan of the vertebrate extracellular matrix (ECM). With its glycosaminoglycuronan chain, it is responsible for stabilizing inter-fibrillar organization. Type I collagen is the predominant member of the fibrillar collagen family, fulfilling both organizational and structural roles in animal ECMs. In this study, interactions between decoron (the decorin core protein) and binding sites in the d and e(1) bands of the type I collagen fibril were investigated through molecular modeling of their respective X-ray diffraction structures. Previously, it was proposed that a model-based, highly curved concave decoron interacts with a single collagen molecule, which would form extensive van der Waals contacts and give rise to strong non-specific binding. However, the large well-ordered aggregate that is the collagen fibril places significant restraints on modes of ligand binding and necessitates multi-collagen molecular contacts. We present here a relatively high-resolution model of the decoron-fibril collagen complex. We find that the respective crystal structures complement each other well, although it is the monomeric form of decoron that shows the most appropriate shape complementarity with the fibril surface and favorable calculated energies of interaction. One molecule of decoron interacts with four to six collagen molecules, and the binding specificity relies on a large number of hydrogen bonds and electrostatic interactions, primarily with the collagen motifs KXGDRGE and AKGDRGE (d and e(1) bands). This work helps us to understand collagen-decorin interactions and the molecular architecture of the fibrillar ECM in health and disease.

  2. Decorin core protein (decoron shape complements collagen fibril surface structure and mediates its binding.

    Directory of Open Access Journals (Sweden)

    Joseph P R O Orgel

    2009-09-01

    Full Text Available Decorin is the archetypal small leucine rich repeat proteoglycan of the vertebrate extracellular matrix (ECM. With its glycosaminoglycuronan chain, it is responsible for stabilizing inter-fibrillar organization. Type I collagen is the predominant member of the fibrillar collagen family, fulfilling both organizational and structural roles in animal ECMs. In this study, interactions between decoron (the decorin core protein and binding sites in the d and e(1 bands of the type I collagen fibril were investigated through molecular modeling of their respective X-ray diffraction structures. Previously, it was proposed that a model-based, highly curved concave decoron interacts with a single collagen molecule, which would form extensive van der Waals contacts and give rise to strong non-specific binding. However, the large well-ordered aggregate that is the collagen fibril places significant restraints on modes of ligand binding and necessitates multi-collagen molecular contacts. We present here a relatively high-resolution model of the decoron-fibril collagen complex. We find that the respective crystal structures complement each other well, although it is the monomeric form of decoron that shows the most appropriate shape complementarity with the fibril surface and favorable calculated energies of interaction. One molecule of decoron interacts with four to six collagen molecules, and the binding specificity relies on a large number of hydrogen bonds and electrostatic interactions, primarily with the collagen motifs KXGDRGE and AKGDRGE (d and e(1 bands. This work helps us to understand collagen-decorin interactions and the molecular architecture of the fibrillar ECM in health and disease.

  3. Adherence issues in inherited metabolic disorders treated by low natural protein diets

    DEFF Research Database (Denmark)

    MaCdonald, A; van Rijn, M; Feillet, F

    2012-01-01

    Common inborn errors of metabolism treated by low natural protein diets [amino acid (AA) disorders, organic acidemias and urea cycle disorders] are responsible for a collection of diverse clinical symptoms, each condition presenting at different ages with variable severity. Precursor......-free or essential L-AAs are important in all these conditions. Optimal long-term outcome depends on early diagnosis and good metabolic control, but because of the rarity and severity of conditions, randomized controlled trials are scarce. In all of these disorders, it is commonly described that dietary adherence...... on their neuropsychological profile. There are little data about their ability to self-manage their own diet or the success of any formal educational programs that may have been implemented. Trials conducted in non-phenylketonuria (PKU) patients are rare, and the development of specialist L-AAs for non-PKU AA disorders has...

  4. Effects of Radiation and Dietary Iron on Expression of Genes and Proteins Involved in Drug Metabolism

    Science.gov (United States)

    Faust, K. M.; Wotring, V. E.

    2014-01-01

    Liver function, especially the rate of metabolic enzyme activities, determines the concentration of circulating drugs and the duration of their efficacy. Most pharmaceuticals are metabolized by the liver, and clinically-used medication doses are given with normal liver function in mind. A drug overdose can result in the case of a liver that is damaged and removing pharmaceuticals from the circulation at a rate slower than normal. Alternatively, if liver function is elevated and removing drugs from the system more quickly than usual, it would be as if too little drug had been given for effective treatment. Because of the importance of the liver in drug metabolism, we want to understand any effects of spaceflight on the enzymes of the liver. Dietary factors and exposure to radiation are aspects of spaceflight that are potential oxidative stressors and both can be modeled in ground experiments. In this experiment, we examined the effects of high dietary iron and low dose gamma radiation (individually and combined) on the gene expression of enzymes involved in drug metabolism, redox homeostasis, and DNA repair. METHODS All procedures were approved by the JSC Animal Care and Use Committee. Male Sprague-Dawley rats were divided into 4 groups (n=8); control, high Fe diet (650 mg iron/kg), radiation (fractionated 3 Gy exposure from a Cs- 137 source) and combined high Fe diet + radiation exposure. Animals were euthanized 24h after the last treatment of radiation; livers were removed immediately and flash -frozen in liquid nitrogen. Expression of genes thought to be involved in redox homeostasis, drug metabolism and DNA damage repair was measured by RT-qPCR. Where possible, protein expression of the same genes was measured by western blotting. All data are expressed as % change in expression normalized to reference gene expression; comparisons were then made of each treatment group to the sham exposed/ normal diet control group. Data was considered significant at phigh Fe

  5. A role for 12/15 lipoxygenase in the amyloid beta precursor protein metabolism.

    Science.gov (United States)

    Succol, Francesca; Praticò, Domenico

    2007-10-01

    12/15 Lipoxygenase (12/15LO) protein levels and activity are increased in pathologically affected regions of Alzheimer's disease (AD) brains, compared with controls. Its metabolic products are elevated in cerebrospinal fluid of patients with AD and individuals with mild cognitive impairment, suggesting that this enzyme may be involved early in AD pathogenesis. Herein, we investigate the effect of pharmacologic inhibition of 12/15LO on the amyloid beta precursor protein (APP) metabolism. To this end, we used CHO and N2A cells stably expressing human APP with the Swedish mutant, and two structurally distinct and selective 12/15LO inhibitors, PD146176 and CDC. Our results demonstrated that both drugs dose-dependently reduced Abeta formation without affecting total APP levels. Interestingly, in the same cells we observed a significant reduction in secreted (s)APPbeta and beta-secretase (BACE), but not sAPPalpha and ADAM10 protein levels. Together, these data show for the first time that this enzymatic pathway influences Abeta formation whereby modulating the BACE proteolytic cascade. We conclude that specific pharmacologic inhibition of 12/15LO could represent a novel therapeutic target for treating or preventing AD pathology in humans.

  6. Trehalose Alters Subcellular Trafficking and the Metabolism of the Alzheimer-associated Amyloid Precursor Protein.

    Science.gov (United States)

    Tien, Nguyen T; Karaca, Ilker; Tamboli, Irfan Y; Walter, Jochen

    2016-05-13

    The disaccharide trehalose is commonly considered to stimulate autophagy. Cell treatment with trehalose could decrease cytosolic aggregates of potentially pathogenic proteins, including mutant huntingtin, α-synuclein, and phosphorylated tau that are associated with neurodegenerative diseases. Here, we demonstrate that trehalose also alters the metabolism of the Alzheimer disease-related amyloid precursor protein (APP). Cell treatment with trehalose decreased the degradation of full-length APP and its C-terminal fragments. Trehalose also reduced the secretion of the amyloid-β peptide. Biochemical and cell biological experiments revealed that trehalose alters the subcellular distribution and decreases the degradation of APP C-terminal fragments in endolysosomal compartments. Trehalose also led to strong accumulation of the autophagic marker proteins LC3-II and p62, and decreased the proteolytic activation of the lysosomal hydrolase cathepsin D. The combined data indicate that trehalose decreases the lysosomal metabolism of APP by altering its endocytic vesicular transport. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Protein metabolism in the small intestine during cancer cachexia and chemotherapy in mice.

    Science.gov (United States)

    Samuels, S E; Knowles, A L; Tilignac, T; Debiton, E; Madelmont, J C; Attaix, D

    2000-09-01

    The impact of cancer cachexia and chemotherapy on small intestinal protein metabolism and its subsequent recovery was investigated. Cancer cachexia was induced in mice with colon 26 adenocarcinoma, which is a small and slow-growing tumor characteristic of the human condition, and can be cured with 100% efficacy using an experimental nitrosourea, cystemustine (C6H12ClN3O4S). Both healthy mice and tumor-bearing mice were given a single i.p. injection of cystemustine (20 mg/kg) 3 days after the onset of cachexia. Cancer cachexia led to a reduced in vivo rate of protein synthesis in the small intestine relative to healthy mice (-13 to -34%; P synthesis compared with healthy mice (23-34%; P < 0.05). Northern hybridizations of mRNA encoding components of the major proteolytic systems suggested that proteolysis may not have mediated intestinal wasting or recovery. A major clinical goal should be to design methods to improve small intestinal protein metabolism before the initiation of chemotherapy.

  8. Effects of rumen undegradable protein supplementation on productive performance and indicators of protein and energy metabolism in Holstein fresh cows.

    Science.gov (United States)

    Amanlou, H; Farahani, T Amirabadi; Farsuni, N Eslamian

    2017-05-01

    The objective of this study was to determine the effects of feeding increased dietary crude protein (CP) on productive performance and indicators of protein and energy metabolism during 21 d postpartum. Thirty multiparous Holstein dairy cows were balanced by previous lactation milk yield, body condition score (BCS) at calving, and parity and randomly allocated to 1 of 3 dietary treatments from calving until 21 d postpartum. Dietary treatments were 16.0% CP with 5.0% rumen undegradable protein (RUP) based on dry matter (DM) (16CP), 18.7% CP with 7.0% RUP based on DM (19CP), and 21.4% CP with 9.0% RUP based on DM (21CP). Diets were similar in net energy for lactation (approximately 1.7 Mcal/kg of DM) and CP levels were increased with corn gluten meal and fish meal. Dry matter intake (DMI) was increased by increasing dietary CP levels from 16.0 to 19.0% of DM, but dietary CP beyond 19.0% had no effect on DMI. Milk yields were 4.7 and 6.5 kg/d greater in cows fed the 19CP and 21CP diets versus those fed the 16CP diet, whereas 4% fat-corrected milk was greater for cows fed the 21CP than the 16CP diet (36.0 vs. 31.4 kg/d). Milk protein content and yield, lactose yield, and milk urea nitrogen were elevated by increased dietary CP. Milk lactose content and fat yield were not different among dietary treatments, but milk fat content tended to decline with increasing content of CP in diets. High CP levels increased milk N secretion but decreased milk N efficiency. Apparent digestibility of DM, CP, and neutral detergent fiber was greater on the 19CP and 21CP diets compared with the 16CP diet. Cows fed the 19CP and 21CP diets lost less body condition relative to those fed the 16CP diet over 21 d postpartum. Feeding higher CP levels increased the concentrations of serum albumin, albumin to globulin ratio, and urea nitrogen and decreased aspartate aminotransferase, nonesterified fatty acids, and β-hydroxybutyrate, but had no effect on globulin, glucose, cholesterol, or

  9. Effect of protein provision via milk replacer or solid feed on protein metabolism in veal calves

    DEFF Research Database (Denmark)

    Berends, H.; van den Borne, J. J G C; Røjen, B. A.

    2015-01-01

    recycling but urea reused for anabolism remained unaffected. Total-tract neutral detergent fiber digestibility decreased (-9%) with increasing low-N SF intake, indicating reduced rumen fermentation. Increasing the N content of SF at equal total N intake resulted in decreased urea production, excretion......The current study evaluated the effects of protein provision to calves fed a combination of solid feed (SF) and milk replacer (MR) at equal total N intake on urea recycling and N retention. Nitrogen balance traits and [15N2]urea kinetics were measured in 30 calves (23 wk of age, 180±3.7kg of body...... of calves for 5 d, and for the assessment of urea recycling from [15N2]urea kinetics. Increasing low-N SF intake at equal total N intake resulted in a shift from urinary to fecal N excretion but did not affect protein retention (0.71g of N·kg of BW-0.75·d-1). Increasing low-N SF intake increased urea...

  10. Hepatitis C virus core protein expression leads to biphasic regulation of the p21 cdk inhibitor and modulation of hepatocyte cell cycle

    International Nuclear Information System (INIS)

    Nguyen, Hau; Mudryj, Maria; Guadalupe, Moraima; Dandekar, Satya

    2003-01-01

    Hepatitis C virus (HCV) Core protein is implicated in viral pathogenesis by the modulation of hepatocyte gene expression and function. To determine the effect of Core protein on the cell-cycle control of hepatocytes, a HepG2 cell line containing a Flag-tagged Core under the control of an inducible promoter was generated. Initial Core protein expression included the presence of unprocessed (191 aa) and processed (173 aa) forms of the Core proteins with the processed form becoming dominant later. Expression of the 191 aa form of Core protein corresponded to an increase in the expression of the p21, a decrease in cdk2-dependent kinase activity, and a decrease in the percentage of cells in S-phase along with an accumulation of cells in the G 0 /G 1 phase of the cell cycle. As the processed form accumulated, the p21 levels started to decline, suggesting that Core protein regulates p21 expression in a biphasic manner. These findings implicate Core protein in potentially modulating hepatocyte cell cycle differentially in the early stages of infection through biphasic regulation of p21 cdk kinase inhibitor

  11. Quantitative proteome and phosphoproteome analyses of Streptomyces coelicolor reveal proteins and phosphoproteins modulating differentiation and secondary metabolism

    DEFF Research Database (Denmark)

    Rioseras, Beatriz; Sliaha, Pavel V; Gorshkov, Vladimir

    2018-01-01

    identified and quantified 3461 proteins corresponding to 44.3% of the S. coelicolor proteome across three developmental stages: vegetative hypha (MI); secondary metabolite producing hyphae (MII); and sporulating hyphae. A total of 1350 proteins exhibited more than 2-fold expression changes during....../Thr/Tyr kinases, making this genus an outstanding model for the study of bacterial protein phosphorylation events. We used mass spectrometry based quantitative proteomics and phosphoproteomics to characterize bacterial differentiation and activation of secondary metabolism of Streptomyces coelicolor. We...... the bacterial differentiation process. These proteins include 136 regulators (transcriptional regulators, transducers, Ser/Thr/Tyr kinases, signalling proteins), as well as 542 putative proteins with no clear homology to known proteins which are likely to play a role in differentiation and secondary metabolism...

  12. Perturbed rhythmic activation of signaling pathways in mice deficient for Sterol Carrier Protein 2-dependent diurnal lipid transport and metabolism.

    Science.gov (United States)

    Jouffe, Céline; Gobet, Cédric; Martin, Eva; Métairon, Sylviane; Morin-Rivron, Delphine; Masoodi, Mojgan; Gachon, Frédéric

    2016-04-21

    Through evolution, most of the living species have acquired a time keeping system to anticipate daily changes caused by the rotation of the Earth. In all of the systems this pacemaker is based on a molecular transcriptional/translational negative feedback loop able to generate rhythmic gene expression with a period close to 24 hours. Recent evidences suggest that post-transcriptional regulations activated mostly by systemic cues play a fundamental role in the process, fine tuning the time keeping system and linking it to animal physiology. Among these signals, we consider the role of lipid transport and metabolism regulated by SCP2. Mice harboring a deletion of the Scp2 locus present a modulated diurnal accumulation of lipids in the liver and a perturbed activation of several signaling pathways including PPARα, SREBP, LRH-1, TORC1 and its upstream regulators. This defect in signaling pathways activation feedbacks upon the clock by lengthening the circadian period of animals through post-translational regulation of core clock regulators, showing that rhythmic lipid transport is a major player in the establishment of rhythmic mRNA and protein expression landscape.

  13. Clofazimine modulates the expression of lipid metabolism proteins in Mycobacterium leprae-infected macrophages.

    Science.gov (United States)

    Degang, Yang; Akama, Takeshi; Hara, Takeshi; Tanigawa, Kazunari; Ishido, Yuko; Gidoh, Masaichi; Makino, Masahiko; Ishii, Norihisa; Suzuki, Koichi

    2012-01-01

    Mycobacterium leprae (M. leprae) lives and replicates within macrophages in a foamy, lipid-laden phagosome. The lipids provide essential nutrition for the mycobacteria, and M. leprae infection modulates expression of important host proteins related to lipid metabolism. Thus, M. leprae infection increases the expression of adipophilin/adipose differentiation-related protein (ADRP) and decreases hormone-sensitive lipase (HSL), facilitating the accumulation and maintenance of lipid-rich environments suitable for the intracellular survival of M. leprae. HSL levels are not detectable in skin smear specimens taken from leprosy patients, but re-appear shortly after multidrug therapy (MDT). This study examined the effect of MDT components on host lipid metabolism in vitro, and the outcome of rifampicin, dapsone and clofazimine treatment on ADRP and HSL expression in THP-1 cells. Clofazimine attenuated the mRNA and protein levels of ADRP in M. leprae-infected cells, while those of HSL were increased. Rifampicin and dapsone did not show any significant effects on ADRP and HSL expression levels. A transient increase of interferon (IFN)-β and IFN-γ mRNA was also observed in cells infected with M. leprae and treated with clofazimine. Lipid droplets accumulated by M. leprae-infection were significantly decreased 48 h after clofazimine treatment. Such effects were not evident in cells without M. leprae infection. In clinical samples, ADRP expression was decreased and HSL expression was increased after treatment. These results suggest that clofazimine modulates lipid metabolism in M. leprae-infected macrophages by modulating the expression of ADRP and HSL. It also induces IFN production in M. leprae-infected cells. The resultant decrease in lipid accumulation, increase in lipolysis, and activation of innate immunity may be some of the key actions of clofazimine.

  14. Whole body and forearm substrate metabolism in hyperthyroidism: evidence of increased basal muscle protein breakdown.

    Science.gov (United States)

    Riis, Anne Lene Dalkjaer; Jørgensen, Jens Otto Lunde; Gjedde, Signe; Nørrelund, Helene; Jurik, Anne Grethe; Nair, K S; Ivarsen, Per; Weeke, Jørgen; Møller, Niels

    2005-06-01

    Thyroid hormones have significant metabolic effects, and muscle wasting and weakness are prominent clinical features of chronic hyperthyroidism. To assess the underlying mechanisms, we examined seven hyperthyroid women with Graves' disease before (Ht) and after (Eut) medical treatment and seven control subjects (Ctr). All subjects underwent a 3-h study in the postabsorptive state. After regional catheterization, protein dynamics of the whole body and of the forearm muscles were measured by amino acid tracer dilution technique using [15N]phenylalanine and [2H4]tyrosine. Before treatment, triiodothyronine was elevated (6.6 nmol/l) and whole body protein breakdown was increased 40%. The net forearm release of phenylalanine was increased in hyperthyroidism (microg.100 ml(-1).min(-1)): -7.0 +/- 1.2 Ht vs. -3.8 +/- 0.8 Eut (P = 0.04), -4.2 +/- 0.3 Ctr (P = 0.048). Muscle protein breakdown, assessed by phenylalanine rate of appearance, was increased (microg.100 ml(-1).min(-1)): 15.5 +/- 2.0 Ht vs. 9.6 +/- 1.4 Eut (P = 0.03), 9.9 +/- 0.6 Ctr (P = 0.02). Muscle protein synthesis rate did not differ significantly. Muscle mass and muscle function were decreased 10-20% before treatment. All abnormalities were normalized after therapy. In conclusion, our results show that hyperthyroidism is associated with increased muscle amino acid release resulting from increased muscle protein breakdown. These abnormalities can explain the clinical manifestations of sarcopenia and myopathy.

  15. Protein metabolism in slow- and fast-twitch skeletal muscle during turpentine-induced inflammation.

    Science.gov (United States)

    Muthny, Tomas; Kovarik, Miroslav; Sispera, Ludek; Tilser, Ivan; Holecek, Milan

    2008-02-01

    The aim of our study was to evaluate the differences in protein and amino acid metabolism after subcutaneous turpentine administration in the soleus muscle (SOL), predominantly composed of red fibres, and the extensor digitorum longus muscle (EDL) composed of white fibres. Young rats (40-60 g) were injected subcutaneously with 0.2 ml of turpentine oil/100 g body weight (inflammation) or with the same volume of saline solution (control). Twenty-four hours later SOL and EDL were dissected and incubated in modified Krebs-Heinseleit buffer to estimate total and myofibrillar proteolysis, chymotrypsin-like activity of proteasome (CHTLA), leucine oxidation, protein synthesis and amino acid release into the medium. The data obtained demonstrate that in intact rats, all parameters measured except protein synthesis are significantly higher in SOL than in EDL. In turpentine treated animals, CHTLA increased and protein synthesis decreased significantly more in EDL. Release of leucine was inhibited significantly more in SOL. We conclude that turpentine-induced inflammation affects more CHTLA, protein synthesis and leucine release in EDL compared to SOL.

  16. Soy Germ Protein With or Without-Zn Improve Plasma Lipid Profile in Metabolic Syndrome Women

    Directory of Open Access Journals (Sweden)

    SIWI PRAMATAMA MARS WIJAYANTI

    2012-03-01

    Full Text Available The aim of this research was to determine the effect of soy germ protein on lipid profile of metabolic syndrome (MetS patients. Respondents were 30 women with criteria, i.e. blood glucose level > normal, body mass index > 25 kg/m2, hypertriglyceridemia, low cholesterol-HDL level, 40-65 years old, living in Purwokerto, and signed the informed consent. The project was approved by the ethics committee of the Medical Faculty from Gadjah Mada University-Yogyakarta. Respondents were divided into three randomly chosen groups consisting of ten women each. The first, second, and third groups were treated, respectively, with milk enriched soy germ protein plus Zn, milk enriched soy germ protein (without Zn, and placebo for two months. Blood samples were taken at baseline, one and two months after observation. Two months after observation the groups consuming milk enriched with soy germ protein, both with or without Zn, had their level of cholesterol-total decrease from 215.8 to 180.2 mg/dl (P = 0.03, triglyceride from 240.2 to 162.5 mg/dl (P = 0.02, and LDL from 154.01 to 93.85 mg/dl (P = 0.03. In contrast, HDL increased from 38.91 to 49.49 mg/dl (P = 0.0008. In conclusion, soy germ protein can improve lipid profile, thus it can inhibit atherosclerosis incident.

  17. Changes in protein metabolism after gastric resection studied by 125I-albumin

    International Nuclear Information System (INIS)

    Beno, I.; Cerven, J.

    1976-01-01

    The changes were studied in the metabolism of protein using albumin- 125 I in seven patients with benign conditions of the stomach or duodenum before gastrectomy and starting with the second week after the surgery. In the postoperative period body weight was found to be significantly reduced, there was a drop in erythrocyte count, and blood hemoglobin and plasma albumin concentration were decreased. There was a significant rise of plasma volume during this period. Compared with the preoperative findings, the intravascular albumin pool was diminished by 11%, the extravascular albumin pool by 19.%, so that the overall albumin pool was postoperatively found to be reduced by 1/6. (author)

  18. Conformational Changes in the Hepatitis B Virus Core Protein Are Consistent with a Role for Allostery in Virus Assembly

    Energy Technology Data Exchange (ETDEWEB)

    Packianathan, Charles; Katen, Sarah P.; Dann, III, Charles E.; Zlotnick, Adam (Indiana)

    2010-01-12

    In infected cells, virus components must be organized at the right place and time to ensure assembly of infectious virions. From a different perspective, assembly must be prevented until all components are available. Hypothetically, this can be achieved by allosterically controlling assembly. Consistent with this hypothesis, here we show that the structure of the hepatitis B virus (HBV) core protein dimer, which can spontaneously self-assemble, is incompatible with capsid assembly. Systematic differences between core protein dimer and capsid conformations demonstrate linkage between the intradimer interface and interdimer contact surface. These structures also provide explanations for the capsid-dimer selectivity of some antibodies and the activities of assembly effectors. Solution studies suggest that the assembly-inactive state is more accurately an ensemble of conformations. Simulations show that allostery supports controlled assembly and results in capsids that are resistant to dissociation. We propose that allostery, as demonstrated in HBV, is common to most self-assembling viruses.

  19. [Low-molecular-weight regulators of biogenic polyamine metabolism affect cytokine production and expression of hepatitis С virus proteins in Huh7.5 human hepatocarcinoma cells].

    Science.gov (United States)

    Masalova, O V; Lesnova, E I; Samokhvalov, E I; Permyakova, K Yu; Ivanov, A V; Kochetkov, S N; Kushch, A A

    2017-01-01

    Hepatitis C virus (HCV) induces the expression of the genes of proinflammatory cytokines, the excessive production of which may cause cell death, and contribute to development of liver fibrosis and hepatocarcinoma. The relationship between cytokine production and metabolic disorders in HCV-infected cells remains obscure. The levels of biogenic polyamines, spermine, spermidine, and their precursor putrescine, may be a potential regulator of these processes. The purpose of the present work was to study the effects of the compounds which modulate biogenic polyamines metabolism on cytokine production and HCV proteins expression. Human hepatocarcinoma Huh7.5 cells have been transfected with the plasmids that encode HCV proteins and further incubated with the following low-molecular compounds that affect different stages of polyamine metabolism: (1) difluoromethylornithine (DFMO), the inhibitor of ornithine decarboxylase, the enzyme that catalyzes the biosynthesis of polyamines; (2) N,N'-bis(2,3-butane dienyl)-1,4-diaminobutane (MDL72.527), the inhibitor of proteins involved in polyamine degradation; and (3) synthetic polyamine analog N^(I),N^(II)-diethylnorspermine (DENSpm), an inducer of polyamine degradation enzyme. The intracellular accumulation and secretion of cytokines (IL-6, IL-1β, TNF-α, and TGF-β) was assessed by immunocytochemistry and in the immunoenzyme assay, while the cytokine gene expression was studied using reverse transcription and PCR. The effects of the compounds under analysis on the expression of HCV proteins were analyzed using the indirect immunofluorescence with anti-HCV monoclonal antibodies. It has been demonstrated that, in cells transfected with HCV genes, DFMO reduces the production of three out of four tested cytokines, namely, TNF-α and TGF-β in cells that express HCV core, Е1Е2, NS3, NS5A, and NS5B proteins, and IL-1β in the cells that express HCV core, Е1Е2, and NS3 proteins. MDL72527 and DENSpm decreased cytokine production

  20. Investigation of carbohydrate and protein metabolism in the digestive organs of the rabbit under the combined influence of vibration, acceleration and irradiation

    Science.gov (United States)

    Yuy, R. I.

    1975-01-01

    During spaceflight, the organism is subjected to the influence of various extremal factors such as acceleration, vibration, irradiation, etc. The study of the influence of these factors on metabolism, especially carbohydrate and protein metabolism, in young rabbits is of great significance in simulation experiments. Dynamic factors and irradiation, depending on dose and duration, lead to reduced RNA and protein metabolism.

  1. Multi-omic profiling of EPO-producing Chinese hamster ovary cell panel reveals metabolic adaptation to heterologous protein production

    DEFF Research Database (Denmark)

    Ley, Daniel; Kazemi Seresht, Ali; Engmark, Mikael

    2015-01-01

    Chinese hamster ovary (CHO) cells are the preferred production host for many therapeutic proteins. The production of heterologous proteins in CHO cells imposes a burden on the host cell metabolism and impact cellular physiology on a global scale. In this work, a multi-omics approach was applied...

  2. Energy metabolism in young mink kits (Neovison vison) affected by protein and carbohydrate level in the diet

    DEFF Research Database (Denmark)

    Hellwing, Anne Louise Frydendahl; Hansen, Niels Enggaard; Tauson, Anne-Helene

    2010-01-01

    The mink is a strict carnivore and mink diets usually have a high content of protein. The energy metabolism in young minks in the transition period from milk to solid food is not investigated in detail, and the protein requirement is poorly defined. The substrate oxidation can give useful...

  3. Protein metabolism in the rat cerebral cortex in vivo and in vitro as affected by the acquisition enhancing drug piracetam

    NARCIS (Netherlands)

    Nickolson, V.J.; Wolthuis, O.L.

    1976-01-01

    The effect of Piracetam on rat cerebral protein metabolism in vivo and in vitro was studied. It was found that the drug stimulates the uptake of labelled leucine by cerebral cortex slices, has no effect on the incorporation of leucine into cerebral protein, neither in slices nor in vivo, but

  4. An increase level of acylation stimulating protein is correlated with metabolic risk markers in North Indian obese women.

    Science.gov (United States)

    Mishra, Supriya; Gupta, Vani; Mishra, Sameeksha; Gupta, Vandana; Mahdi, Abbas Ali; Sachan, Rekha

    2017-12-01

    The present study was to investigate the association between serum acylation stimulating protein (ASP) level with metabolic risk factors in North Indian obese women. This is a case control study, total n=322 women aged between 20 and 45 years (n=162 with metabolic syndrome & n=160 without metabolic syndrome) were recruited for the study according to National Cholesterol Education Program Treatment Panel (NCEPATP) guidelines. Serum ASP level were determined by enzyme linked immunosorbent assay. Results indicated that circulating ASP and other metabolic risk factors (waist circumference, triglycerides, fasting plasma glucose etc) were significantly higher in women with metabolic syndrome (WmetS) than in women without syndrome (WometS) (pwomen with metabolic syndrome. Conclusively circulating ASP was found to be significantly associated with hyperlipidemia, obesity and obesity related disorders in North Indian obese women. Copyright © 2017 Diabetes India. Published by Elsevier Ltd. All rights reserved.

  5. Multifunctional Fe3O4/Au core/satellite nanocubes: an efficient chemical synthesis, characterization and functionalization of streptavidin protein.

    Science.gov (United States)

    Abbas, Mohamed; RamuluTorati, Sri; Kim, CheolGi

    2017-02-14

    A novel and efficient chemical approach for the synthesis of Fe 3 O 4 /Au core/satellite nanocubes is reported. In a one-pot reaction, metallic Au nanodots were successfully deposited on the polyvinylpyrrolidone (PVP) functionalized Fe 3 O 4 nanocube surface for the fabrication of a core/satellite structure (Fe 3 O 4 /Au) by the reduction of HAuCl 4 using ammonia. Transmission electron microscopy and energy dispersive spectroscopy mapping revealed that small Au nanodots of about 2 nm average size decorated the surface of Fe 3 O 4 nanocubes. X-ray diffraction data was used to confirm the formation of both the phases of a cubic inverse spinel structure for Fe 3 O 4 and a bcc structure for Au in the core/satellite structure of Fe 3 O 4 /Au nanocubes. The magnetic properties of the seed Fe 3 O 4 nanocubes and Fe 3 O 4 /Au core/satellite nanocubes were measured by using a superconducting quantum interference device at 300 K. For biological application purposes, the as-synthesized Fe 3 O 4 /Au core/satellite nanocubes were functionalized by cysteamine followed by successful immobilization of streptavidin protein as confirmed through the fluorescence confocal microscopy images.

  6. Abomasal protein infusion in postpartum transition dairy cows: Effect on performance and mammary metabolism

    DEFF Research Database (Denmark)

    Larsen, Mogens; Lapierre, H; Kristensen, Niels Bastian

    2014-01-01

    The effect of increasing the postpartum metabolizable protein (MP) supply on performance and mammary metabolism was studied using 8 Holstein cows in a complete randomized design. At parturition, cows were assigned to abomasal infusion of water (CTRL) or casein (CAS). Arterial and epigastric venous....../d for CTRL, but did not differ at 29 DIM (1,383 ± 48 g/d). The ratio of MP total supply to requirement was numerically greater at 4 DIM for CAS compared with CTRL, indicating less postpartum protein deficiency. In contrast, a greater net energy deficiency tended to be induced with CAS, but the greater milk...... of glucose, lactate, and β-hydroxybutyrate, whereas uptakes of volatile fatty acids were unaffected. Despite similar MP supply by 29 DIM, milk and lactose yields were greater with CAS indicating a persistent response to increased postpartum MP supply. In conclusion, the postpartum MP deficiency can have...

  7. Cytosolic fatty acid-binding proteins: subjects and tools in metabolic research

    Energy Technology Data Exchange (ETDEWEB)

    Binas, B. [Max Delbrueck Center for Molecular Medicine, Berlin-Buch (Germany)

    1998-12-31

    Fatty acid-binding proteins (FABPs) are major targets for specific binding of fatty acids in vivo. They constitute a widely expressed family of genetically related, small cytosolic proteins which very likely mediate intracellular transport of free long chain fatty acids. Genetic inhibition of FABP expression in vivo should therefore provide a useful tool to investigate and engineer fatty acid metabolism. (orig.) [Deutsch] Fettsaeurebindungsproteine (FABPs) sind wichtige Bindungsstellen fuer Fettsaeuren in vivo; sie bilden eine breit exprimierte Familie genetisch verwandter kleiner Zytosoleiweisse, die sehr wahrscheinlich den intrazellulaeren Transport unveresterter langkettiger Fettsaeuren vermitteln. Die genetische Hemmung der FABP-Expanssion in vivo bietet sich deshalb als Werkzeug zur Erforschung und gezielten Veraenderung des Fettsaeurestoffwechsels an. (orig.)

  8. Ecdysteroids affect in vivo protein metabolism of the flight muscle of the tobacco hornworm (Manduca sexta)

    Science.gov (United States)

    Tischler, M. E.; Wu, M.; Cook, P.; Hodsden, S.

    1990-01-01

    Ecdysteroid growth promotion of the dorsolongitudinal flight muscle of Manduca sexta was studied by measuring in vivo protein metabolism using both "flooding-dose" and "non-carrier" techniques. These procedures differ in that the former method includes injection of non-labelled phenylalanine (30 micromoles/insect) together with the [3H]amino acid. Injected radioactivity plateaued in the haemolymph within 7 min. With the flooding-dose method, haemolymph and intramuscular specific radioactivities were similar between 15 min and 2 h. Incorporation of [3H]phenylalanine into muscle protein was linear with either method between 30 and 120 min. Fractional rates (%/12 h) of synthesis with the flooding-dose technique were best measured after 1 h because of the initial delay in radioactivity equilibration. Estimation of body phenylalanine turnover with the non-carrier method showed 24-53%/h which was negligible with the flooding-dose method. Since the two methods yielded similar rates of protein synthesis, the large injection of non-labelled amino acid did not alter the rate of synthesis. Because the flooding-dose technique requires only a single time point measurement, it is the preferred method. The decline and eventual cessation of flight-muscle growth was mostly a consequence of declining protein synthesis though degradation increased between 76-86 h before eclosion and was relatively rapid. This decline in muscle growth could be prevented by treating pupae with 20-hydroxyecdysone (10 micrograms/insect). Protein accretion was promoted by a decline of up to 80% in protein breakdown, which was offset in part by a concurrent though much smaller decrease in protein synthesis. Therefore, ecdysteroids may increase flight-muscle growth by inhibiting proteolysis.

  9. Anesthesia with halothane and nitrous oxide alters protein and amino acid metabolism in dogs

    International Nuclear Information System (INIS)

    Horber, F.F.; Krayer, S.; Rehder, K.; Haymond, M.W.

    1988-01-01

    General anesthesia in combination with surgery is known to result in negative nitrogen balance. To determine whether general anesthesia without concomitant surgery decreases whole body protein synthesis and/or increases whole body protein breakdown, two groups of dogs were studied: Group 1 (n = 6) in the conscious state and Group 2 (n = 8) during general anesthesia employing halothane (1.5 MAC) in 50% nitrous oxide and oxygen. Changes in protein metabolism were estimated by isotope dilution techniques employing simultaneous infusions of [4,53H]leucine and alpha-[1-14C]-ketoisocaproate (KIC). Total leucine carbon flux was unchanged or slightly increased in the anesthetized animals when compared to the conscious controls, indicating only a slight increase in the rate of proteolysis. However, leucine oxidation was increased (P less than 0.001) by more than 80% in the anesthetized animals when compared with their conscious controls, whereas whole body nonoxidative leucine disappearance, an indicator of whole body protein synthesis, was decreased. The ratio of leucine oxidation to the nonoxidative rate of leucine disappearance, which provides an index of the catabolism of at least one essential amino acid in the postabsorptive state, was more than twofold increased (P less than 0.001) in the anesthetized animals regardless of the tracer employed. These studies suggest that the administration of anesthesia alone, without concomitant surgery, is associated with a decreased rate of whole body protein synthesis and increased leucine oxidation, resulting in increased leucine and protein catabolism, which may be underlying or initiating some of the protein wasting known to occur in patients undergoing surgery

  10. The impact of visual media to encourage low protein cooking in inherited metabolic disorders.

    Science.gov (United States)

    Evans, S; Daly, A; Hopkins, V; Davies, P; MacDonald, A

    2009-10-01

    The use of educational visual aids is one way to help children with inherited metabolic disorders (IMD) understand and develop a positive attitude towards their low protein diet. However, it is difficult to establish their effectiveness in the clinical setting. The present study aimed to evaluate the impact of a low protein recipe book and accompanying DVD for children with IMD. One hundred and five children (53% female; median age = 6-8 years) with IMD on low protein diets were each given a low protein recipe book and DVD. After 6 months, children and carers were posted a questionnaire asking whether they used these resources; identifying any change in frequency of low protein cooking; and the outcome when preparing recipes. One hundred and two questionnaires were returned, representing 105 patients. Seventy percent (n = 71) of questionnaires were from carers. Ninety-three percent (n = 66) of carers acknowledged receipt of the resource; one-third (n = 22) had not watched the DVD and 23% (n = 15) had not opened the recipe book; 55% (n = 36) had tried the recipes; and 71% (n = 47) said the recipe book and/or DVD motivated them to try new recipes. Children were more likely to have watched the DVD (75%; n = 21/28) and read the recipe book (86%; n = 24/28) than carers. Although a helpful educational tool, just over one-half of respondents had used the resource. Identifying visual media that, by itself, will motivate most families of children with IMD to prepare low protein recipes may be unrealistic. The combined approach of visual aids and 'hands-on' practical experience, such as low protein cooking workshops and individual counselling, may be more beneficial.

  11. The hepatitis C virus Core protein is a potent nucleic acid chaperone that directs dimerization of the viral (+) strand RNA in vitro.

    Science.gov (United States)

    Cristofari, Gaël; Ivanyi-Nagy, Roland; Gabus, Caroline; Boulant, Steeve; Lavergne, Jean-Pierre; Penin, François; Darlix, Jean-Luc

    2004-01-01

    The hepatitis C virus (HCV) is an important human pathogen causing chronic hepatitis, liver cirrhosis and hepatocellular carcinoma. HCV is an enveloped virus with a positive-sense, single-stranded RNA genome encoding a single polyprotein that is processed to generate viral proteins. Several hundred molecules of the structural Core protein are thought to coat the genome in the viral particle, as do nucleocapsid (NC) protein molecules in Retroviruses, another class of enveloped viruses containing a positive-sense RNA genome. Retroviral NC proteins also possess nucleic acid chaperone properties that play critical roles in the structural remodelling of the genome during retrovirus replication. This analogy between HCV Core and retroviral NC proteins prompted us to investigate the putative nucleic acid chaperoning properties of the HCV Core protein. Here we report that Core protein chaperones the annealing of complementary DNA and RNA sequences and the formation of the most stable duplex by strand exchange. These results show that the HCV Core is a nucleic acid chaperone similar to retroviral NC proteins. We also find that the Core protein directs dimerization of HCV (+) RNA 3' untranslated region which is promoted by a conserved palindromic sequence possibly involved at several stages of virus replication.

  12. Clinical and protein metabolic efficacy of glutamine granules-supplemented enteral nutrition in severely burned patients.

    Science.gov (United States)

    Peng, Xi; Yan, Hong; You, Zhongyi; Wang, Pei; Wang, Shiliang

    2005-05-01

    As an abundant amino acid in the human body, glutamine has many important metabolic roles that may protect or promote tissue integrity and enhance the immune system. A relative deficiency of glutamine in such patients could compromise recovery and result in prolonged illness and an increase in late mortality. The purpose of this clinical study is to observe the effects of enteral supplement with glutamine granules on protein metabolism in severely burned patients. Forty-eight severe burn patients (total burn surface area 30-75%, full thickness burn area 20-58%) who met the requirements of the protocol joined this double-blind randomized controlled clinical trial. Patients were randomly divided into two groups: burn control group (B group, 23 patients) and glutamine treated group (Gln group, 25 patients). There was isonitrogenous and isocaloric intake in both groups, glutamine and B group patents were supplemented with glutamine granules or placebo (glycine) at 0.5 g/kg per day for 14 days with oral feeding or tube feeding, respectively. The level of plasma glutamine, plasma protein content, urine nitrogen and urine 3-methylhistidine (3-MTH) excretion were determined, wound healing rate of the burned area and hospital stay were recorded. The results showed that there were significant reductions in plasma glutamine level and abnormal protein metabolism. After supplement with glutamine granules for 14 days, the plasma glutamine concentration was significantly higher than that in B group (607.86+/-147.25 micromol/L versus 447.63+/-132.38 micromol/L, P0.05). On the other hand, the amount of urine nitrogen and 3-MTH excreted in Gln group were significantly lower than that in B group. In addition, wound healing was faster and hospital stay days were shorter in Gln group than B group (46.59+/-12.98 days versus 55.68+/-17.36 days, P<0.05). These indicated that supplement glutamine granules with oral feeding or tube feeding could abate the degree of glutamine depletion

  13. A new fluorescence-based method identifies protein phosphatases regulating lipid droplet metabolism.

    Directory of Open Access Journals (Sweden)

    Bruno L Bozaquel-Morais

    Full Text Available In virtually every cell, neutral lipids are stored in cytoplasmic structures called lipid droplets (LDs and also referred to as lipid bodies or lipid particles. We developed a rapid high-throughput assay based on the recovery of quenched BODIPY-fluorescence that allows to quantify lipid droplets. The method was validated by monitoring lipid droplet turnover during growth of a yeast culture and by screening a group of strains deleted in genes known to be involved in lipid metabolism. In both tests, the fluorimetric assay showed high sensitivity and good agreement with previously reported data using microscopy. We used this method for high-throughput identification of protein phosphatases involved in lipid droplet metabolism. From 65 yeast knockout strains encoding protein phosphatases and its regulatory subunits, 13 strains revealed to have abnormal levels of lipid droplets, 10 of them having high lipid droplet content. Strains deleted for type I protein phosphatases and related regulators (ppz2, gac1, bni4, type 2A phosphatase and its related regulator (pph21 and sap185, type 2C protein phosphatases (ptc1, ptc4, ptc7 and dual phosphatases (pps1, msg5 were catalogued as high-lipid droplet content strains. Only reg1, a targeting subunit of the type 1 phosphatase Glc7p, and members of the nutrient-sensitive TOR pathway (sit4 and the regulatory subunit sap190 were catalogued as low-lipid droplet content strains, which were studied further. We show that Snf1, the homologue of the mammalian AMP-activated kinase, is constitutively phosphorylated (hyperactive in sit4 and sap190 strains leading to a reduction of acetyl-CoA carboxylase activity. In conclusion, our fast and highly sensitive method permitted us to catalogue protein phosphatases involved in the regulation of LD metabolism and present evidence indicating that the TOR pathway and the SNF1/AMPK pathway are connected through the Sit4p-Sap190p pair in the control of lipid droplet biogenesis.

  14. Effects of Dietary Protein Source and Quantity during Weight Loss on Appetite, Energy Expenditure, and Cardio-Metabolic Responses.

    Science.gov (United States)

    Li, Jia; Armstrong, Cheryl L H; Campbell, Wayne W

    2016-01-26

    Higher protein meals increase satiety and the thermic effect of feeding (TEF) in acute settings, but it is unclear whether these effects remain after a person becomes acclimated to energy restriction or a given protein intake. This study assessed the effects of predominant protein source (omnivorous, beef/pork vs. lacto-ovo vegetarian, soy/legume) and quantity (10%, 20%, or 30% of energy from protein) on appetite, energy expenditure, and cardio-metabolic indices during energy restriction (ER) in overweight and obese adults. Subjects were randomly assigned to one protein source and then consumed diets with different quantities of protein (4 weeks each) in a randomized crossover manner. Perceived appetite ratings (free-living and in-lab), TEF, and fasting cardio-metabolic indices were assessed at the end of each 4-week period. Protein source and quantity did not affect TEF, hunger, or desire to eat, other than a modestly higher daily composite fullness rating with 30% vs. 10% protein diet (p = 0.03). While the 20% and 30% protein diets reduced cholesterol, triacylglycerol, and APO-B vs. 10% protein (p appetite control, energy expenditure and cardio-metabolic risk factors during ER-induced weight loss.

  15. Effect of beta-hydroxy-beta-methylbutyrate (HMB) on protein metabolism in whole body and in selected tissues.

    Science.gov (United States)

    Holecek, M; Muthny, T; Kovarik, M; Sispera, L

    2009-01-01

    Beta-hydroxy-beta-methylbutyrate (HMB) is a leucine metabolite with protein anabolic effect. The aim of the study was to examine the role of exogenous HMB on leucine and protein metabolism in whole body and selected tissues. Rats were administered by HMB (0.1 g/kg b.w.) or by saline. The parameters of whole-body protein metabolism were evaluated 24 h later using L-[1-14C]leucine and L-[3,4,5-3H]phenylalanine. Changes in proteasome dependent proteolysis and protein synthesis were determined according the "chymotrypsin-like" enzyme activity and labeled leucine and phenylalanine incorporation into the protein. A decrease in leucine clearance and whole-body protein turnover (i.e., a decrease in whole-body proteolysis and protein synthesis) was observed in HMB treated rats. Proteasome-dependent proteolysis decreased significantly in skeletal muscle, changes in heart, liver, jejunum, colon, kidney, and spleen were insignificant. Decrease in protein synthesis was observed in the heart, colon, kidney, and spleen, while an increase was observed in the liver. There were no significant changes in leucine oxidation. We conclude that protein anabolic effect of HMB in skeletal muscle is related to inhibition of proteolysis in proteasome. Alterations in protein synthesis in visceral tissues may affect several important functions and the metabolic status of the whole body.

  16. Core Data of Yeast Interacting Proteins Database (Original Version) - Yeast Interacting Proteins Database | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available y are in the reverse direction. *1 A comprehensive two-hybrid analysis to explore the yeast protein interact...s. 2000 Jan 1;28(1):73-6. *2 The yeast proteome database (YPD) and Caenorhabditis elegans proteome database (WormPD): comprehensive...000 Jan 1;28(1):73-6. *3 A comprehensive analysis of protein-protein interactions in Saccharomyces cerevisia

  17. Energy requirements, protein-energy metabolism and balance, and carbohydrates in preterm infants.

    Science.gov (United States)

    Hay, William W; Brown, Laura D; Denne, Scott C

    2014-01-01

    Energy is necessary for all vital functions of the body at molecular, cellular, organ, and systemic levels. Preterm infants have minimum energy requirements for basal metabolism and growth, but also have requirements for unique physiology and metabolism that influence energy expenditure. These include body size, postnatal age, physical activity, dietary intake, environmental temperatures, energy losses in the stool and urine, and clinical conditions and diseases, as well as changes in body composition. Both energy and protein are necessary to produce normal rates of growth. Carbohydrates (primarily glucose) are principle sources of energy for the brain and heart until lipid oxidation develops over several days to weeks after birth. A higher protein/energy ratio is necessary in most preterm infants to approximate normal intrauterine growth rates. Lean tissue is predominantly produced during early gestation, which continues through to term. During later gestation, fat accretion in adipose tissue adds increasingly large caloric requirements to the lean tissue growth. Once protein intake is sufficient to promote net lean body accretion, additional energy primarily produces more body fat, which increases almost linearly at energy intakes >80-90 kcal/kg/day in normal, healthy preterm infants. Rapid gains in adiposity have the potential to produce later life obesity, an increasingly recognized risk of excessive energy intake. In addition to fundamental requirements for glucose, protein, and fat, a variety of non-glucose carbohydrates found in human milk may have important roles in promoting growth and development, as well as production of a gut microbiome that could protect against necrotizing enterocolitis. © 2014 S. Karger AG, Basel.

  18. A review of some metabolic changes in protein-energy malnutrition.

    Science.gov (United States)

    Akuyam, S A

    2007-06-01

    Protein-energy malnutrition (PEM) is a major public health problem in the tropical and subtropical regions of the world and often arises during protein and / or energy deficit due to nutritional inadequacy, infections and poor socio-economic and environmental conditions. It is the most common nutritional disorder affecting children in developing countries and the third most common disease of childhood in such countries. PEM has a lasting effect on immune functions, growth and development of children, learning ability, social adjustment, work efficiency and productivity of labour. It seems that many deaths from PEM occur as a result of outdated clinical practices and that improving these practices reduces the rate of morbidity and mortality. This paper reviews various metabolic changes in protein-energy malnutrition (PEM). It gives an overview of the theoretical basis for the understanding of the biochemical derangements in PEM. It aims at stimulating the paediatricians and clinical chemists to read more on the recent advances in this broad subject with the view to improving the understanding of the current laboratory investigation of PEM. This review demonstrates that the metabolic changes in PEM include water and electrolytes imbalance, amino acids and proteins deficiencies, carbohydrates and energy deficiencies, hypolipidaemias, hypolipoproteinaemias, hormonal imbalance, deficiency of anti-oxidant vitamins and enzymes, depression of cell-mediated immune complexes and decrease in amino acids and trace elements in skin and hair. The review therefore suggests that assessment of these conditions in PEM patients could improve the management of this group of patients and hence reduce the rate of morbidity and mortality from PEM.

  19. Cocoa and Whey Protein Differentially Affect Markers of Lipid and Glucose Metabolism and Satiety.

    Science.gov (United States)

    Campbell, Caroline L; Foegeding, E Allen; Harris, G Keith

    2016-03-01

    Food formulation with bioactive ingredients is a potential strategy to promote satiety and weight management. Whey proteins are high in leucine and are shown to decrease hunger ratings and increase satiety hormone levels; cocoa polyphenolics moderate glucose levels and slow digestion. This study examined the effects of cocoa and whey proteins on lipid and glucose metabolism and satiety in vitro and in a clinical trial. In vitro, 3T3-L1 preadipocytes were treated with 0.5-100 μg/mL cocoa polyphenolic extract (CPE) and/or 1-15 mM leucine (Leu) and assayed for lipid accumulation and leptin production. In vivo, a 6-week clinical trial consisted of nine panelists (age: 22.6 ± 1.7; BMI: 22.3 ± 2.1) consuming chocolate-protein beverages once per week, including placebo, whey protein isolate (WPI), low polyphenolic cocoa (LP), high polyphenolic cocoa (HP), LP-WPI, and HP-WPI. Measurements included blood glucose and adiponectin levels, and hunger ratings at baseline and 0.5-4.0 h following beverage consumption. At levels of 50 and 100 μg/mL, CPE significantly inhibited preadipocyte lipid accumulation by 35% and 50%, respectively, and by 22% and 36% when combined with 15 mM Leu. Leu treatment increased adipocyte leptin production by 26-37%. In the clinical trial, all beverages significantly moderated blood glucose levels 30 min postconsumption. WPI beverages elicited lowest peak glucose levels and HP levels were significantly lower than LP. The WPI and HP beverage treatments significantly increased adiponectin levels, but elicited no significant changes in hunger ratings. These trends suggest that combinations of WPI and cocoa polyphenols may improve markers of metabolic syndrome and satiety.

  20. Dissociation of the effects of epinephrine and insulin on glucose and protein metabolism

    International Nuclear Information System (INIS)

    Castellino, P.; Luzi, L.; Del Prato, S.; DeFronzo, R.A.

    1990-01-01

    The separate and combined effects of insulin and epinephrine on leucine metabolism were examined in healthy young volunteers. Subjects participated in four experimental protocols: (1) euglycemic insulin clamp (+80 microU/ml), (2) epinephrine infusion (50 ng.kg-1.min-1) plus somatostatin with basal replacement of insulin and glucagon, (3) combined epinephrine (50 ng.kg-1.min-1) plus insulin (+80 microU/ml) infusion, and (4) epinephrine and somatostatin as in study 2 plus basal amino acid replacement. Studies were performed with a prime-continuous infusion of [1-14C]leucine and indirect calorimetry. Our results indicate that (1) hyperinsulinemia causes a generalized decrease in plasma amino acid concentrations, including leucine; (2) the reduction in plasma leucine concentration is primarily due to an inhibition of endogenous leucine flux; nonoxidative leucine disposal decreases after insulin infusion; (3) epinephrine, without change in plasma insulin concentration, reduces plasma amino acid levels; (4) combined epinephrine-insulin infusion causes a greater decrease in plasma amino levels than observed with either hormone alone; this is because of a greater inhibition of endogenous leucine flux; and (5) when basal amino acid concentrations are maintained constant with a balanced amino acid infusion, epinephrine inhibits the endogenous leucine flux. In conclusion, the present results do not provide support for the concept that epinephrine is a catabolic hormone with respect to amino acid-protein metabolism. In contrast, epinephrine markedly inhibits insulin-mediated glucose metabolism

  1. Role of the Irr protein in the regulation of iron metabolism in Rhodobacter sphaeroides.

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    Verena Peuser

    Full Text Available In Rhizobia the Irr protein is an important regulator for iron-dependent gene expression. We studied the role of the Irr homolog RSP_3179 in the photosynthetic alpha-proteobacterium Rhodobacter sphaeroides. While Irr had little effect on growth under iron-limiting or non-limiting conditions its deletion resulted in increased resistance to hydrogen peroxide and singlet oxygen. This correlates with an elevated expression of katE for catalase in the Irr mutant compared to the wild type under non-stress conditions. Transcriptome studies revealed that Irr affects the expression of genes for iron metabolism, but also has some influence on genes involved in stress response, citric acid cycle, oxidative phosphorylation, transport, and photosynthesis. Most genes showed higher expression levels in the wild type than in the mutant under normal growth conditions indicating an activator function of Irr. Irr was however not required to activate genes of the iron metabolism in response to iron limitation, which showed even stronger induction in the absence of Irr. This was also true for genes mbfA and ccpA, which were verified as direct targets for Irr. Our results suggest that in R. sphaeroides Irr diminishes the strong induction of genes for iron metabolism under iron starvation.

  2. Heterogeneity of elderly depression: increased risk of Alzheimer's disease and Aβ protein metabolism.

    Science.gov (United States)

    Namekawa, Yuki; Baba, Hajime; Maeshima, Hitoshi; Nakano, Yoshiyuki; Satomura, Emi; Takebayashi, Naoko; Nomoto, Hiroshi; Suzuki, Toshihito; Arai, Heii

    2013-06-03

    Epidemiological studies have proposed that depression may increase the risk for Alzheimer's disease (AD), even in patients with early-onset depression. Although metabolism of amyloid β protein (Aβ) in elderly depression received attention in terms of their correlation, there is a serious heterogeneity in elderly depression in terms of age at onset of depression. Moreover, it is unknown whether early-onset major depressive disorder (MDD) has a long-term effect on the involvement of Aβ metabolism and later development of AD. Thus, we evaluated serum Aβ40 and Aβ42 levels, the Aβ40/Aβ42 ratio in 89 elderly (≥60 years of age) inpatients with MDD and 81 age-matched healthy controls, and compared them among patients with early-onset (great interest that the serum Aβ40/Aβ42 ratio was negatively correlated with the age at MDD onset (R=-0.201, p=0.032). These results suggest that an earlier onset of MDD may have a more serious abnormality in Aβ metabolism, possibly explaining a biological mechanism underlying the link between depression and AD. Copyright © 2012 Elsevier Inc. All rights reserved.

  3. Impairment of interferon regulatory factor-3 activation by hepatitis C virus core protein basic amino acid region 1.

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    Inoue, Kazuaki; Tsukiyama-Kohara, Kyoko; Matsuda, Chiho; Yoneyama, Mitsutoshi; Fujita, Takashi; Kuge, Shusuke; Yoshiba, Makoto; Kohara, Michinori

    2012-11-30

    Interferon regulatory factor-3 (IRF-3), a key transcriptional factor in the type I interferon system, is frequently impaired by hepatitis C virus (HCV), in order to establish persistent infection. However, the exact mechanism by which the virus establishes persistent infection has not been fully understood yet. The present study aimed to investigate the effects of various HCV proteins on IRF-3 activation, and elucidate the underlying mechanisms. To achieve this, full-length HCV and HCV subgenomic constructs corresponding to structural and each of the nonstructural proteins were transiently transfected into HepG2 cells. IFN-β induction, plaque formation, and IRF-3 dimerization were elicited by Newcastle disease virus (NDV) infection. The expressions of IRF-3 homodimer and its monomer, Ser386-phosphorylated IRF-3, and HCV core protein were detected by immunofluorescence and western blotting. IFN-β mRNA expression was quantified by real-time PCR (RT-PCR), and IRF-3 activity was measured by the levels of IRF-3 dimerization and phosphorylation, induced by NDV infection or polyriboinosinic:polyribocytidylic acid [poly(I:C)]. Switching of the expression of the complete HCV genome as well as the core proteins, E1, E2, and NS2, suppressed IFN-β mRNA levels and IRF-3 dimerization, induced by NDV infection. Our study revealed a crucial region of the HCV core protein, basic amino acid region 1 (BR1), to inhibit IRF-3 dimerization as well as its phosphorylation induced by NDV infection and poly (I:C), thus interfering with IRF-3 activation. Therefore, our study suggests that rescue of the IRF-3 pathway impairment may be an effective treatment for HCV infection. Copyright © 2012 Elsevier Inc. All rights reserved.

  4. Rotavirus NSP2 interferes with the core lattice protein VP2 in initiation of minus-strand synthesis

    International Nuclear Information System (INIS)

    Vende, Patrice; Tortorici, M. Alejandra; Taraporewala, Zenobia F.; Patton, John T.

    2003-01-01

    The rotavirus nonstructural protein NSP2 self-assembles into stable octameric structures that possess nonspecific affinity for single-stranded (ss)RNA and RNA-RNA helix-destabilizing and NTPase activities. Furthermore, NSP2 is a component of replication intermediates with replicase activity and plays a critical role in the packaging and replication of the segmented dsRNA genome of rotavirus. To better understand the function of the protein in genome replication, we examined the effect that purified recombinant NSP2 had on the synthesis of dsRNA by the open core replication system. The results showed that NSP2 inhibited the synthesis of dsRNA from viral mRNA in vitro, in a concentration-dependent manner. The inhibition was overcome by adding increasing amounts of viral mRNA or nonviral ssRNA to the system, indicating that the inhibition was mediated by the nonspecific RNA-binding activity of NSP2. Further analysis revealed that NSP2 interfered with the ability of the open core proteins, GTP, and viral mRNA to form the initiation complex for (-) strand synthesis. Additional experiments indicated that NSP2 did not perturb recognition of viral mRNA by the viral RNA polymerase VP1, but rather interfered with the function of VP2, a protein that is essential for (-) strand initiation and dsRNA synthesis and that forms the T = 1 lattice of the virion core. In contrast to initiation, NSP2 did not inhibit (-) strand elongation. Collectively, the findings provide evidence that the temporal order of interaction of RNA-binding proteins with viral mRNA is a crucial factor impacting the formation of replication intermediates

  5. A Systematic Review of the Effects of Plant Compared with Animal Protein Sources on Features of Metabolic Syndrome.

    Science.gov (United States)

    Chalvon-Demersay, Tristan; Azzout-Marniche, Dalila; Arfsten, Judith; Egli, Léonie; Gaudichon, Claire; Karagounis, Leonidas G; Tomé, Daniel

    2017-03-01

    Dietary protein may play an important role in the prevention of metabolic dysfunctions. However, the way in which the protein source affects these dysfunctions has not been clearly established. The aim of the current systematic review was to compare the impact of plant- and animal-sourced dietary proteins on several features of metabolic syndrome in humans. The PubMed database was searched for both chronic and acute interventional studies, as well as observational studies, in healthy humans or those with metabolic dysfunctions, in which the impact of animal and plant protein intake was compared while using the following variables: cholesterolemia and triglyceridemia, blood pressure, glucose homeostasis, and body composition. Based on data extraction, we observed that soy protein consumption (with isoflavones), but not soy protein alone (without isoflavones) or other plant proteins (pea and lupine proteins, wheat gluten), leads to a 3% greater decrease in both total and LDL cholesterol compared with animal-sourced protein ingestion, especially in individuals with high fasting cholesterol concentrations. This observation was made when animal proteins were provided as a whole diet rather than given supplementally. Some observational studies reported an inverse association between plant protein intake and systolic and diastolic blood pressure, but this was not confirmed by intervention studies. Moreover, plant protein (wheat gluten, soy protein) intake as part of a mixed meal resulted in a lower postprandial insulin response than did whey. This systematic review provides some evidence that the intake of soy protein associated with isoflavones may prevent the onset of risk factors associated with cardiovascular disease, i.e., hypercholesterolemia and hypertension, in humans. However, we were not able to draw any further conclusions from the present work on the positive effects of plant proteins relating to glucose homeostasis and body composition. © 2017 American

  6. Synthetic protein scaffolds based on peptide motifs and cognate adaptor domains for improving metabolic productivity

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    Anselm H.C. Horn

    2015-11-01

    Full Text Available The efficiency of many cellular processes relies on the defined interaction among different proteins within the same metabolic or signaling pathway. Consequently, a spatial colocalization of functionally interacting proteins has frequently emerged during evolution. This concept has been adapted within the synthetic biology community for the purpose of creating artificial scaffolds. A recent advancement of this concept is the use of peptide motifs and their cognate adaptor domains. SH2, SH3, GBD, and PDZ domains have been used most often in research studies to date. The approach has been successfully applied to the synthesis of a variety of target molecules including catechin, D-glucaric acid, H2, hydrochinone, resveratrol, butyrate, gamma-aminobutyric acid, and mevalonate. Increased production levels of up to 77-fold have been observed compared to non-scaffolded systems. A recent extension of this concept is the creation of a covalent linkage between peptide motifs and adaptor domains, which leads to a more stable association of the scaffolded systems and thus bears the potential to further enhance metabolic productivity.

  7. Lack of the Lysosomal Membrane Protein, GLMP, in Mice Results in Metabolic Dysregulation in Liver.

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    Xiang Yi Kong

    Full Text Available Ablation of glycosylated lysosomal membrane protein (GLMP, formerly known as NCU-G1 has been shown to cause chronic liver injury which progresses into liver fibrosis in mice. Both lysosomal dysfunction and chronic liver injury can cause metabolic dysregulation. Glmp gt/gt mice (formerly known as Ncu-g1gt/gt mice were studied between 3 weeks and 9 months of age. Body weight gain and feed efficiency of Glmp gt/gt mice were comparable to wild type siblings, only at the age of 9 months the Glmp gt/gt siblings had significantly reduced body weight. Reduced size of epididymal fat pads was accompanied by hepatosplenomegaly in Glmp gt/gt mice. Blood analysis revealed reduced levels of blood glucose, circulating triacylglycerol and non-esterified fatty acids in Glmp gt/gt mice. Increased flux of glucose, increased de novo lipogenesis and lipid accumulation were detected in Glmp gt/gt primary hepatocytes, as well as elevated triacylglycerol levels in Glmp gt/gt liver homogenates, compared to hepatocytes and liver from wild type mice. Gene expression analysis showed an increased expression of genes involved in fatty acid uptake and lipogenesis in Glmp gt/gt liver compared to wild type. Our findings are in agreement with the metabolic alterations observed in other mouse models lacking lysosomal proteins, and with alterations characteristic for advanced chronic liver injury.

  8. Impaired mitochondrial metabolism and protein synthesis in streptozotocin diabetic rat hepatocytes

    International Nuclear Information System (INIS)

    Memon, R.A.; Bessman, S.P.; Mohan, C.

    1990-01-01

    Isolated hepatocytes prepared from control, streptozotocin diabetic rats were incubated at 30 degrees C in Krebs-Henseleit bicarbonate buffer, pH 7.4, containing 0.5 mM concentration of each of the 20 natural amino acids. Effect of insulin on the oxidation of 2,3- 14 C and 1,4- 14 C succinate (suc) carbons and their incorporation into hepatocyte protein, lipid and various metabolic intermediates was studied. Mitochondrial oxidation of suc carbons and their incorporation into protein and lipid was significantly lower in diabetic and insulin treated diabetic rats. Diabetic rats failed to exhibit any significant insulin effect on the oxidation of either 2,3 or 1,4- 14 C suc carbons. Amphibolic channeling of 2,3- 14 C suc carbons into amino acids was significantly reduced in hepatocytes of diabetic rats, however, more of these carbons were diverted into the gluconeogenesis pathway. Diabetes caused a far greater decrease in the oxidation of 2,3- 14 C suc carbons as compared to 1,4- 14 C suc. Based on an earlier report that insulin stimulates only the intramitochondrial Krebs cycle reactions, the authors conclude that the diminished level of anabolic activities in the diabetic rat hepatocytes is due to the subsequent reduction in amphibolic channeling of metabolic intermediates

  9. A Protein Scaffold Coordinates SRC-Mediated JNK Activation in Response to Metabolic Stress.

    Science.gov (United States)

    Kant, Shashi; Standen, Claire L; Morel, Caroline; Jung, Dae Young; Kim, Jason K; Swat, Wojciech; Flavell, Richard A; Davis, Roger J

    2017-09-19

    Obesity is a major risk factor for the development of metabolic syndrome and type 2 diabetes. How obesity contributes to metabolic syndrome is unclear. Free fatty acid (FFA) activation of a non-receptor tyrosine kinase (SRC)-dependent cJun NH 2 -terminal kinase (JNK) signaling pathway is implicated in this process. However, the mechanism that mediates SRC-dependent JNK activation is unclear. Here, we identify a role for the scaffold protein JIP1 in SRC-dependent JNK activation. SRC phosphorylation of JIP1 creates phosphotyrosine interaction motifs that bind the SH2 domains of SRC and the guanine nucleotide exchange factor VAV. These interactions are required for SRC-induced activation of VAV and the subsequent engagement of a JIP1-tethered JNK signaling module. The JIP1 scaffold protein, therefore, plays a dual role in FFA signaling by coordinating upstream SRC functions together with downstream effector signaling by the JNK pathway. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  10. Energy dense, protein restricted diet increases adiposity and perturbs metabolism in young, genetically lean pigs.

    Science.gov (United States)

    Fisher, Kimberly D; Scheffler, Tracy L; Kasten, Steven C; Reinholt, Brad M; van Eyk, Gregory R; Escobar, Jeffery; Scheffler, Jason M; Gerrard, David E

    2013-01-01

    Animal models of obesity and metabolic dysregulation during growth (or childhood) are lacking. Our objective was to increase adiposity and induce metabolic syndrome in young, genetically lean pigs. Pre-pubertal female pigs, age 35 d, were fed a high-energy diet (HED; n = 12), containing 15% tallow, 35% refined sugars and 9.1-12.9% crude protein, or a control corn-based diet (n = 11) with 12.2-19.2% crude protein for 16 wk. Initially, HED pigs self-regulated energy intake similar to controls, but by wk 5, consumed more (Pblood glucose increased (Pblood glucose did not return to baseline (P = 0.01), even 4 h post-challenge. During OGTT, glucose area under the curve (AUC) was higher and insulin AUC was lower in HED pigs compared to controls (P = 0.001). Chronic HED intake increased (PAUC and insulin AUC did not improve, supporting that dietary intervention was not sufficient to recover glucose tolerance or insulin production. These data suggest a HED may be used to increase adiposity and disrupt glucose homeostasis in young, growing pigs.

  11. A Protein Scaffold Coordinates SRC-Mediated JNK Activation in Response to Metabolic Stress

    Directory of Open Access Journals (Sweden)

    Shashi Kant

    2017-09-01

    Full Text Available Obesity is a major risk factor for the development of metabolic syndrome and type 2 diabetes. How obesity contributes to metabolic syndrome is unclear. Free fatty acid (FFA activation of a non-receptor tyrosine kinase (SRC-dependent cJun NH2-terminal kinase (JNK signaling pathway is implicated in this process. However, the mechanism that mediates SRC-dependent JNK activation is unclear. Here, we identify a role for the scaffold protein JIP1 in SRC-dependent JNK activation. SRC phosphorylation of JIP1 creates phosphotyrosine interaction motifs that bind the SH2 domains of SRC and the guanine nucleotide exchange factor VAV. These interactions are required for SRC-induced activation of VAV and the subsequent engagement of a JIP1-tethered JNK signaling module. The JIP1 scaffold protein, therefore, plays a dual role in FFA signaling by coordinating upstream SRC functions together with downstream effector signaling by the JNK pathway.

  12. Osteocalcin: The extra-skeletal role of a vitamin K-dependent protein in glucose metabolism

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    Eibhlís M. O'Connor

    2017-03-01

    Full Text Available The role of vitamin K in the body has long been associated with blood clotting and coagulation. In more recent times, its role in a range of physiological processes has been described including the regulation of bone and soft tissue calcification, cell growth and proliferation, cognition, inflammation, various oxidative processes and fertility, where osteocalcin is thought to up-regulate the synthesis of the enzymes needed for the biosynthesis of testosterone thereby increasing male fertility. Vitamin K dependent proteins (VKDP contain γ-carboxyglutamic acid residues which require post-translational, gamma-glutamyl carboxylation by the vitamin K-dependent (VKD gamma-glutamyl carboxylase enzyme for full functionality. These proteins are present both hepatically and extrahepatically. The role of bone-derived osteocalcin has many physiological roles including, maintenance of bone mass with more recent links to energy metabolism due to the role of the skeleton as an endocrine organ. It has been proposed that insulin binds to bone forming cells (osteoblasts promoting osteocalcin production which in turn promotes β-cell proliferation, insulin secretion and glucose control. However much of this research has been conducted in animal models with equivocal findings in human studies. This review will discuss the role of osteocalcin in relation to its role in human health, focusing specifically on glucose metabolism.

  13. The choice of label and measurement technique in tracer studies of body protein metabolism in man

    International Nuclear Information System (INIS)

    James, W.P.T.; Sender, P.M.; Garlick, P.J.; Waterlow, J.C.

    1975-01-01

    The turnover of non-serum proteins in man has had limited study despite the physiological importance of maintaining the balance between synthesis and breakdown of body proteins. Body protein is usually considered as a single pool and breakdown rates are often measured by monitoring excreted label at intervals after pulse labelling with radioactive or 15 N amino acids. No label has yet been used for measuring tissue protein breakdown in man which is free from the major problem of label re-utilization. All measurements of breakdown rates, eg. with 75 Se-selenomethionine, 15 N- or 14 C-glycine, give rate constants which are too low. The heterogeneity of body proteins also means that an estimate of the weighted average breakdown rate can only be obtained after following the excretion of isotope for a long period, perhaps of the order of 3-4 half-lives which, for man, would be 100 days after labelling. We therefore use infusions with either 14 C- or 15 N-labelled amino acids to measure breakdown and synthesis rates: these values are less affected by problems of protein heterogeneity. Single injection techniques are subject to more error than constant infusions of label because of the difficulty of defining the precursor activity. 15 N labelling need not be confined to essential amino acids if total protein rather than amino acid turnover is studied: the latter involves measurements of the labelled amino acid itself which is difficult with 15 N because of the small amounts of free amino acid nitrogen available. Carbon labelling of non-essential amino acids is unsuitable for studies of protein turnover and the choice of the position of the label on the molecule is important when labelled essential amino acids are employed. Short-term changes in protein metabolism are evaluated better with amino acids with a small pool size; the equilibration time in the excretory bicarbonate pool is also shorter than in the urea pool so that 15 N is less useful than carbon labelling. We

  14. Protein O-linked ß-N-acetylglucosamine: A novel effector of cardiomyocyte metabolism and function

    Science.gov (United States)

    Darley-Usmar, Victor M.; Ball, Lauren E.; Chatham, John C.

    2014-01-01

    The post-translational modification of serine and threonine residues of nuclear and cytoplasmic proteins by the O-linked attachment of the monosaccharide ß-N-acetyl-glucosamine (O-GlcNAc) is emerging as an important mechanism for the regulation of numerous biological processes critical for normal cell function. Active synthesis of O-GlcNAc is essential for cell viability and acute activation of pathways resulting in increased protein O-GlcNAc levels improves the tolerance of cells to a wide range of stress stimuli. Conversely sustained increases in O-GlcNAc levels have been implicated in numerous chronic disease states, especially as a pathogenic contributor to diabetic complications. There has been increasing interest in the role of O-GlcNAc in the heart and vascular system and acute activation of O-GlcNAc levels have been shown to reduce ischemia/reperfusion injury attenuate vascular injury responses as well mediate some of the detrimental effects of diabetes and hypertension on cardiac and vascular function. Here we provide an overview of our current understanding of pathways regulating protein O-GlcNAcylation, summarize the different methodologies for identifying and characterizing O-GlcNAcylated proteins and subsequently focus on two emerging areas: 1) the role of O-GlcNAc as a potential regulator of cardiac metabolism and 2) the cross talk between O-GlcNAc and reactive oxygen species. PMID:21878340

  15. Protein kinase N2 regulates AMP kinase signaling and insulin responsiveness of glucose metabolism in skeletal muscle.

    Science.gov (United States)

    Ruby, Maxwell A; Riedl, Isabelle; Massart, Julie; Åhlin, Marcus; Zierath, Juleen R

    2017-10-01

    Insulin resistance is central to the development of type 2 diabetes and related metabolic disorders. Because skeletal muscle is responsible for the majority of whole body insulin-stimulated glucose uptake, regulation of glucose metabolism in this tissue is of particular importance. Although Rho GTPases and many of their affecters influence skeletal muscle metabolism, there is a paucity of information on the protein kinase N (PKN) family of serine/threonine protein kinases. We investigated the impact of PKN2 on insulin signaling and glucose metabolism in primary human skeletal muscle cells in vitro and mouse tibialis anterior muscle in vivo. PKN2 knockdown in vitro decreased insulin-stimulated glucose uptake, incorporation into glycogen, and oxidation. PKN2 siRNA increased 5'-adenosine monophosphate-activated protein kinase (AMPK) signaling while stimulating fatty acid oxidation and incorporation into triglycerides and decreasing protein synthesis. At the transcriptional level, PKN2 knockdown increased expression of PGC-1α and SREBP-1c and their target genes. In mature skeletal muscle, in vivo PKN2 knockdown decreased glucose uptake and increased AMPK phosphorylation. Thus, PKN2 alters key signaling pathways and transcriptional networks to regulate glucose and lipid metabolism. Identification of PKN2 as a novel regulator of insulin and AMPK signaling may provide an avenue for manipulation of skeletal muscle metabolism. Copyright © 2017 the American Physiological Society.

  16. Protein-energy malnutrition induces an aberrant acute-phase response and modifies the circadian rhythm of core temperature.

    Science.gov (United States)

    Smith, Shari E; Ramos, Rafaela Andrade; Refinetti, Roberto; Farthing, Jonathan P; Paterson, Phyllis G

    2013-08-01

    Protein-energy malnutrition (PEM), present in 12%-19% of stroke patients upon hospital admission, appears to be a detrimental comorbidity factor that impairs functional outcome, but the mechanisms are not fully elucidated. Because ischemic brain injury is highly temperature-sensitive, the objectives of this study were to investigate whether PEM causes sustained changes in temperature that are associated with an inflammatory response. Activity levels were recorded as a possible explanation for the immediate elevation in temperature upon introduction to a low protein diet. Male, Sprague-Dawley rats (7 weeks old) were fed a control diet (18% protein) or a low protein diet (PEM, 2% protein) for either 7 or 28 days. Continuous core temperature recordings from bioelectrical sensor transmitters demonstrated a rapid increase in temperature amplitude, sustained over 28 days, in response to a low protein diet. Daily mean temperature rose transiently by day 2 (p = 0.01), falling to normal by day 4 (p = 0.08), after which mean temperature continually declined as malnutrition progressed. There were no alterations in activity mean (p = 0.3) or amplitude (p = 0.2) that were associated with the early rise in mean temperature. Increased serum alpha-2-macroglobulin (p protein diet had no effect on the signaling pathway of the pro-inflammatory transcription factor, NFκB, in the hippocampus. In conclusion, PEM induces an aberrant and sustained acute-phase response coupled with long-lasting effects on body temperature.

  17. Expression of Hepatitis C Virus Core and E2 antigenic recombinant proteins and their use for development of diagnostic assays.

    Science.gov (United States)

    Ali, Amjad; Nisar, Muhammad; Idrees, Muhammad; Rafique, Shazia; Iqbal, Muhammad

    2015-05-01

    Early diagnosis of HCV infection is based on detection of antibodies against HCV proteins using recombinant viral antigens. The present study was designed to select, clone and express the antigenic regions of Core and E2 genes from local HCV-3a genotype and to utilize the antigenic recombinant proteins (Core & E2) to develop highly sensitive, specific and economical diagnostic assays for detection of HCV infection. The antigenic sites were determined within Core and E2 genes and were then cloned in pET-28a expression vector. The right orientation of the desired inserted fragments of Core and E2 were confirmed via sequencing prior to expression and were then transformed in BL21 (DE3) pLysS strains of E. coli and induced with 0.5mM Isopropyl-b-D-thiogalactopyranoside (IPTG) for the production of antigenic recombinant proteins. The produced truncated antigens were then purified by Nickel affinity chromatography and were confirmed by western blotting, immunoblotting and enzyme-linked immunosorbent assay (ELISA). The expressed Core and E2 recombinant antigens were used to develop immunoblotting assay for the detection of anti-HCV antibodies in sera. With immunoblotting, a total of 93-HCV infected sera and 35-HCV negative individuals were tested for the presence of anti-HCV antibodies to the Core and E2 antigens. Recombinant antigen showed 100% reactivity against HCV infected sera, with no cross reactivity against HCV-negative sera. The immunoblot assay mixture of recombinant antigens (Core+E2) showed a strong reaction intensity in the test area (TA) as compared to the individual truncated Core and E2 recombinant antigens. In the in-house ELISA assay, mixed Core and E2 recombinant antigens showed 100% reactivity against a standardized panel of 150-HCV-positive sera and non reactivity against a standardized panel of 150 HCV-negative sera while also being non reactive to sera positive for other viral infections. The antigenic recombinant antigens also were tested for the

  18. The Replacement of 10 Non-Conserved Residues in the Core Protein of JFH-1 Hepatitis C Virus Improves Its Assembly and Secretion.

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    Loïc Etienne

    Full Text Available Hepatitis C virus (HCV assembly is still poorly understood. It is thought that trafficking of the HCV core protein to the lipid droplet (LD surface is essential for its multimerization and association with newly synthesized HCV RNA to form the viral nucleocapsid. We carried out a mapping analysis of several complete HCV genomes of all genotypes, and found that the genotype 2 JFH-1 core protein contained 10 residues different from those of other genotypes. The replacement of these 10 residues of the JFH-1 strain sequence with the most conserved residues deduced from sequence alignments greatly increased virus production. Confocal microscopy of the modified JFH-1 strain in cell culture showed that the mutated JFH-1 core protein, C10M, was present mostly at the endoplasmic reticulum (ER membrane, but not at the surface of the LDs, even though its trafficking to these organelles was possible. The non-structural 5A protein of HCV was also redirected to ER membranes and colocalized with the C10M core protein. Using a Semliki forest virus vector to overproduce core protein, we demonstrated that the C10M core protein was able to form HCV-like particles, unlike the native JFH-1 core protein. Thus, the substitution of a few selected residues in the JFH-1 core protein modified the subcellular distribution and assembly properties of the protein. These findings suggest that the early steps of HCV assembly occur at the ER membrane rather than at the LD surface. The C10M-JFH-1 strain will be a valuable tool for further studies of HCV morphogenesis.

  19. A diet containing whey protein, glutamine, and TGFbeta modulates gut protein metabolism during chemotherapy-induced mucositis in rats.

    Science.gov (United States)

    Boukhettala, Nabile; Ibrahim, Ayman; Claeyssens, Sophie; Faure, Magali; Le Pessot, Florence; Vuichoud, Jacques; Lavoinne, Alain; Breuillé, Denis; Déchelotte, Pierre; Coëffier, Moïse

    2010-08-01

    Mucositis, a common side effect of chemotherapy, is characterized by compromised digestive function, barrier integrity and immune competence. Our aim was to evaluate the impact of a specifically designed diet Clinutren Protect (CP), which contains whey proteins, TGFbeta-rich casein, and free glutamine, on mucositis in rats. Mucositis was induced by three consecutive injections (day 0, day 1, day 2) of methotrexate (2.5 mg/kg). Rats had free access to CP or placebo diets from days -7 to 9. In the placebo diet, whey proteins and TGFbeta-rich casein were replaced by TGFbeta-free casein and glutamine by alanine. Intestinal parameters were assessed at day 3 and 9. Values, expressed as mean +/- SEM, were compared using two-way ANOVA. At day 3, villus height was markedly decreased in the placebo (296 +/- 11 microm) and CP groups (360 +/- 10 microm) compared with controls (464 +/- 27 microm), but more markedly in the placebo as compared to CP group. The intestinal damage score was also reduced in the CP compared with the placebo group. Glutathione content increased in the CP compared with the placebo group (2.2 +/- 0.2 vs. 1.7 +/- 0.2 micromol/g tissue). Gut protein metabolism was more affected in the placebo than in the CP group. The fractional synthesis rate was decreased in the placebo group (93.8 +/- 4.9%/day) compared with controls (121.5 +/- 12.1, P < 0.05), but not in the CP group (106.0 +/- 13.1). In addition, at day 9, rats exhibited improved body weight and food intake recovery in the CP compared to the placebo group. Clinutren Protect feeding reduces intestinal injury in the acute phase of methotrexate-induced mucositis in rats and improves recovery.

  20. Fluorescent protein-tagged Vpr dissociates from HIV-1 core after viral fusion and rapidly enters the cell nucleus.

    Science.gov (United States)

    Desai, Tanay M; Marin, Mariana; Sood, Chetan; Shi, Jiong; Nawaz, Fatima; Aiken, Christopher; Melikyan, Gregory B

    2015-10-29

    HIV-1 Vpr is recruited into virions during assembly and appears to remain associated with the viral core after the reverse transcription and uncoating steps of entry. This feature has prompted the use of fluorescently labeled Vpr to visualize viral particles and to follow trafficking of post-fusion HIV-1 cores in the cytoplasm. Here, we tracked single pseudovirus entry and fusion and observed that fluorescently tagged Vpr gradually dissociates from post-fusion viral cores over the course of several minutes and accumulates in the nucleus. Kinetics measurements showed that fluorescent Vpr released from the cores very rapidly entered the cell nucleus. More than 10,000 Vpr molecules can be delivered into the cell nucleus within 45 min of infection by HIV-1 particles pseudotyped with the avian sarcoma and leukosis virus envelope glycoprotein. The fraction of Vpr from cell-bound viruses that accumulated in the nucleus was proportional to the extent of virus-cell fusion and was fully blocked by viral fusion inhibitors. Entry of virus-derived Vpr into the nucleus occurred independently of envelope glycoproteins or target cells. Fluorescence correlation spectroscopy revealed two forms of nuclear Vpr-monomers and very large complexes, likely involving host factors. The kinetics of viral Vpr entering the nucleus after fusion was not affected by point mutations in the capsid protein that alter the stability of the viral core. The independence of Vpr shedding of capsid stability and its relatively rapid dissociation from post-fusion cores suggest that this process may precede capsid uncoating, which appears to occur on a slower time scale. Our results thus demonstrate that a bulk of fluorescently labeled Vpr incorporated into HIV-1 particles is released shortly after fusion. Future studies will address the question whether the quick and efficient nuclear delivery of Vpr derived from incoming viruses can regulate subsequent steps of HIV-1 infection.

  1. Purification, crystallization and preliminary X-ray diffraction study of human ribosomal protein L10 core domain

    International Nuclear Information System (INIS)

    Nishimura, Mitsuhiro; Kaminishi, Tatsuya; Kawazoe, Masahito; Shirouzu, Mikako; Takemoto, Chie; Yokoyama, Shigeyuki; Tanaka, Akiko; Sugano, Sumio; Yoshida, Takuya; Ohkubo, Tadayasu; Kobayashi, Yuji

    2007-01-01

    A truncated variant of human ribosomal protien L10 was prepared and crystallized. Diffraction data were collected to 2.5 Å resolution. Eukaryotic ribosomal protein L10 is an essential component of the large ribosomal subunit, which organizes the architecture of the aminoacyl-tRNA binding site. The human L10 protein is also called the QM protein and consists of 214 amino-acid residues. For crystallization, the L10 core domain (L10CD, Phe34–Glu182) was recombinantly expressed in Escherichia coli and purified to homogeneity. A hexagonal crystal of L10CD was obtained by the sitting-drop vapour-diffusion method. The L10CD crystal diffracted to 2.5 Å resolution and belongs to space group P3 1 21 or P3 2 21

  2. Association of cancer metabolism-related proteins with oral carcinogenesis – indications for chemoprevention and metabolic sensitizing of oral squamous cell carcinoma?

    Science.gov (United States)

    2014-01-01

    Background Tumor metabolism is a crucial factor for the carcinogenesis of oral squamous cell carcinoma (OSCC). Methods Expression of IGF-R1, glycolysis-related proteins (GLUT-1, HK 2, PFK-1, LDHA, TKTL1), mitochondrial enzymes (SDHA, SDHB, ATP synthase) were analyzed in normal oral mucosa (n = 5), oral precursor lesions (simple hyperplasia, n = 11; squamous intraepithelial neoplasia, SIN I-III, n = 35), and OSCC specimen (n = 42) by immunohistochemistry and real-time polymerase chain reaction (qPCR) analysis in OSCC cell lines. Metabolism-related proteins were correlated with proliferation activity (Ki-67) and apoptotic properties (TUNEL assay) in OSCC. Specificity of antibodies was confirmed by western blotting in cancer cell lines. Results Expression of IGF-R1, glycolysis-related proteins (GLUT-1, HK 2, LDHA, TKTL1), and mitochondrial enzymes (SDHA, SDHB, ATP synthase) were significantly increased in the carcinogenesis of OSCC. Metabolic active regions of OSCC were strongly correlated with proliferating cancer (Ki-67+) cells without detection of apoptosis (TUNEL assay). Conclusions This study provides the first evidence of the expression of IGF-R1, glycolysis-related proteins GLUT-1, HK 2, PFK-1, LDHA, and TKTL1, as well as mitochondrial enzymes SDHA, SDHB, and ATP synthase in the multi-step carcinogenesis of OSCC. Both, hypoxia-related glucose metabolism and mitochondrial oxidative phosphorylation characteristics are associated with the carcinogenesis of OSCC. Acidosis and OXPHOS may drive a metabolic shift towards the pentose phosphate pathway (PPP). Therefore, inhibition of the PPP, glycolysis, and targeted anti-mitochondrial therapies (ROS generation) by natural compounds or synthetic vitamin derivatives may act as sensitizer for apoptosis in cancer cells mediated by adjuvant therapies in OSCC. PMID:25048361

  3. Combined enteral infusion of glutamine, carbohydrates, and antioxidants modulates gut protein metabolism in humans.

    Science.gov (United States)

    Coëffier, Moïse; Claeyssens, Sophie; Lecleire, Stéphane; Leblond, Jonathan; Coquard, Aude; Bôle-Feysot, Christine; Lavoinne, Alain; Ducrotté, Philippe; Déchelotte, Pierre

    2008-11-01

    Available data suggest that nutrients can affect intestinal protein metabolism, which contributes to the regulation of gut barrier function. We aimed to assess whether an oral nutritional supplement (ONS) containing glutamine (as the dipeptide Ala-Gln), carbohydrates, and antioxidants would modulate duodenal protein metabolism in healthy humans. Thirty healthy control subjects were included and, over a period of 5 h, received by nasogastric tube either saline or ONS providing 11.7 kcal/kg as 0.877 g Ala-Gln/kg, 3.9 g carbohydrates/kg, and antioxidants (29.25 mg vitamin C/kg, 9.75 mg vitamin E/kg, 195 microg beta-carotene/kg, 5.85 mg Se/kg, and 390 microg Zn/kg) or glutamine (0.585 g/kg, 2.34 kcal/kg). Simultaneously, a continuous intravenous infusion of l-[1-(13)C]-leucine was done until endoscopy. Leucine enrichment was assessed by using gas chromatography-mass spectrometric analysis, and mucosal fractional synthesis rate was calculated by using intracellular amino acid enrichment as precursor. Mucosal proteolytic pathways were also evaluated. ONS infusion resulted in a doubling increase (P < 0.01) of duodenal fractional synthesis rate and a significant (P < 0.05) decrease in cathepsin D-mediated proteolysis compared with saline, whereas proteasome and Ca(2+)-dependent activities were unaffected. ONS infusion significantly (P < 0.01) decreased duodenal glutathione but not glutathione disulfide concentrations or the ratio of glutathione to glutathione disulfide. Insulinemia increased after ONS infusion, whereas plasma essential amino acids decreased. Infusion of glutamine alone did not reproduce ONS effects. ONS infusion improves duodenal protein balance in healthy humans. Further investigations are needed to study the origin of these effects and to evaluate ONS supply in stressed persons.

  4. Comparative proteome analysis of metabolic proteins from seeds of durum wheat (cv. Svevo) subjected to heat stress

    DEFF Research Database (Denmark)

    Laino, Paolo; Shelton, Dale; Finnie, Christine

    2010-01-01

    of nonprolamin proteins were monitored to identify polypeptides affected by heat stress during grain fill. This study shows that heat stress alters significantly the durum wheat seed proteome, although the changes range is only between 1.2- and 2.2-fold. This analysis revealed 132 differentially expressed...... include proteins with metabolic activity or structural function. In order to investigate the consequences of heat stress on the accumulation of nonprolamin proteins in mature durum wheat kernels, the Italian cultivar Svevo was subjected to two thermal regimes (heat stress versus control). The 2-D patterns...... polypeptides, 47 of which were identified by MALDI-TOF and MALDI-TOF-TOF MS and included HSPs, proteins involved in the glycolysis and carbohydrate metabolism, as well as stress-related proteins. Many of the heat-induced polypeptides are considered to be allergenic for sensitive individuals....

  5. A 7-day high protein hypocaloric diet promotes cellular metabolic adaptations and attenuates lean mass loss in healthy males

    OpenAIRE

    Matthew Furber; Ana Anton-Solanas; Emma Koppe; Charlotte Ashby; Michael Roberts; Justin Roberts

    2017-01-01

    Mitochondrial quantity and density are associated with increased oxidative metabolism. It has been demonstrated that a hypocaloric high fat/low carbohydrate (HF/LC) diet can up-regulate transcriptional markers of mitochondrial biogenesis; this was yet to be explored in vivo subsequent to a high protein/low carbohydrate (HP/LC) diet. Thus the aims of the study were to explore such diets on transcriptional markers or mitochondrial biogenesis, body composition and resting metabolic rate (RMR). F...

  6. Global Structural Flexibility of Metalloproteins Regulates Reactivity of Transition Metal Ion in the Protein Core: An Experimental Study Using Thiol-subtilisin as a Model Protein.

    Science.gov (United States)

    Matsuo, Takashi; Kono, Takamasa; Shobu, Isamu; Ishida, Masaya; Gonda, Katsuya; Hirota, Shun

    2018-02-21

    The functions of metal-containing proteins (metalloproteins) are determined by the reactivities of transition metal ions at their active sites. Because protein macromolecular structures have several molecular degrees of freedom, global structural flexibility may also regulate the properties of metalloproteins. However, the influence of this factor has not been fully delineated in mechanistic studies of metalloproteins. Accordingly, we have investigated the relationship between global protein flexibility and the characteristics of a transition metal ion in the protein core using thiol-subtilisin (tSTL) with a Cys-coordinated Cu 2+ ion as a model system. Although tSTL has two Ca 2+ -binding sites, the Ca 2+ -binding status hardly affects its secondary structure. Nevertheless, guanidinium-induced denaturation and amide H/D exchange indicated the increase in the structural flexibility of tSTL by the removal of bound Ca 2+ ions. Electron paramagnetic resonance and absorption spectral changes have revealed that the protein flexibility determines the characteristics of a Cu 2+ ion in tSTL. Therefore, global protein flexibility should be recognized as an important factor that regulates the properties of metalloproteins. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Hibiscus chlorotic ringspot virus coat protein upregulates sulfur metabolism genes for enhanced pathogen defense.

    Science.gov (United States)

    Gao, Ruimin; Ng, Florence Kai Lin; Liu, Peng; Wong, Sek-Man

    2012-12-01

    In both Hibiscus chlorotic ringspot virus (HCRSV)-infected and HCRSV coat protein (CP) agroinfiltrated plant leaves, we showed that sulfur metabolism pathway related genes-namely, sulfite oxidase (SO), sulfite reductase, and adenosine 5'-phosphosulfate kinase-were upregulated. It led us to examine a plausible relationship between sulfur-enhanced resistance (SED) and HCRSV infection. We broadened an established method to include different concentrations of sulfur (0S, 1S, 2S, and 3S) to correlate them to symptom development of HCRSV-infected plants. We treated plants with glutathione and its inhibitor to verify the SED effect. Disease resistance was induced through elevated glutathione contents during HCRSV infection. The upregulation of SO was related to suppression of symptom development induced by sulfur treatment. In this study, we established that HCRSV-CP interacts with SO which, in turn, triggers SED and leads to enhanced plant resistance. Thus, we have discovered a new function of SO in the SED pathway. This is the first report to demonstrate that the interaction of a viral protein and host protein trigger SED in plants. It will be interesting if such interaction applies generally to other host-pathogen interactions that will lead to enhanced pathogen defense.

  8. Sorbitol dehydrogenase is a cytosolic protein required for sorbitol metabolism in Arabidopsis thaliana.

    Science.gov (United States)

    Aguayo, María Francisca; Ampuero, Diego; Mandujano, Patricio; Parada, Roberto; Muñoz, Rodrigo; Gallart, Marta; Altabella, Teresa; Cabrera, Ricardo; Stange, Claudia; Handford, Michael

    2013-05-01

    Sorbitol is converted to fructose in Rosaceae species by SORBITOL DEHYDROGENASE (SDH, EC 1.1.1.14), especially in sink organs. SDH has also been found in non-Rosaceae species and here we show that the protein encoded by At5g51970 in Arabidopsis thaliana (L.) Heynh. possesses the molecular characteristics of an SDH. Using a green fluorescent protein-tagged version and anti-SDH antisera, we determined that SDH is cytosolically localized, consistent with bioinformatic predictions. We also show that SDH is widely expressed, and that SDH protein accumulates in both source and sink organs. In the presence of NAD+, recombinant SDH exhibited greatest oxidative activity with sorbitol, ribitol and xylitol as substrates; other sugar alcohols were oxidized to a lesser extent. Under standard growth conditions, three independent sdh- mutants developed as wild-type. Nevertheless, all three exhibited reduced dry weight and primary root length compared to wild-type when grown in the presence of sorbitol. Additionally, under short-day conditions, the mutants were more resistant to dehydration stress, as shown by a reduced loss of leaf water content when watering was withheld, and a greater survival rate on re-watering. This evidence suggests that limitations in the metabolism of sugar alcohols alter the growth of Arabidopsis and its response to drought. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  9. Acute dim light at night increases body mass, alters metabolism, and shifts core body temperature circadian rhythms.

    Science.gov (United States)

    Borniger, Jeremy C; Maurya, Santosh K; Periasamy, Muthu; Nelson, Randy J

    2014-10-01

    The circadian system is primarily entrained by the ambient light environment and is fundamentally linked to metabolism. Mounting evidence suggests a causal relationship among aberrant light exposure, shift work, and metabolic disease. Previous research has demonstrated deleterious metabolic phenotypes elicited by chronic (>4 weeks) exposure to dim light at night (DLAN) (∼ 5 lux). However, the metabolic effects of short-term (dim light would gain more body mass, alter whole body metabolism, and display altered body temperature (Tb) and activity rhythms compared to mice maintained in dark nights. Our data largely support these predictions; DLAN mice gained significantly more mass, reduced whole body energy expenditure, increased carbohydrate over fat oxidation, and altered temperature circadian rhythms. Importantly, these alterations occurred despite similar activity locomotor levels (and rhythms) and total food intake between groups. Peripheral clocks are potently entrained by body temperature rhythms, and the deregulation of body temperature we observed may contribute to metabolic problems due to "internal desynchrony" between the central circadian oscillator and temperature sensitive peripheral clocks. We conclude that even relatively short-term exposure to low levels of nighttime light can influence metabolism to increase mass gain.

  10. Alcohol Induced Hepatic Degeneration in a Hepatitis C Virus Core Protein Transgenic Mouse Model

    Directory of Open Access Journals (Sweden)

    Dong-Hyung Noh

    2014-03-01

    Full Text Available Hepatitis C virus (HCV has become a major public health issue. It is prevalent in most countries. HCV infection frequently begins without clinical symptoms, before progressing to persistent viremia, chronic hepatitis, cirrhosis and hepatocellular carcinoma (HCC in the majority of patients (70% to 80%. Alcohol is an independent cofactor that accelerates the development of HCC in chronic hepatitis C patients. The purpose of the current study was to evaluate ethanol-induced hepatic changes in HCV core-Tg mice and mutant core Tg mice. Wild type (NTG, core wild-Tg mice (TG-K, mutant core 116-Tg mice (TG-116 and mutant core 99-Tg mice (TG-99 were used in this investigation. All groups were given drinking water with 10% ethanol and 5% sucrose for 13 weeks. To observe liver morphological changes, we performed histopathological and immunohistochemical examinations. Histopathologically, NTG, TG-K and TG-116 mice showed moderate centrilobular necrosis, while severe centrilobular necrosis and hepatocyte dissociation were observed in TG-99 mice with increasing lymphocyte infiltration and piecemeal necrosis. In all groups, a small amount of collagen fiber was found, principally in portal areas. None of the mice were found to have myofibroblasts based on immunohistochemical staining specific for α-SMA. CYP2E1-positive cells were clearly detected in the centrilobular area in all groups. In the TG-99 mice, we also observed cells positive for CK8/18, TGF-β1 and phosphorylated (p-Smad2/3 and p21 around the necrotic hepatocytes in the centrilobular area (p < 0.01. Based on our data, alcohol intake induced piecemeal necrosis and hepatocyte dissociation in the TG-99 mice. These phenomena involved activation of the TGF-β1/p-Smad2/3/p21 signaling pathway in hepatocytes. Data from this study will be useful for elucidating the association between alcohol intake and HCV infection.

  11. Getting to the core of protein pharmaceuticals – comprehensive structure analysis by mass spectrometry

    DEFF Research Database (Denmark)

    Leurs, Ulrike; Mistarz, Ulrik Hvid; Rand, Kasper Dyrberg

    2015-01-01

    . Mass spectrometry has evolved as a powerful tool for the characterization of both primary and higher order structures of protein pharmaceuticals. Furthermore, the chemical and physical stability of protein drugs, as well as their pharmacokinetics are nowadays routinely determined by mass spectrometry...

  12. Charge neutralization as the major factor for the assembly of nucleocapsid-like particles from C-terminal truncated hepatitis C virus core protein

    OpenAIRE

    Theo Luiz Ferraz de Souza; Sheila Maria Barbosa de Lima; Vanessa L. de Azevedo Braga; David S. Peabody; Davis Fernandes Ferreira; M. Lucia Bianconi; Andre Marco de Oliveira Gomes; Jerson Lima Silva; Andréa Cheble de Oliveira

    2016-01-01

    Background Hepatitis C virus (HCV) core protein, in addition to its structural role to form the nucleocapsid assembly, plays a critical role in HCV pathogenesis by interfering in several cellular processes, including microRNA and mRNA homeostasis. The C-terminal truncated HCV core protein (C124) is intrinsically unstructured in solution and is able to interact with unspecific nucleic acids, in the micromolar range, and to assemble into nucleocapsid-like particles (NLPs) in vitro. The specific...

  13. [Relationship between high-sensitivity C-reactive protein and obesity/metabolic syndrome in children].

    Science.gov (United States)

    Chen, Fangfang; Wang, Wenpeng; Teng, Yue; Hou, Dongqing; Zhao, Xiaoyuan; Yang, Ping; Yan, Yinkun; Mi, Jie

    2014-06-01

    To explore the relationship between high-sensitivity C-reactive protein (hsCRP) and obesity/metabolic syndrome (MetS) related factors in children. 403 children aged 10-14 and born in Beijing were involved in this study. Height, weight, waist circumference, fat mass percentage (Fat%), blood pressure (BP), hsCRP, triglyceride (TG), total cholesterol (TC), fasting plasma glucose (FPG), high and low density lipoprotein cholesterol (HDL-C, LDL-C) were observed among these children. hsCRP was transformed with base 10 logarithm (lgCRP). MetS was defined according to the International Diabetes Federation 2007 definition. Associations between MetS related components and hsCRP were tested using partial correlation analysis, analysis of covariance and linear regression models. 1) lgCRP was positively correlated with BMI, waist circumference, Fat%,BP, FPG, LDL-C and TC while negatively correlated with HDL-C. With BMI under control, the relationships disappeared, but LDL-C (r = 0.102). 2) The distributions of lgCRP showed obvious differences in all the metabolic indices, in most groups, respectively. With BMI under control, close relationships between lgCRP and high blood pressure/high TG disappeared and the relationship with MetS weakened. 3) Through linear regression models, factors as waist circumference, BMI, Fat% were the strongest factors related to hsCRP, followed by systolic BP, HDL-C, diastolic BP, TG and LDL-C. With BMI under control, the relationships disappeared, but LDL-C(β = 0.045). hsCRP was correlated with child obesity, lipid metabolism and MetS. Waist circumference was the strongest factors related with hsCRP. Obesity was the strongest and the independent influencing factor of hsCRP.

  14. Relative Abundance of Integral Plasma Membrane Proteins in Arabidopsis Leaf and Root Tissue Determined by Metabolic Labeling and Mass Spectrometry

    Science.gov (United States)

    Bernfur, Katja; Larsson, Olaf; Larsson, Christer; Gustavsson, Niklas

    2013-01-01

    Metabolic labeling of proteins with a stable isotope (15N) in intact Arabidopsis plants was used for accurate determination by mass spectrometry of differences in protein abundance between plasma membranes isolated from leaves and roots. In total, 703 proteins were identified, of which 188 were predicted to be integral membrane proteins. Major classes were transporters, receptors, proteins involved in membrane trafficking and cell wall-related proteins. Forty-one of the integral proteins, including nine of the 13 isoforms of the PIP (plasma membrane intrinsic protein) aquaporin subfamily, could be identified by peptides unique to these proteins, which made it possible to determine their relative abundance in leaf and root tissue. In addition, peptides shared between isoforms gave information on the proportions of these isoforms. A comparison between our data for protein levels and corresponding data for mRNA levels in the widely used database Genevestigator showed an agreement for only about two thirds of the proteins. By contrast, localization data available in the literature for 21 of the 41 proteins show a much better agreement with our data, in particular data based on immunostaining of proteins and GUS-staining of promoter activity. Thus, although mRNA levels may provide a useful approximation for protein levels, detection and quantification of isoform-specific peptides by proteomics should generate the most reliable data for the proteome. PMID:23990937

  15. Heparan sulfate proteoglycan from the extracellular matrix of human lung fibroblasts. Isolation, purification, and core protein characterization

    International Nuclear Information System (INIS)

    Heremans, A.; Cassiman, J.J.; Van den Berghe, H.; David, G.

    1988-01-01

    Confluent cultured human lung fibroblasts were labeled with 35SO4(2-). After 48 h of labeling, the pericellular matrix was prepared by Triton X-100 and deoxycholate extraction of the monolayers. Heparan sulfate proteoglycan (HSPG) accounted for nearly 80% of the total matrix [35S]proteoglycans. After solubilization in 6 M guanidinium HCl and cesium chloride density gradient centrifugation, the majority (78%) of these [35S] HSPG equilibrated at an average buoyant density of 1.35 g/ml. This major HSPG fraction was purified by ion-exchange chromatography on Mono Q and by gel filtration on Sepharose CL-4B, and further characterized by gel electrophoresis and immunoblotting. Intact [35S]HSPG eluted with Kav 0.1 from Sepharose CL-4B, whereas the protein-free [35S]heparan sulfate chains, obtained by alkaline borohydride treatment of the proteoglycan fractions, eluted with Kav 0.45 (Mr approximately 72,000). When analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography, core (protein) preparations, obtained by heparitinase digestion of 125I-labeled HSPG fractions, yielded one major labeled band with apparent molecular mass of approximately 300 kDa. Reduction with beta-mercaptoethanol slightly increased the apparent Mr of the labeled band, suggesting a single polypeptide structure and the presence of intrachain disulfide bonds. Immunoadsorption experiments and immunostaining of electrophoretically separated heparitinase-digested core proteins with monoclonal antibodies raised against matrix and cell surface-associated HSPG suggested that the major matrix-associated HSPG of cultured human lung fibroblasts is distinct from the HSPG that are anchored in the membranes of these cells. Binding studies suggested that this matrix HSPG interacts with several matrix components, both through its glycosaminoglycan chains and through its heparitinase-resistant core. (Abstract Truncated)

  16. AMP-activated protein kinase: Role in metabolism and therapeutic implications.

    Science.gov (United States)

    Schimmack, Greg; Defronzo, Ralph A; Musi, Nicolas

    2006-11-01

    AMP-activated protein kinase (AMPK) is an enzyme that works as a fuel gauge which becomes activated in situations of energy consumption. AMPK functions to restore cellular ATP levels by modifying diverse metabolic and cellular pathways. In the skeletal muscle, AMPK is activated during exercise and is involved in contraction-stimulated glucose transport and fatty acid oxidation. In the heart, AMPK activity increases during ischaemia and functions to sustain ATP, cardiac function and myocardial viability. In the liver, AMPK inhibits the production of glucose, cholesterol and triglycerides and stimulates fatty acid oxidation. Recent studies have shown that AMPK is involved in the mechanism of action of metformin and thiazolidinediones, and the adipocytokines leptin and adiponectin. These data, along with evidence that pharmacological activation of AMPK in vivo improves blood glucose homeostasis, cholesterol concentrations and blood pressure in insulin-resistant rodents, make this enzyme an attractive pharmacological target for the treatment of type 2 diabetes, ischaemic heart disease and other metabolic diseases.

  17. Saliva C-reactive protein as a biomarker of metabolic syndrome in diabetic patients.

    Science.gov (United States)

    Dezayee, Zhian Mahmood Ibrahim; Al-Nimer, Marwan Salih Mohamad

    2016-01-01

    Human C-reactive protein (CRP) has been used in the risk assessment of coronary events. Human saliva mirrors the body's health and well-being and is noninvasive, easy to collect, and ideal for third-world countries as well as for large patient screening. This study aimed to screen the saliva CRP qualitatively in patients with diabetes (Type 1 and 2) taking in considerations, the diagnostic criteria of metabolic syndrome. Center for diabetes mellitus, prospective study. A total number of 50 Type 2 diabetes (T2D) patients, 25 Type 1 diabetes (T1D) patients, and 25 healthy subjects were recruited from the center for diabetes mellitus. Each patient was assessed clinically, and the anthropometric measures, glycemic status, and lipid profiles were determined. Stimulated salivary flow rate and saliva CRP were determined. All calculations analysis was made using Excel 2003 program for Windows. The results showed that the salivary flow rate in T1D was less than healthy subjects and T2D and CRP was found positive (6 mg/L) in 36% and 56% of patients with T1D and T2D, respectively. Saliva CRP was found to be related to the anthropometric measurement, blood pressure, and glycemic control. We conclude that saliva CRP may be used as a biomarker for metabolic syndrome and its value is obvious in T2D rather than in T1D.

  18. Influence of multidrug resistance and drug transport proteins on chemotherapy drug metabolism.

    Science.gov (United States)

    Joyce, Helena; McCann, Andrew; Clynes, Martin; Larkin, Annemarie

    2015-05-01

    Chemotherapy involving the use of anticancer drugs remains an important strategy in the overall management of patients with metastatic cancer. Acquisition of multidrug resistance remains a major impediment to successful chemotherapy. Drug transporters in cell membranes and intracellular drug metabolizing enzymes contribute to the resistance phenotype and determine the pharmacokinetics of anticancer drugs in the body. ATP-binding cassette (ABC) transporters mediate the transport of endogenous metabolites and xenobiotics including cytotoxic drugs out of cells. Solute carrier (SLC) transporters mediate the influx of cytotoxic drugs into cells. This review focuses on the substrate interaction of these transporters, on their biology and what role they play together with drug metabolizing enzymes in eliminating therapeutic drugs from cells. The majority of anticancer drugs are substrates for the ABC transporter and SLC transporter families. Together, these proteins have the ability to control the influx and the efflux of structurally unrelated chemotherapeutic drugs, thereby modulating the intracellular drug concentration. These interactions have important clinical implications for chemotherapy because ultimately they determine therapeutic efficacy, disease progression/relapse and the success or failure of patient treatment.

  19. Proteome-Level Analysis of Metabolism- and Stress-Related Proteins during Seed Dormancy and Germination in Gnetum parvifolium.

    Science.gov (United States)

    Chang, Ermei; Deng, Nan; Zhang, Jin; Liu, Jianfeng; Chen, Lanzhen; Zhao, Xiulian; Abbas, M; Jiang, Zeping; Shi, Shengqing

    2018-03-21

    Gnetum parvifolium is a rich source of materials for traditional medicines, food, and oil, but little is known about the mechanism underlying its seed dormancy and germination. In this study, we analyzed the proteome-level changes in its seeds during germination using isobaric tags for relative and absolute quantitation. In total, 1,040 differentially expressed proteins were identified, and cluster analysis revealed the distinct time points during which signal transduction and oxidation-reduction activity changed. Gene Ontology analysis showed that "carbohydrate metabolic process" and "response to oxidative stress" were the main enriched terms. Proteins associated with starch degradation and antioxidant enzymes were important for dormancy-release, while proteins associated with energy metabolism and protein synthesis were up-regulated during germination. Moreover, protein-interaction networks were mainly associated with heat-shock proteins. Furthermore, in accord with changes in the energy metabolism- and antioxidant-related proteins, indole-3-acetic acid, Peroxidase, and soluble sugar content increased, and the starch content decreased in almost all six stages of dormancy and germination analyzed (S1-S6). The activity of superoxide dismutase, abscisic acid, and malondialdehyde content increased in the dormancy stages (S1-S3) and then decreased in the germination stages (S4-S6). Our results provide new insights into G. parvifolium seed dormancy and germination at the proteome and physiological levels, with implications for improving seed propagation.

  20. Energy dense, protein restricted diet increases adiposity and perturbs metabolism in young, genetically lean pigs.

    Directory of Open Access Journals (Sweden)

    Kimberly D Fisher

    Full Text Available Animal models of obesity and metabolic dysregulation during growth (or childhood are lacking. Our objective was to increase adiposity and induce metabolic syndrome in young, genetically lean pigs. Pre-pubertal female pigs, age 35 d, were fed a high-energy diet (HED; n = 12, containing 15% tallow, 35% refined sugars and 9.1-12.9% crude protein, or a control corn-based diet (n = 11 with 12.2-19.2% crude protein for 16 wk. Initially, HED pigs self-regulated energy intake similar to controls, but by wk 5, consumed more (P<0.001 energy per kg body weight. At wk 15, pigs were subjected to an oral glucose tolerance test (OGTT; blood glucose increased (P<0.05 in control pigs and returned to baseline levels within 60 min. HED pigs were hyperglycemic at time 0, and blood glucose did not return to baseline (P = 0.01, even 4 h post-challenge. During OGTT, glucose area under the curve (AUC was higher and insulin AUC was lower in HED pigs compared to controls (P = 0.001. Chronic HED intake increased (P<0.05 subcutaneous, intramuscular, and perirenal fat deposition, and induced hyperglycemia, hypoinsulinemia, and low-density lipoprotein hypercholesterolemia. A subset of HED pigs (n = 7 was transitioned back to a control diet for an additional six weeks. These pigs were subjected to an additional OGTT at 22 wk. Glucose AUC and insulin AUC did not improve, supporting that dietary intervention was not sufficient to recover glucose tolerance or insulin production. These data suggest a HED may be used to increase adiposity and disrupt glucose homeostasis in young, growing pigs.

  1. A recruiting protein of geranylgeranyl diphosphate synthase controls metabolic flux toward chlorophyll biosynthesis in rice.

    Science.gov (United States)

    Zhou, Fei; Wang, Cheng-Yuan; Gutensohn, Michael; Jiang, Ling; Zhang, Peng; Zhang, Dabing; Dudareva, Natalia; Lu, Shan

    2017-06-27

    In plants, geranylgeranyl diphosphate (GGPP) is produced by plastidic GGPP synthase (GGPPS) and serves as a precursor for vital metabolic branches, including chlorophyll, carotenoid, and gibberellin biosynthesis. However, molecular mechanisms regulating GGPP allocation among these biosynthetic pathways localized in the same subcellular compartment are largely unknown. We found that rice contains only one functionally active GGPPS, OsGGPPS1, in chloroplasts. A functionally active homodimeric enzyme composed of two OsGGPPS1 subunits is located in the stroma. In thylakoid membranes, however, the GGPPS activity resides in a heterodimeric enzyme composed of one OsGGPPS1 subunit and GGPPS recruiting protein (OsGRP). OsGRP is structurally most similar to members of the geranyl diphosphate synthase small subunit type II subfamily. In contrast to members of this subfamily, OsGRP enhances OsGGPPS1 catalytic efficiency and specificity of GGPP production on interaction with OsGGPPS1. Structural biology and protein interaction analyses demonstrate that affinity between OsGRP and OsGGPPS1 is stronger than between two OsGGPPS1 molecules in homodimers. OsGRP determines OsGGPPS1 suborganellar localization and directs it to a large protein complex in thylakoid membranes, consisting of geranylgeranyl reductase (OsGGR), light-harvesting-like protein 3 (OsLIL3), protochlorophyllide oxidoreductase (OsPORB), and chlorophyll synthase (OsCHLG). Taken together, genetic and biochemical analyses suggest OsGRP functions in recruiting OsGGPPS1 from the stroma toward thylakoid membranes, thus providing a mechanism to control GGPP flux toward chlorophyll biosynthesis.

  2. Metabolic syndrome, C-reactive protein and cardiovascular risk in psoriasis patients: a cross-sectional study*

    Science.gov (United States)

    Paschoal, Renato Soriani; Silva, Daniela Antoniali; Cardili, Renata Nahas; Souza, Cacilda da Silva

    2018-01-01

    Background Psoriasis has been associated with co-morbidities and elevated cardiovascular risk. Objectives To analyze the relationships among metabolic syndrome, cardiovascular risk, C-reactive protein, gender, and Psoriasis severity. Methods In this cross-sectional study, plaque Psoriasis patients (n=90), distributed equally in gender, were analyzed according to: Psoriasis Area and Severity Index, cardiovascular risk determined by the Framingham risk score and global risk assessment, C-reactive protein and metabolic syndrome criteria (NCEPT-ATP III). Results Metabolic syndrome frequency was 43.3% overall, without significance between genders (P=0.14); but women had higher risk for obesity (OR 2.56, 95%CI 1.02-6.41; P=0.04) and systemic arterial hypertension (OR 3.29, 95%CI 1.39-7.81; P=0.006). The increase in the Psoriasis Area and Severity Index also increased the risk for metabolic syndrome (OR 1.060, 95%CI 1.006-1.117; P=0.03). Absolute 10-year cardiovascular risk was higher in males (P=0.002), but after global risk assessment, 51.1% patients, 52.2% women, were re-classified as high-intermediate cardiovascular risk; without significance between genders (P=0.83). C-reactive protein level was elevated nearly six-fold overall, higher in metabolic syndrome (P=0.05), systemic arterial hypertension (P=0.004), and high-intermediate 10-year cardiovascular risk patients (Preactive protein patients (t=1.98; P=0.05). Study limitations Restricted sample, hospital-based and representative of a single center and no specification of psoriatic arthritis. Conclusions Psoriasis, metabolic syndrome, systemic arterial hypertension and age share the increase in C-reactive protein, which could implicate in additional burden for increasing the cardiovascular risk and be an alert for effective interventions. PMID:29723366

  3. Exercise training and work task induced metabolic and stress-related mRNA and protein responses in myalgic muscles

    DEFF Research Database (Denmark)

    Sjøgaard, Gisela; Zebis, Mette Kreutzfeldt; Kiilerich, Kristian

    2013-01-01

    healthy controls. Those with myalgia performed similar to 7 hrs repetitive stressful work and were subsequently randomized to 10 weeks of specific strength training, general fitness training, or reference intervention. Muscles biopsies were taken from the trapezius muscle at baseline, after work and after...... 10 weeks intervention. The main findings are that the capacity of carbohydrate oxidation was reduced in myalgic compared with healthy muscle. Repetitive stressful work increased mRNA content for heat shock proteins and decreased levels of key regulators for growth and oxidative metabolism......The aim was to assess mRNA and/or protein levels of heat shock proteins, cytokines, growth regulating, and metabolic proteins in myalgic muscle at rest and in response to work tasks and prolonged exercise training. A randomized controlled trial included 28 females with trapezius myalgia and 16...

  4. The adverse effect of 4-tert-octylphenol on fat metabolism in pregnant rats via regulation of lipogenic proteins.

    Science.gov (United States)

    Kim, Jun; Kang, Eun-Jin; Park, Mee-Na; Kim, Ji-Eun; Kim, Seung-Chul; Jeung, Eui-Bae; Lee, Geun-Shik; Hwang, Dae-Youn; An, Beum-Soo

    2015-07-01

    Alkylphenols such as 4-tert-octylphenol (OP), nonylphenol, and bisphenol A are classified as endocrine-disrupting chemicals (EDCs). Digestion and metabolism of food are controlled by many endocrine factors, including insulin, glucagon, and estrogen. These factors are differentially regulated during pregnancy. The alteration of nutritional intake and fat metabolism may affect the maintenance of pregnancy and supplementation of nutrients to the fetus, and therefore can cause severe metabolic diseases such as ketosis, marasmus and diabetes mellitus in pregnant individuals. In this study, we examined the effects of OP on fat metabolism in pregnant rats. Ethinyl estradiol (EE) was also administered as an estrogenic positive control. In our results, rats treated with OP showed significantly reduced body weights compared to the control group. In addition, histological analysis showed that the amount of fat deposited in adipocytes was reduced by OP treatment. To study the mechanism of action of OP in fat metabolism, we examined the expression levels of fat metabolism-associated genes in rat adipose tissue and liver by real-time PCR. OP and EE negatively regulated the expression of lipogenic enzymes, including FAS (fatty acid synthase), ACC-1 (acetyl-CoA carboxylase-1), and SCD-1 (stearoyl-CoA desaturase-1). The levels of lipogenic enzyme-associated transcription factors such as C/EBP-α (CAAT enhancer binding protein alpha) and SREBP-1c (sterol regulatory element binding protein-1c) were also reduced in both liver and adipose tissue. In summary, these findings suggest that OP has adverse effects on fat metabolism in pregnant rats and inhibits fat deposition via regulating lipogenic genes in the liver and adipose tissue. The altered fat metabolism by OP may affect the nutrition balance during pregnancy and can cause metabolism-related diseases. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. Enhancement of curcumin water dispersibility and antioxidant activity using core-shell protein-polysaccharide nanoparticles.

    Science.gov (United States)

    Huang, Xiaoxia; Huang, Xulin; Gong, Yushi; Xiao, Hang; McClements, David Julian; Hu, Kun

    2016-09-01

    Curcumin has strong antioxidant activity, but poor water-solubility and chemical stability, which limits its utilization as a nutraceutical in many applications. Previously, we developed a core-shell (zein-pectin) nanoparticle delivery system with high curcumin loading efficiency, high particle yield, and good water dispersibility. However, this system was unstable to aggregation around neutral pH and moderate ionic strengths due to weakening of electrostatic repulsion between nanoparticles. In the current study, we used a combination of alginate (high charge density) and pectin (low charge density) to form the shell around zein nanoparticles. Replacement of 30% of pectin with alginate greatly improved aggregation stability at pH 5 to 7 and at high ionic strengths (2000mM NaCl). Curcumin encapsulated within these core-shell nanoparticles exhibited higher antioxidant and radical scavenging activities than curcumin solubilized in ethanol solutions as determined by Fe (III) reducing power, 1, 1-Diphenyl-2-picrylhydrazyl free radical (DPPH·), and 2, 2'-azinobis-(3-ethylbenzothiazoline)-6-sulfonic acid radical cation (ABTS· + ) scavenging analysis. These core-shell nanoparticles may be useful for incorporating chemically unstable hydrophobic nutraceuticals such as curcumin into functional foods, dietary supplements, and pharmaceuticals. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Metabolism of leucine and alanine in growing rats fed the diets with various protein to energy ratios

    International Nuclear Information System (INIS)

    Tanaka, Hideyuki; Yamaguchi, Michio; Kametaka, Masao

    1975-01-01

    In order to clarify the nutritional significance of metabolism of the carbon skeleton of individual amino acids, the metabolic fates of L-leucine-U- 14 C and L-alanine-U- 14 C were investigated in growing rats fed the diets with various protein calories percents (PC%) at 410 kcal of metabolizable energy. The incorporation of 14 C into body protein in 12 hr after the injection of leucine- 14 C was about 73% of the dose in the 0 and 5 PC% groups, though it decreased with increasing the levels of dietary protein from 10 to 30 PC%. The value of 14 C recovery in body protein almost agreed with the net protein utilization (NPU) determined for the whole egg protein in a similar experimental condition. The 14 C recovery in expired CO 2 and body lipid suggested that the carbon skeleton of leucine is well utilized as an energy source when the dietary carbohydrate is extensively replaced by protein. While, the incorporation of 14 C into body protein from alanine- 14 C was less than about 11% of the dose in all the dietary groups, and the majority of 14 C was recovered in expired CO 2 and body lipid in a remarked contrast to leucine. A similar pattern in urinary excretion of 14 C was obtained for these amino acids, and the refracted rise of 14 C from 10 PC% may give an indication for minimum protein requirements. (auth.)

  7. AMP-Activated Protein Kinase (AMPK) Regulates Energy Metabolism through Modulating Thermogenesis in Adipose Tissue

    Science.gov (United States)

    Wu, Lingyan; Zhang, Lina; Li, Bohan; Jiang, Haowen; Duan, Yanan; Xie, Zhifu; Shuai, Lin; Li, Jia; Li, Jingya

    2018-01-01

    Obesity occurs when excess energy accumulates in white adipose tissue (WAT), whereas brown adipose tissue (BAT), which is specialized in dissipating energy through thermogenesis, potently counteracts obesity. White adipocytes can be converted to thermogenic “brown-like” cells (beige cells; WAT browning) under various stimuli, such as cold exposure. AMP-activated protein kinase (AMPK) is a crucial energy sensor that regulates energy metabolism in multiple tissues. However, the role of AMPK in adipose tissue function, especially in the WAT browning process, is not fully understood. To illuminate the effect of adipocyte AMPK on energy metabolism, we generated Adiponectin-Cre-driven adipose tissue-specific AMPK α1/α2 KO mice (AKO). These AKO mice were cold intolerant and their inguinal WAT displayed impaired mitochondrial integrity and biogenesis, and reduced expression of thermogenic markers upon cold exposure. High-fat-diet (HFD)-fed AKO mice exhibited increased adiposity and exacerbated hepatic steatosis and fibrosis and impaired glucose tolerance and insulin sensitivity. Meanwhile, energy expenditure and oxygen consumption were markedly decreased in the AKO mice both in basal conditions and after stimulation with a β3-adrenergic receptor agonist, CL 316,243. In contrast, we found that in HFD-fed obese mouse model, chronic AMPK activation by A-769662 protected against obesity and related metabolic dysfunction. A-769662 alleviated HFD-induced glucose intolerance and reduced body weight gain and WAT expansion. Notably, A-769662 increased energy expenditure and cold tolerance in HFD-fed mice. A-769662 treatment also induced the browning process in the inguinal fat depot of HFD-fed mice. Likewise, A-769662 enhanced thermogenesis in differentiated inguinal stromal vascular fraction (SVF) cells via AMPK signaling pathway. In summary, a lack of adipocyte AMPKα induced thermogenic impairment and obesity in response to cold and nutrient-overload, respectively

  8. Analysis of hepatitis C virus core/NS5A protein co-localization using novel cell culture systems expressing core-NS2 and NS5A of genotypes 1-7

    DEFF Research Database (Denmark)

    Galli, Andrea; Scheel, Troels K H; Prentoe, Jannick C

    2013-01-01

    Hepatitis C virus (HCV) is an important human pathogen infecting hepatocytes. With the advent of infectious cell culture systems, the HCV particle assembly and release processes are finally being uncovered. The HCV core and NS5A proteins co-localize on cytoplasmic lipid droplets (c......LDs) or on the endoplasmic reticulum (ER) at different stages of particle assembly. Current knowledge on assembly and release is primarily based on studies in genotype 2a cell culture systems; however, given the high genetic heterogeneity of HCV, variations might exist among genotypes. Here, we developed novel HCV strain...... JFH1-based recombinants expressing core-NS2 and NS5A from genotypes 1-7, and analysed core and NS5A co-localization in infected cells. Huh7.5 cells were transfected with RNA of core-NS2/NS5A recombinants and putative adaptive mutations were analysed by reverse genetics. Adapted core-NS2/NS5A...

  9. Comparative proteomic analyses reveal that the regulators of G-protein signaling proteins regulate amino acid metabolism of the rice blast fungus Magnaporthe oryzae.

    Science.gov (United States)

    Zhang, Haifeng; Ma, Hongyu; Xie, Xin; Ji, Jun; Dong, Yanhan; Du, Yan; Tang, Wei; Zheng, Xiaobo; Wang, Ping; Zhang, Zhengguang

    2014-11-01

    The rice blast fungus Magnaporthe oryzae encodes eight regulators of G-protein (GTP-binding protein) signaling (RGS) proteins MoRgs1-MoRgs8 that orchestrate the growth, asexual/sexual production, appressorium differentiation, and pathogenicity. To address the mechanisms by which MoRgs proteins function, we conducted a 2DE proteome study and identified 82 differentially expressed proteins by comparing five ∆Morgs mutants with wild-type Guy11 strain. We found that the abundances of eight amino acid (AA) biosynthesis or degradation associated proteins were markedly altered in five ∆Morgs mutants, indicating one of the main collective roles for the MoRgs proteins is to influence AA metabolism. We showed that MoRgs proteins have distinct roles in AA metabolism and nutrient responses from growth assays. In addition, we characterized MoLys20 (Lys is lysine), a homocitrate synthase, whose abundance was significantly decreased in the ∆Morgs mutants. The ∆Molys20 mutant is auxotrophic for lys and exogenous lys could partially rescue its auxotrophic defects. Deletion of MoLYS20 resulted in defects in conidiation and infection, as well as pathogenicity on rice. Overall, our results indicate that one of the critical roles for MoRgs proteins is to regulate AA metabolism, and that MoLys20 may be directly or indirectly regulated by MoRgs and participated in lys biosynthesis, thereby affecting fungal development and pathogenicity. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Significance of glycolytic metabolism-related protein expression in colorectal cancer, lymph node and hepatic metastasis

    International Nuclear Information System (INIS)

    Martins, Sandra Fernandes; Amorim, Ricardo; Viana-Pereira, Marta; Pinheiro, Céline; Costa, Ricardo Filipe Alves; Silva, Patrícia; Couto, Carla; Alves, Sara; Fernandes, Sara; Vilaça, Sónia; Falcão, Joaquim; Marques, Herlander; Pardal, Fernando; Rodrigues, Mesquita; Preto, Ana; Reis, Rui Manuel; Longatto-Filho, Adhemar; Baltazar, Fátima

    2016-01-01

    Colorectal cancer (CRC) is one of the most common malignancies and a leading cause of cancer death worldwide. Most cancer cells display high rates of glycolysis with production of lactic acid, which is then exported to the microenvironment by monocarboxylate transporters (MCTs). The main aim of this study was to evaluate the significance of MCT expression in a comprehensive series of primary CRC cases, lymph node and hepatic metastasis. Expressions of MCT1, MCT4, CD147 and GLUT1 were studied in human samples of CRC, lymph node and hepatic metastasis, by immunohistochemistry. All proteins were overexpressed in primary CRC, lymph node and hepatic metastasis, when compared with non-neoplastic tissue, with exception of MCT1 in lymph node and hepatic metastasis. MCT1 and MCT4 expressions were associated with CD147 and GLUT1 in primary CRC. These markers were associated with clinical pathological features, reflecting the putative role of these metabolism-related proteins in the CRC setting. These findings provide additional evidence for the pivotal role of MCTs in CRC maintenance and progression, and support the use of MCTs as biomarkers and potential therapeutic targets in primary and metastatic CRC

  11. Contribution of cellular retinol-binding protein type 1 to retinol metabolism during mouse development.

    Science.gov (United States)

    Matt, Nicolas; Schmidt, Carsten K; Dupé, Valérie; Dennefeld, Christine; Nau, Heinz; Chambon, Pierre; Mark, Manuel; Ghyselinck, Norbert B

    2005-05-01

    Within cells, retinol (ROL) is bound to cytoplasmic proteins (cellular retinol-binding proteins [CRBPs]), whose proposed function is to protect it from unspecific enzymes through channeling to retinoid-metabolizing pathways. We show that, during development, ROL and retinyl ester levels are decreased in CRBP type 1 (CRBP1) -deficient embryos and fetuses by 50% and 80%, respectively. The steady state level of retinoic acid (RA) is also decreased but to a lesser extent. However, CRBP1-null fetuses do not exhibit the abnormalities characteristic of a vitamin A-deficiency syndrome. Neither CRBP1 deficiency alters the expression patterns of RA-responding genes during development, nor does CRBP1 availability modify the expression of an RA-dependent gene in primary embryonic fibroblasts treated with ROL. Therefore, CRBP1 is required in prenatal life to maintain normal amounts of ROL and to ensure its efficient storage but seems of secondary importance for RA synthesis, at least under conditions of maternal vitamin A sufficiency. Copyright 2005 Wiley-Liss, Inc.

  12. Nitrogen metabolism and protozoa production rate in cattle fed on diet containing protected protein

    International Nuclear Information System (INIS)

    Singh, G.P.; Gupta, B.N.

    1992-01-01

    Nitrogen metabolism and protozoa production rate using 14 C-choline as marker were studied on 9 adult male crossbred (Tharparker x Brown Swiss) rumen fistulated animals divided into 3 groups (A, B and C). All the animals were fed concentrate mixture and wheatstraw. However, groundnut cake (GNC) in concentrate mixture was untreated in group A, 50 per cent formaldehyde treated in group B and 100 per cent formaldehyde treated in group C. Although, DM intake was similar in these groups but water intake was significantly (P<0.05) higher in control group. Total-N, ammonia-N and blood urea were significantly lower in group B and C as compared to group A. Apparent CP digestibility was not affected by addition of formaldehyde treated GNC at 50 and 100 per cent levels. However, N balances increased significantly (P<0.05) due to addition of protected protein in diet. Protozoal pool as well as production rate were significantly (P<0.01) decreased due to formaldehyde treatment of GNC protein. Thus addition of formaldehyde treated GNC in diets decreased ammonia and protozoa production but increased N retention in groups B and C. (author). 27 refs., 3 tabs., 2 figs

  13. Aroclor 1254, a developmental neurotoxicant, alters energy metabolism- and intracellular signaling-associated protein networks in rat cerebellum and hippocampus

    International Nuclear Information System (INIS)

    Kodavanti, Prasada Rao S.; Osorio, Cristina; Royland, Joyce E.; Ramabhadran, Ram; Alzate, Oscar

    2011-01-01

    The vast literature on the mode of action of polychlorinated biphenyls (PCBs) indicates that PCBs are a unique model for understanding the mechanisms of toxicity of environmental mixtures of persistent chemicals. PCBs have been shown to adversely affect psychomotor function and learning and memory in humans. Although the molecular mechanisms for PCB effects are unclear, several studies indicate that the disruption of Ca 2+ -mediated signal transduction plays significant roles in PCB-induced developmental neurotoxicity. Culminating events in signal transduction pathways include the regulation of gene and protein expression, which affects the growth and function of the nervous system. Our previous studies showed changes in gene expression related to signal transduction and neuronal growth. In this study, protein expression following developmental exposure to PCB is examined. Pregnant rats (Long Evans) were dosed with 0.0 or 6.0 mg/kg/day of Aroclor-1254 from gestation day 6 through postnatal day (PND) 21, and the cerebellum and hippocampus from PND14 animals were analyzed to determine Aroclor 1254-induced differential protein expression. Two proteins were found to be differentially expressed in the cerebellum following PCB exposure while 18 proteins were differentially expressed in the hippocampus. These proteins are related to energy metabolism in mitochondria (ATP synthase, sub unit β (ATP5B), creatine kinase, and malate dehydrogenase), calcium signaling (voltage-dependent anion-selective channel protein 1 (VDAC1) and ryanodine receptor type II (RyR2)), and growth of the nervous system (dihydropyrimidinase-related protein 4 (DPYSL4), valosin-containing protein (VCP)). Results suggest that Aroclor 1254-like persistent chemicals may alter energy metabolism and intracellular signaling, which might result in developmental neurotoxicity. -- Highlights: ► We performed brain proteomic analysis of rats exposed to the neurotoxicant, Aroclor 1254. ► Cerebellum and

  14. Protein-energy malnutrition at mid-adulthood does not imprint long-term metabolic consequences in male rats.

    Science.gov (United States)

    Malta, Ananda; de Moura, Egberto Gaspar; Ribeiro, Tatiane Aparecida; Tófolo, Laize Peron; Abdennebi-Najar, Latifa; Vieau, Didier; Barella, Luiz Felipe; de Freitas Mathias, Paulo Cezar; Lisboa, Patrícia Cristina; de Oliveira, Júlio Cezar

    2016-06-01

    The long-term effects of the development of chronic metabolic diseases such as type 2 diabetes and obesity have been associated with nutritional insults in critical life stages. In this study, we evaluated the effect of a low-protein diet on metabolism in mid-adulthood male rats. At 90 days of age, Wistar male rats were fed a low-protein diet (4.0 %, LP group) for 30 days, whereas control rats were fed a normal-protein diet (20.5 %, NP group) throughout their lifetimes. To allow for dietary rehabilitation, from 120 to 180 days of age, the LP rats were fed a normal-protein diet. Then, we measured body composition, fat stores, glucose-insulin homeostasis and pancreatic islet function. At 120 days of age, just after low-protein diet treatment, the LP rats displayed a strong lean phenotype, hypoinsulinemia, as assessed under fasting and glucose tolerance test conditions, as well as weak pancreatic islet insulinotropic response to glucose and acetylcholine (p protein diet rehabilitation, the LP rats displayed a slight lean phenotype (p  0.05). Taken together, the present data suggest that the effects of dietary restriction as a stressor in adulthood are reversible with dietary rehabilitation, indicating that adulthood is not a sensitive or critical time window for metabolic programming.

  15. Pokemon siRNA Delivery Mediated by RGD-Modified HBV Core Protein Suppressed the Growth of Hepatocellular Carcinoma.

    Science.gov (United States)

    Kong, Jing; Liu, Xiaoping; Jia, Jianbo; Wu, Jinsheng; Wu, Ning; Chen, Jun; Fang, Fang

    2015-10-01

    Hepatocellular carcinoma (HCC) is a deadly human malignant tumor that is among the most common cancers in the world, especially in Asia. Hepatitis B virus (HBV) infection has been well established as a high risk factor for hepatic malignance. Studies have shown that Pokemon is a master oncogene for HCC growth, suggesting it as an ideal therapeutic target. However, efficient delivery system is still lacking for Pokemon targeting treatment. In this study, we used core proteins of HBV, which is modified with RGD peptides, to construct a biomimetic vector for the delivery of Pokemon siRNAs (namely, RGD-HBc-Pokemon siRNA). Quantitative PCR and Western blot assays revealed that RGD-HBc-Pokemon siRNA possessed the highest efficiency of Pokemon suppression in HCC cells. In vitro experiments further indicated that RGD-HBc-Pokemon-siRNA exerted a higher tumor suppressor activity on HCC cell lines, evidenced by reduced proliferation and attenuated invasiveness, than Pokemon-siRNA or RGD-HBc alone. Finally, animal studies demonstrated that RGD-HBc-Pokemon siRNA suppressed the growth of HCC xenografts in mice by a greater extent than Pokemon-siRNA or RGD-HBc alone. Based on the above results, Pokemon siRNA delivery mediated by RGD-modified HBV core protein was shown to be an effective strategy of HCC gene therapy.

  16. Human Cytomegalovirus Nuclear Capsids Associate with the Core Nuclear Egress Complex and the Viral Protein Kinase pUL97.

    Science.gov (United States)

    Milbradt, Jens; Sonntag, Eric; Wagner, Sabrina; Strojan, Hanife; Wangen, Christina; Lenac Rovis, Tihana; Lisnic, Berislav; Jonjic, Stipan; Sticht, Heinrich; Britt, William J; Schlötzer-Schrehardt, Ursula; Marschall, Manfred

    2018-01-13

    The nuclear phase of herpesvirus replication is regulated through the formation of regulatory multi-component protein complexes. Viral genomic replication is followed by nuclear capsid assembly, DNA encapsidation and nuclear egress. The latter has been studied intensely pointing to the formation of a viral core nuclear egress complex (NEC) that recruits a multimeric assembly of viral and cellular factors for the reorganization of the nuclear envelope. To date, the mechanism of the association of human cytomegalovirus (HCMV) capsids with the NEC, which in turn initiates the specific steps of nuclear capsid budding, remains undefined. Here, we provide electron microscopy-based data demonstrating the association of both nuclear capsids and NEC proteins at nuclear lamina budding sites. Specifically, immunogold labelling of the core NEC constituent pUL53 and NEC-associated viral kinase pUL97 suggested an intranuclear NEC-capsid interaction. Staining patterns with phospho-specific lamin A/C antibodies are compatible with earlier postulates of targeted capsid egress at lamina-depleted areas. Important data were provided by co-immunoprecipitation and in vitro kinase analyses using lysates from HCMV-infected cells, nuclear fractions, or infectious virions. Data strongly suggest that nuclear capsids interact with pUL53 and pUL97. Combined, the findings support a refined concept of HCMV nuclear trafficking and NEC-capsid interaction.

  17. Human Cytomegalovirus Nuclear Capsids Associate with the Core Nuclear Egress Complex and the Viral Protein Kinase pUL97

    Directory of Open Access Journals (Sweden)

    Jens Milbradt

    2018-01-01

    Full Text Available The nuclear phase of herpesvirus replication is regulated through the formation of regulatory multi-component protein complexes. Viral genomic replication is followed by nuclear capsid assembly, DNA encapsidation and nuclear egress. The latter has been studied intensely pointing to the formation of a viral core nuclear egress complex (NEC that recruits a multimeric assembly of viral and cellular factors for the reorganization of the nuclear envelope. To date, the mechanism of the association of human cytomegalovirus (HCMV capsids with the NEC, which in turn initiates the specific steps of nuclear capsid budding, remains undefined. Here, we provide electron microscopy-based data demonstrating the association of both nuclear capsids and NEC proteins at nuclear lamina budding sites. Specifically, immunogold labelling of the core NEC constituent pUL53 and NEC-associated viral kinase pUL97 suggested an intranuclear NEC-capsid interaction. Staining patterns with phospho-specific lamin A/C antibodies are compatible with earlier postulates of targeted capsid egress at lamina-depleted areas. Important data were provided by co-immunoprecipitation and in vitro kinase analyses using lysates from HCMV-infected cells, nuclear fractions, or infectious virions. Data strongly suggest that nuclear capsids interact with pUL53 and pUL97. Combined, the findings support a refined concept of HCMV nuclear trafficking and NEC-capsid interaction.

  18. Combined metabonomic and quantitative real-time PCR analyses reveal systems metabolic changes in Jurkat T-cells treated with HIV-1 Tat protein.

    Science.gov (United States)

    Liao, Wenting; Tan, Guangguo; Zhu, Zhenyu; Chen, Qiuli; Lou, Ziyang; Dong, Xin; Zhang, Wei; Pan, Wei; Chai, Yifeng

    2012-11-02

    HIV-1 Tat protein is released by infected cells and can affect bystander uninfected T cells and induce numerous biological responses which contribute to its pathogenesis. To elucidate the complex pathogenic mechanism, we conducted a comprehensive investigation on Tat protein-related extracellular and intracellular metabolic changes in Jurkat T-cells using combined gas chromatography-mass spectrometry (GC-MS), reversed-phase liquid chromatography-mass spectrometry (RPLC-MS) and a hydrophilic interaction liquid chromatography-mass spectrometry (HILIC-MS)-based metabonomics approach. Quantitative real-time PCR (qRT-PCR) analyses were further employed to measure expressions of several relevant enzymes together with perturbed metabolic pathways. Combined metabonomic and qRT-PCR analyses revealed that HIV-1 Tat caused significant and comprehensive metabolic changes, as represented by significant changes of 37 metabolites and 10 relevant enzymes in HIV-1 Tat-treated cells. Using MetaboAnalyst 2.0, it was found that 11 pathways (Impact-value >0.10) among the regulated pathways were acutely perturbed, including sphingolipid metabolism, glycine, serine and threonine metabolism, pyruvate metabolism, inositol phosphate metabolism, arginine and proline metabolism, citrate cycle, phenylalanine metabolism, tryptophan metabolism, pentose phosphate pathway, glycerophospholipid metabolism, glycolysis or gluconeogenesis. These results provide metabolic evidence of the complex pathogenic mechanism of HIV-1 Tat protein as a "viral toxin", and would help obligate Tat protein as "an important target" for therapeutic intervention and vaccine development.

  19. Influence of some simulated factors of cosmic flight on mineral and protein metabolism in the maxillodental system of rats

    International Nuclear Information System (INIS)

    Khristova, M.N.

    1985-01-01

    Mineral and protein metabolic changes in hard tissues of the maxillodental system, as opposed to skeletal bone, were studied in model experiments simulating space flight, with extreme factors given solely or combined: hypergravitation, ionizing radiation, magnetically activated water. Alterations occurring in protein synthesis proved to be the key events, whereas those in individual mineral components were either insignificant or of secondary nature. Long-term ingestion of magnetically activated water reduced radiation susceptibility of metabolic processes in mineralized tissues, but affected unfavorably the complex of signs resulting from combined exposure to extreme factors. Administration of a radioprotective drug, Adeturone, in treatments involving acceleration plus radiation led to normalization or partial balancing of metabolic processes in the mineral and organic components of maxillodental hard tissues and skeletal bone. The evidence obtained adds thus an anti-osteolathyrogenic feature to the radioprotective characteristics of Adeturone

  20. An extrahepatic receptor-associated protein-sensitive mechanism is involved in the metabolism of triglyceride-rich lipoproteins

    NARCIS (Netherlands)

    Vlijmen, B.J.M. van; Rohlmann, A.; Page, S.T.; Bensadoun, A.; Bos, I.S.T.; Berkel, T.J.C. van; Havekes, L.M.; Herz, J.

    1999-01-01

    We have used adenovirus-mediated gene transfer in mice to investigate low density lipoprotein receptor (LDLR) and LDLR-related protein (LRP)- independent mechanisms that control the metabolism of chylomicron and very low density lipoprotein (VLDL) remnants in vivo. Overexpression of receptor-

  1. Multi-omic profiling of EPO producing Chinese hamster ovary cell panel reveals metabolic adaptation to heterologous protein production

    DEFF Research Database (Denmark)

    Ley, Daniel; Kazemi Seresht, Ali; Engmark, Mikael

    Heterologous protein production in CHO cells imposes a burden on the host cell metabolism and impact cellular physiology on a global scale. In this work, a multi-omics approach was applied to characterize the physiological impact of erythropoietin production, and discover production bottlenecks, ...

  2. Hepatitis C virus core protein targets 4E-BP1 expression and phosphorylation and potentiates Myc-induced liver carcinogenesis in transgenic mice.

    Science.gov (United States)

    Abdallah, Cosette; Lejamtel, Charlène; Benzoubir, Nassima; Battaglia, Serena; Sidahmed-Adrar, Nazha; Desterke, Christophe; Lemasson, Matthieu; Rosenberg, Arielle R; Samuel, Didier; Bréchot, Christian; Pflieger, Delphine; Le Naour, François; Bourgeade, Marie-Françoise

    2017-08-22

    Hepatitis C virus (HCV) is a leading cause of liver diseases including the development of hepatocellular carcinoma (HCC). Particularly, core protein has been involved in HCV-related liver pathologies. However, the impact of HCV core on signaling pathways supporting the genesis of HCC remains largely elusive. To decipher the host cell signaling pathways involved in the oncogenic potential of HCV core, a global quantitative phosphoproteomic approach was carried out. This study shed light on novel differentially phosphorylated proteins, in particular several components involved in translation. Among the eukaryotic initiation factors that govern the translational machinery, 4E-BP1 represents a master regulator of protein synthesis that is associated with the development and progression of cancers due to its ability to increase protein expression of oncogenic pathways. Enhanced levels of 4E-BP1 in non-modified and phosphorylated forms were validated in human hepatoma cells and in mouse primary hepatocytes expressing HCV core, in the livers of HCV core transgenic mice as well as in HCV-infected human primary hepatocytes. The contribution of HCV core in carcinogenesis and the status of 4E-BP1 expression and phosphorylation were studied in HCV core/Myc double transgenic mice. HCV core increased the levels of 4E-BP1 expression and phosphorylation and significantly accelerated the onset of Myc-induced tumorigenesis in these double transgenic mice. These results reveal a novel function of HCV core in liver carcinogenesis potentiation. They position 4E-BP1 as a tumor-specific target of HCV core and support the involvement of the 4E-BP1/eIF4E axis in hepatocarcinogenesis.

  3. Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels

    Directory of Open Access Journals (Sweden)

    M. Ryan Smith

    2016-08-01

    Full Text Available Many cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes involved in metabolism possess a reactive thiolate group, which can be modified by certain oxidants. In the current study, we show that modification of mitochondrial protein thiols by a model compound, iodobutyl triphenylphosphonium (IBTP, decreased mitochondrial metabolism and ATP in MDA-MB 231 (MB231 breast adenocarcinoma cells up to 6 days after an initial 24 h treatment. Mitochondrial thiol modification also depressed oxygen consumption rates (OCR in a dose-dependent manner to a greater extent than a non-thiol modifying analog, suggesting that thiol reactivity is an important factor in the inhibition of cancer cell metabolism. In non-tumorigenic MCF-10A cells, IBTP also decreased OCR; however the extracellular acidification rate was significantly increased at all but the highest concentration (10 µM of IBTP indicating that thiol modification can have significantly different effects on bioenergetics in tumorigenic versus non-tumorigenic cells. ATP and other adenonucleotide levels were also decreased by thiol modification up to 6 days post-treatment, indicating a decreased overall energetic state in MB231 cells. Cellular proliferation of MB231 cells was also inhibited up to 6 days post-treatment with little change to cell viability. Targeted metabolomic analyses revealed that thiol modification caused depletion of both Krebs cycle and glutaminolysis intermediates. Further experiments revealed that the activity of the Krebs cycle enzyme, aconitase, was attenuated in response to thiol modification. Additionally, the inhibition of glutaminolysis corresponded to decreased glutaminase C (GAC protein levels, although other protein levels were

  4. Identifying molecular effects of diet through systems biology: influence of herring diet on sterol metabolism and protein turnover in mice.

    Directory of Open Access Journals (Sweden)

    Intawat Nookaew

    Full Text Available BACKGROUND: Changes in lifestyle have resulted in an epidemic development of obesity-related diseases that challenge the healthcare systems worldwide. To develop strategies to tackle this problem the focus is on diet to prevent the development of obesity-associated diseases such as cardiovascular disease (CVD. This will require methods for linking nutrient intake with specific metabolic processes in different tissues. METHODOLOGY/PRINCIPAL FINDING: Low-density lipoprotein receptor-deficient (Ldlr -/- mice were fed a high fat/high sugar diet to mimic a westernized diet, being a major reason for development of obesity and atherosclerosis. The diets were supplemented with either beef or herring, and matched in macronutrient contents. Body composition, plasma lipids and aortic lesion areas were measured. Transcriptomes of metabolically important tissues, e.g. liver, muscle and adipose tissue were analyzed by an integrated approach with metabolic networks to directly map the metabolic effects of diet in these different tissues. Our analysis revealed a reduction in sterol metabolism and protein turnover at the transcriptional level in herring-fed mice. CONCLUSION: This study shows that an integrated analysis of transcriptome data using metabolic networks resulted in the identification of signature pathways. This could not have been achieved using standard clustering methods. In particular, this systems biology analysis could enrich the information content of biomedical or nutritional data where subtle changes in several tissues together affects body metabolism or disease progression. This could be applied to improve diets for subjects exposed to health risks associated with obesity.

  5. Coupled cell-free synthesis, segregation, and core glycosylation of a secretory protein.

    Science.gov (United States)

    Lingappa, V R; Lingappa, J R; Prasad, R; Ebner, K E; Blobel, G

    1978-05-01

    mRNA from rat mammary glands 13-15 days post partum was translated in a wheat germ cell-free system either in the absence or in the presence of ribosome-denuded membranes prepared from isolated rough microsomes of dog pancreas. Newly synthesized alpha-lactalbumin was identified by immunoprecipitation with a monospecific rabbit antiserum against rat alpha-lactalbumin and was characterized by partial amino-terminal sequence determination and by lectin affinity chromatography. In the absence of membranes a presumably unglycosylated form of alpha-lactalbumin was synthesized that bound neither to concanavalin A-Sepharose nor to Ricinus communis lectin-agarose and that contained an amino-terminal signal peptide region comprising 19 amino acid residues. In the presence of membranes a processed form was synthesized that lacked the signal peptide portion and that had an amino-terminal sequence identical to that of mature alpha-lactalbumin. Furthermore, this processed form was found to be segregated, presumably within the microsomal vesicles, because it was resistant to post-translational proteolysis. It was also found to be glycosylated, and because it bound to concanavalin A-Sepharose, from which it could be eluted specifically by alpha-methyl mannoside, but not to R. communis lectin-agarose, it was presumably core-glycosylated. Processing, segregation, and core glycosylation were observed to proceed only when membranes were present during translation and not when they were added after translation.

  6. Fatty acid transport protein 1 regulates retinoid metabolism and photoreceptor development in mouse retina.

    Directory of Open Access Journals (Sweden)

    Aurélie Cubizolle

    Full Text Available In retinal pigment epithelium (RPE, RPE65 catalyzes the isomerization of all-trans-retinyl fatty acid esters to 11-cis-retinol in the visual cycle and controls the rhodopsin regeneration rate. However, the mechanisms by which these processes are regulated are still unclear. Fatty Acid Transport Protein 1 (FATP1 is involved in fatty acid uptake and lipid metabolism in a variety of cell types. FATP1 co-localizes with RPE65 in RPE and inhibits its isomerase activity in vitro. Here, we further investigated the role of FATP1 in the visual cycle using transgenic mice that overexpress human FATP1 specifically in the RPE (hFATP1TG mice. The mice displayed no delay in the kinetics of regeneration of the visual chromophore 11-cis-retinal after photobleaching and had no defects in light sensitivity. However, the total retinoid content was higher in the hFATP1TG mice than in wild type mice, and the transgenic mice also displayed an age-related accumulation (up to 40% of all-trans-retinal and retinyl esters that was not observed in control mice. Consistent with these results, hFATP1TG mice were more susceptible to light-induced photoreceptor degeneration. hFATP1 overexpression also induced an ~3.5-fold increase in retinosome autofluorescence, as measured by two-photon microscopy. Interestingly, hFATP1TG retina contained ~25% more photoreceptor cells and ~35% longer outer segments than wild type mice, revealing a non-cell-autonomous effect of hFATP1 expressed in the RPE. These data are the first to show that FATP1-mediated fatty acid uptake in the RPE controls both retinoid metabolism in the outer retina and photoreceptor development.

  7. Efficacy, Pharmacokinetics, and Metabolism of Tetrahydroquinoline Inhibitors of Plasmodium falciparum Protein Farnesyltransferase▿ †

    Science.gov (United States)

    Van Voorhis, Wesley C.; Rivas, Kasey L.; Bendale, Pravin; Nallan, Laxman; Hornéy, Carolyn; Barrett, Lynn K.; Bauer, Kevin D.; Smart, Brian P.; Ankala, Sudha; Hucke, Oliver; Verlinde, Christophe L. M. J.; Chakrabarti, Debopam; Strickland, Corey; Yokoyama, Kohei; Buckner, Frederick S.; Hamilton, Andrew D.; Williams, David K.; Lombardo, Louis J.; Floyd, David; Gelb, Michael H.

    2007-01-01

    New antimalarials are urgently needed. We have shown that tetrahydroquinoline (THQ) protein farnesyltransferase (PFT) inhibitors (PFTIs) are effective against the Plasmodium falciparum PFT and are effective at killing P. falciparum in vitro. Previously described THQ PFTIs had limitations of poor oral bioavailability and rapid clearance from the circulation of rodents. In this paper, we validate both the Caco-2 cell permeability model for predicting THQ intestinal absorption and the in vitro liver microsome model for predicting THQ clearance in vivo. Incremental improvements in efficacy, oral absorption, and clearance rate were monitored by in vitro tests; and these tests were followed up with in vivo absorption, distribution, metabolism, and excretion studies. One compound, PB-93, achieved cure when it was given orally to P. berghei-infected rats every 8 h for a total of 72 h. However, PB-93 was rapidly cleared, and dosing every 12 h failed to cure the rats. Thus, the in vivo results corroborate the in vitro pharmacodynamics and demonstrate that 72 h of continuous high-level exposure to PFTIs is necessary to kill plasmodia. The metabolism of PB-93 was demonstrated by a novel technique that relied on double labeling with a radiolabel and heavy isotopes combined with radiometric liquid chromatography and mass spectrometry. The major liver microsome metabolite of PB-93 has the PFT Zn-binding N-methyl-imidazole removed; this metabolite is inactive in blocking PFT function. By solving the X-ray crystal structure of PB-93 bound to rat PFT, a model of PB-93 bound to malarial PFT was constructed. This model suggests areas of the THQ PFTIs that can be modified to retain efficacy and protect the Zn-binding N-methyl-imidazole from dealkylation. PMID:17606674

  8. Intellectual disabilities, neuronal posttranscriptional RNA metabolism, and RNA-binding proteins: three actors for a complex scenario.

    Science.gov (United States)

    Bardoni, Barbara; Abekhoukh, Sabiha; Zongaro, Samantha; Melko, Mireille

    2012-01-01

    Intellectual disability (ID) is the most frequent cause of serious handicap in children and young adults and interests 2-3% of worldwide population, representing a serious problem from the medical, social, and economic points of view. The causes are very heterogeneous. Genes involved in ID have various functions altering different pathways important in neuronal function. Regulation of mRNA metabolism is particularly important in neurons for synaptic structure and function. Here, we review ID due to alteration of mRNA metabolism. Functional absence of some RNA-binding proteins--namely, FMRP, FMR2P, PQBP1, UFP3B, VCX-A--causes different forms of ID. These proteins are involved in different steps of RNA metabolism and, even if a detailed analysis of their RNA targets has been performed so far only for FMRP, it appears clear that they modulate some aspects (translation, stability, transport, and sublocalization) of a subset of RNAs coding for proteins, whose function must be relevant for neurons. Two other proteins, DYRK1A and CDKL5, involved in Down syndrome and Rett syndrome, respectively, have been shown to have an impact on splicing efficiency of specific mRNAs. Both proteins are kinases and their effect is indirect. Interestingly, both are localized in nuclear speckles, the nuclear domains where splicing factors are assembled, stocked, and recycled and influence their biogenesis and/or their organization. Copyright © 2012 Elsevier B.V. All rights reserved.

  9. Programming Post-Translational Control over the Metabolic Labeling of Cellular Proteins with a Noncanonical Amino Acid.

    Science.gov (United States)

    Thomas, Emily E; Pandey, Naresh; Knudsen, Sarah; Ball, Zachary T; Silberg, Jonathan J

    2017-08-18

    Transcriptional control can be used to program cells to label proteins with noncanonical amino acids by regulating the expression of orthogonal aminoacyl tRNA synthetases (aaRSs). However, we cannot yet program cells to control labeling in response to aaRS and ligand binding. To identify aaRSs whose activities can be regulated by interactions with ligands, we used a combinatorial approach to discover fragmented variants of Escherichia coli methionyl tRNA synthetase (MetRS) that require fusion to associating proteins for maximal activity. We found that these split proteins could be leveraged to create ligand-dependent MetRS using two approaches. When a pair of MetRS fragments was fused to FKBP12 and the FKBP-rapamycin binding domain (FRB) of mTOR and mutations were introduced that direct substrate specificity toward azidonorleucine (Anl), Anl metabolic labeling was significantly enhanced in growth medium containing rapamycin, which stabilizes the FKBP12-FRB complex. In addition, fusion of MetRS fragments to the termini of the ligand-binding domain of the estrogen receptor yielded proteins whose Anl metabolic labeling was significantly enhanced when 4-hydroxytamoxifen (4-HT) was added to the growth medium. These findings suggest that split MetRS can be fused to a range of ligand-binding proteins to create aaRSs whose metabolic labeling activities depend upon post-translational interactions with ligands.

  10. Role and metabolism of free leucine in skeletal muscle in protein sparing action of dietary carbohydrate and fat

    International Nuclear Information System (INIS)

    Nakano, Kiwao; Ishikawa, Tamotsu

    1977-01-01

    Feeding rats with either a carbohydrate meal or a fat meal to the previously fasted rats caused significant decrease in urinary output of urea and total nitrogen. The content of free leucine in skeletal muscle decreased in the rats fed either a carbohydrate meal or a fat meal. Feeding of either a carbohydrate meal or a fat meal stimulated incorporation of L-leucine-1- 14 C into protein fraction of skeletal muscle and reduced its oxidation to 14 CO 2 . These results suggest that the metabolism of leucine is under nutritional regulation and that the decrease in content of free leucine in skeletal muscle might be caused by enhanced reutilization of leucine into protein by the feeding of a carbohydrate meal or a fat meal. The role of free leucine in skeletal muscle as a regulator of protein turnover in the tissue are discussed in relation to the metabolism of this branched chain amino acid. (auth.)

  11. A Proteomic Approach for the Identification of Up-Regulated Proteins Involved in the Metabolic Process of the Leiomyoma.

    Science.gov (United States)

    Ura, Blendi; Scrimin, Federica; Arrigoni, Giorgio; Franchin, Cinzia; Monasta, Lorenzo; Ricci, Giuseppe

    2016-04-09

    Uterine leiomyoma is the most common benign smooth muscle cell tumor of the uterus. Proteomics is a powerful tool for the analysis of complex mixtures of proteins. In our study, we focused on proteins that were upregulated in the leiomyoma compared to the myometrium. Paired samples of eight leiomyomas and adjacent myometrium were obtained and submitted to two-dimensional gel electrophoresis (2-DE) and mass spectrometry for protein identification and to Western blotting for 2-DE data validation. The comparison between the patterns revealed 24 significantly upregulated (p leiomyoma and not with the normal myometrium. The overexpression of seven proteins involved in the metabolic processes of the leiomyoma was further validated by Western blotting and 2D Western blotting. Four of these proteins have never been associated with the leiomyoma before. The 2-DE approach coupled with mass spectrometry, which is among the methods of choice for comparative proteomic studies, identified a number of proteins overexpressed in the leiomyoma and involved in several biological processes, including metabolic processes. A better understanding of the mechanism underlying the overexpression of these proteins may be important for therapeutic purposes.

  12. Effects of immobilization and aerobic training on proteins related to intramuscular substrate storage and metabolism in young and older men.

    Science.gov (United States)

    Vigelsø, Andreas; Gram, Martin; Wiuff, Caroline; Hansen, Christina Neigaard; Prats, Clara; Dela, Flemming; Helge, Jørn Wulff

    2016-03-01

    Aging and inactivity lead to skeletal muscle metabolic inflexibility, but the underlying molecular mechanisms are not entirely elucidated. Therefore, we investigated how muscle lipid and glycogen stores and major regulatory proteins were affected by short-term immobilization followed by aerobic training in young and older men. 17 young (23 ± 1 years, 24 ± 1 kg m(-2), and 20 ± 2% body fat) and 15 older men (68 ± 1 years; 27 ± 1 kg m(-2), and 29 ± 2% body fat) underwent 2 weeks' one leg immobilization followed by 6 weeks' cycle training. Biopsies were obtained from m. vastus lateralis just before immobilization (at inclusion), after immobilization, and the after 6 weeks' training. The biopsies were analyzed for muscle substrates; muscle perilipin protein (PLIN), glycogen synthase (GS), synaptosomal-associated protein of 23 kDa (SNAP23) protein content, and muscle 3-hydroxyacyl-CoA dehydrogenase (HAD) activity The older men had higher intramuscular triglyceride (IMTG) (73 %) and Glycogen (16%) levels compared to the young men, and IMTG tended to increase with immobilization. PLIN2 and 3 protein content increased with immobilization in the older men only. The young men had higher GS (74%) protein compared to the older men. Immobilization decreased and training restored HAD activity, GS and SNAP23 protein content in young and older men. Evidence of age-related metabolic inflexibility is presented, seen as body fat and IMTG accumulation. The question arises as to whether IMTG accumulation in the older men is caused by or leading to the increase in PLIN2 and 3 protein content. Training decreased body fat and IMTG levels in both young and older men; hence, training should be prioritized to reduce the detrimental effect of aging on metabolism.

  13. The duck hepatitis B virus polymerase and core proteins accumulate in different patterns from their common mRNA

    International Nuclear Information System (INIS)

    Yao Ermei; Schaller, Heinz; Tavis, John E.

    2003-01-01

    Hepadnaviral reverse transcription occurs in capsids in which the core (C) protein surrounds the reverse transcriptase (P) and pregenomic RNA (pgRNA). We analyzed the accumulation patterns of duck hepatitis B virus P, C, and pgRNA in transfected LMH cells, infected primary duck hepatocytes (PDH), and infected duck liver. In all three systems, P accumulated over time in a different pattern compared with C, despite translation of both proteins from the pgRNA. Although the accumulation patterns of the proteins varied between the systems, in each case P became detectable at the same time or earlier than C and the ratio of P relative to C dropped with time. These accumulation patterns were consistent with the translation rates and half-lives of P and C. Comparing the translation rates of P and C with the pgRNA level over time revealed that translation of P and C was negatively regulated in LMH cells. These data provide a framework for comparing replication studies performed in LMH cells, PDHs and ducks

  14. Structure of the protein core of translation initiation factor 2 in apo, GTP-bound and GDP-bound forms

    International Nuclear Information System (INIS)

    Simonetti, Angelita; Marzi, Stefano; Fabbretti, Attilio; Hazemann, Isabelle; Jenner, Lasse; Urzhumtsev, Alexandre; Gualerzi, Claudio O.; Klaholz, Bruno P.

    2013-01-01

    The crystal structures of the eubacterial translation initiation factor 2 in apo form and with bound GDP and GTP reveal conformational changes upon nucleotide binding and hydrolysis, notably of the catalytically important histidine in the switch II region. Translation initiation factor 2 (IF2) is involved in the early steps of bacterial protein synthesis. It promotes the stabilization of the initiator tRNA on the 30S initiation complex (IC) and triggers GTP hydrolysis upon ribosomal subunit joining. While the structure of an archaeal homologue (a/eIF5B) is known, there are significant sequence and functional differences in eubacterial IF2, while the trimeric eukaryotic IF2 is completely unrelated. Here, the crystal structure of the apo IF2 protein core from Thermus thermophilus has been determined by MAD phasing and the structures of GTP and GDP complexes were also obtained. The IF2–GTP complex was trapped by soaking with GTP in the cryoprotectant. The structures revealed conformational changes of the protein upon nucleotide binding, in particular in the P-loop region, which extend to the functionally relevant switch II region. The latter carries a catalytically important and conserved histidine residue which is observed in different conformations in the GTP and GDP complexes. Overall, this work provides the first crystal structure of a eubacterial IF2 and suggests that activation of GTP hydrolysis may occur by a conformational repositioning of the histidine residue

  15. Structure of the protein core of translation initiation factor 2 in apo, GTP-bound and GDP-bound forms

    Energy Technology Data Exchange (ETDEWEB)

    Simonetti, Angelita [IGBMC (Institute of Genetics and of Molecular and Cellular Biology), Centre National de la Recherche Scientifique (CNRS) UMR 7104/Institut National de la Santé de la Recherche Médicale - INSERM U964/Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France); Marzi, Stefano [Architecture et Réactivité de l’ARN, UPR 9002 CNRS, IBMC (Institute of Molecular and Cellular Biology), 15 Rue R. Descartes, 67084 Strasbourg, France, Université de Strasbourg, 67000 Strasbourg (France); Fabbretti, Attilio [University of Camerino, 62032 Camerino (Monaco) (Italy); Hazemann, Isabelle; Jenner, Lasse [IGBMC (Institute of Genetics and of Molecular and Cellular Biology), Centre National de la Recherche Scientifique (CNRS) UMR 7104/Institut National de la Santé de la Recherche Médicale -INSERM U964/Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France); Urzhumtsev, Alexandre [IGBMC (Institute of Genetics and of Molecular and Cellular Biology), Centre National de la Recherche Scientifique (CNRS) UMR 7104/Institut National de la Santé de la Recherche Médicale - INSERM U964/Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France); Université de Lorraine, 54506 Vandoeuvre-lès-Nancy (France); Gualerzi, Claudio O. [University of Camerino, 62032 Camerino (Monaco) (Italy); Klaholz, Bruno P., E-mail: klaholz@igbmc.fr [IGBMC (Institute of Genetics and of Molecular and Cellular Biology), Centre National de la Recherche Scientifique (CNRS) UMR 7104/Institut National de la Santé de la Recherche Médicale - INSERM U964/Université de Strasbourg, 1 Rue Laurent Fries, 67404 Illkirch (France)

    2013-06-01

    The crystal structures of the eubacterial translation initiation factor 2 in apo form and with bound GDP and GTP reveal conformational changes upon nucleotide binding and hydrolysis, notably of the catalytically important histidine in the switch II region. Translation initiation factor 2 (IF2) is involved in the early steps of bacterial protein synthesis. It promotes the stabilization of the initiator tRNA on the 30S initiation complex (IC) and triggers GTP hydrolysis upon ribosomal subunit joining. While the structure of an archaeal homologue (a/eIF5B) is known, there are significant sequence and functional differences in eubacterial IF2, while the trimeric eukaryotic IF2 is completely unrelated. Here, the crystal structure of the apo IF2 protein core from Thermus thermophilus has been determined by MAD phasing and the structures of GTP and GDP complexes were also obtained. The IF2–GTP complex was trapped by soaking with GTP in the cryoprotectant. The structures revealed conformational changes of the protein upon nucleotide binding, in particular in the P-loop region, which extend to the functionally relevant switch II region. The latter carries a catalytically important and conserved histidine residue which is observed in different conformations in the GTP and GDP complexes. Overall, this work provides the first crystal structure of a eubacterial IF2 and suggests that activation of GTP hydrolysis may occur by a conformational repositioning of the histidine residue.

  16. Regulation of Brain Glucose Metabolic Patterns by Protein Phosphorlyation and Drug Therapy

    Science.gov (United States)

    2007-03-30

    Tymoczko et al. 2002). Both cardiac muscle and brain contain the necessary enzymes to metabolize either glucose or ketone bodies . The enzymes... metabolic phenotype of astrocytes and neurons in vitro; and to determine whether antipsychotic drug administration affects glucose metabolites in...Cortical Astrocytes and Neurons 20 Abstract 21 v Introduction ~ 22 Results 24 Enriched Astrocyte and Neuronal Cultures Display Unique Metabolic

  17. Insulin responsiveness of protein metabolism in vivo following bedrest in humans

    International Nuclear Information System (INIS)

    Shangraw, R.E.; Stuart, C.A.; Prince, M.J.; Peters, E.J.; Wolfe, R.R.

    1988-01-01

    To test the influence of bedrest on insulin regulation of leucine metabolism, six normal young men were subjected to a five-step hyperinsulinemic euglycemic clamp before and after 7 days of strict bedrest. A primed-constant infusion of [1- 13 C]leucine was used. Before bedrest, the basal rate of appearance (R a ) of intracellular leucine and leucine oxidation were 2.79±0.17 and 0.613±0.070 μmol·kg -1 ·min -1 , respectively. Insulin caused a dose-dependent reduction of the intracellular leucine R a and leucine oxidation to a minimum of 1.64±0.08 and 0.322±0.039 μmol·kg -1 ·min -1 , respectively, in nonbedrested subjects. Insulin also caused a dose-dependent reduction of plasma leucine concentration. After bedrest, subjects exhibited decreased glucose tolerance and increased endogenous insulin secretion, but basal and insulin-suppressed intracellular leucine R a and leucine oxidation rates were not different from control. Magnetic resonance imaging of the back and lower extremities revealed a 1-4% decrease in muscle volume and a 2-5% increase in fat volume secondary to bedrest. Bedrest also resulted in a negative nitrogen balance as compared with the control period. Thus because negative nitrogen balance and skeletal muscle atrophy occurred in six rested subjects in the absence of changes in the two indices of protein breakdown used in this study, it seems likely that muscle protein synthesis was inhibited

  18. Hepatitis C virus core protein regulates p300/CBP co-activation function. Possible role in the regulation of NF-AT1 transcriptional activity

    International Nuclear Information System (INIS)

    Gomez-Gonzalo, Marta; Benedicto, Ignacio; Carretero, Marta; Lara-Pezzi, Enrique; Maldonado-Rodriguez, Alejandra; Moreno-Otero, Ricardo; Lai, Michael M.C.; Lopez-Cabrera, Manuel

    2004-01-01

    Hepatitis C virus (HCV) core is a viral structural protein; it also participates in some cellular processes, including transcriptional regulation. However, the mechanisms of core-mediated transcriptional regulation remain poorly understood. Oncogenic virus proteins often target p300/CBP, a known co-activator of a wide variety of transcription factors, to regulate the expression of cellular and viral genes. Here we demonstrate, for the first time, that HCV core protein interacts with p300/CBP and enhances both its acetyl-transferase and transcriptional activities. In addition, we demonstrate that nuclear core protein activates the NH 2 -terminal transcription activation domain (TAD) of NF-AT1 in a p300/CBP-dependent manner. We propose a model in which core protein regulates the co-activation function of p300/CBP and activates NF-AT1, and probably other p300/CBP-regulated transcription factors, by a novel mechanism involving the regulation of the acetylation state of histones and/or components of the transcriptional machinery

  19. Tandem fusion of hepatitis B core antigen allows assembly of virus-like particles in bacteria and plants with enhanced capacity to accommodate foreign proteins.

    Directory of Open Access Journals (Sweden)

    Hadrien Peyret

    Full Text Available The core protein of the hepatitis B virus, HBcAg, assembles into highly immunogenic virus-like particles (HBc VLPs when expressed in a variety of heterologous systems. Specifically, the major insertion region (MIR on the HBcAg protein allows the insertion of foreign sequences, which are then exposed on the tips of surface spike structures on the outside of the assembled particle. Here, we present a novel strategy which aids the display of whole proteins on the surface of HBc particles. This strategy, named tandem core, is based on the production of the HBcAg dimer as a single polypeptide chain by tandem fusion of two HBcAg open reading frames. This allows the insertion of large heterologous sequences in only one of the two MIRs in each spike, without compromising VLP formation. We present the use of tandem core technology in both plant and bacterial expression systems. The results show that tandem core particles can be produced with unmodified MIRs, or with one MIR in each tandem dimer modified to contain the entire sequence of GFP or of a camelid nanobody. Both inserted proteins are correctly folded and the nanobody fused to the surface of the tandem core particle (which we name tandibody retains the ability to bind to its cognate antigen. This technology paves the way for the display of natively folded proteins on the surface of HBc particles either through direct fusion or through non-covalent attachment via a nanobody.

  20. Aggravation of Irradiation Induced Impairment in Protein Metabolism in Albino Rate Subjected to Oral Injection of Kelthane Miticide

    International Nuclear Information System (INIS)

    Yousri, R.M.; Abu Ghadeer, A.R.M.; Abbady, M.M.; Helmy, A.S.; Abdallah, N.M.

    1998-01-01

    The combined effect of both whole body gamma radiation exposure and administration of organo chlorine miticide k elthane o n protein metabolism was investigated in male albino rats. Kelthane was orally given at a dose level of 100 mg/kg body weight over a period of seven days. Irradiation process permitted the rats to receive one Gray every other day at a weekly cumulative dose of 3 Gy up to a total dose of 15 Gy. The biochemical assays included total proteins, protein fractions, free amino acids (FAAS) and urea level in blood serum as well as protein content and its FAAS in urine . The data revealed significant changes in the protein parameters due to whole body gamma irradiation. These changes were shown to be dose and time dependent which reached their maximum at the end of the experimentation period. The alterations were more pronounced in animal groups exposed to gamma radiation and received keltane pesticide

  1. Differential regulation of lipid and protein metabolism in obese vs. lean subjects before and after a 72-h fast

    DEFF Research Database (Denmark)

    Bak, Ann Mosegaard; Møller, Andreas Buch; Vendelbo, Mikkel Holm

    2016-01-01

    release in obese subjects under basal and fasting conditions. We therefore studied nine lean and nine obese subjects twice, after 12 and 72 h of fasting, using measurements of mRNA and protein expression and phosphorylation of lipolytic and protein metabolic signaling molecules in fat and muscle together...... with whole body and forearm tracer techniques. Obese subjects displayed increased whole body lipolysis, decreased urea production rates, and decreased forearm muscle protein breakdown per 100 ml of forearm tissue, differences that persisted after 72 h of fasting. Lipolysis per fat mass unit was reduced...... in obese subjects and, correspondingly, adipose tissue hormone-sensitive lipase (HSL) phosphorylation and mRNA and protein levels of the adipose triglyceride lipase (ATGL) coactivator CGI58 were decreased. Fasting resulted in higher HSL phosphorylations and lower protein levels of the ATGL inhibitor G0S2...

  2. Granzyme B Disrupts Central Metabolism and Protein Synthesis in Bacteria to Promote an Immune Cell Death Program.

    Science.gov (United States)

    Dotiwala, Farokh; Sen Santara, Sumit; Binker-Cosen, Andres Ariel; Li, Bo; Chandrasekaran, Sriram; Lieberman, Judy

    2017-11-16

    Human cytotoxic lymphocytes kill intracellular microbes. The cytotoxic granule granzyme proteases released by cytotoxic lymphocytes trigger oxidative bacterial death by disrupting electron transport, generating superoxide anion and inactivating bacterial oxidative defenses. However, they also cause non-oxidative cell death because anaerobic bacteria are also killed. Here, we use differential proteomics to identify granzyme B substrates in three unrelated bacteria: Escherichia coli, Listeria monocytogenes, and Mycobacteria tuberculosis. Granzyme B cleaves a highly conserved set of proteins in all three bacteria, which function in vital biosynthetic and metabolic pathways that are critical for bacterial survival under diverse environmental conditions. Key proteins required for protein synthesis, folding, and degradation are also substrates, including multiple aminoacyl tRNA synthetases, ribosomal proteins, protein chaperones, and the Clp system. Because killer cells use a multipronged strategy to target vital pathways, bacteria may not easily become resistant to killer cell attack. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Effects of immobilization and aerobic training on proteins related to intramuscular substrate storage and metabolism in young and older men

    DEFF Research Database (Denmark)

    Vigelsø Hansen, Andreas; Gram, Martin; Wiuff, Caroline

    2016-01-01

    by aerobic training in young and older men. METHODS: 17 young (23 ± 1 years, 24 ± 1 kg m(-2), and 20 ± 2% body fat) and 15 older men (68 ± 1 years; 27 ± 1 kg m(-2), and 29 ± 2% body fat) underwent 2 weeks' one leg immobilization followed by 6 weeks' cycle training. Biopsies were obtained from m. vastus...... lateralis just before immobilization (at inclusion), after immobilization, and the after 6 weeks' training. The biopsies were analyzed for muscle substrates; muscle perilipin protein (PLIN), glycogen synthase (GS), synaptosomal-associated protein of 23 kDa (SNAP23) protein content, and muscle 3-hydroxyacyl...... GS (74%) protein compared to the older men. Immobilization decreased and training restored HAD activity, GS and SNAP23 protein content in young and older men. CONCLUSION: Evidence of age-related metabolic inflexibility is presented, seen as body fat and IMTG accumulation. The question arises...

  4. Genetic Disruption of Protein Kinase STK25 Ameliorates Metabolic Defects in a Diet-Induced Type 2 Diabetes Model

    DEFF Research Database (Denmark)

    Amrutkar, Manoj; Cansby, Emmelie; Chursa, Urszula

    2015-01-01

    Understanding the molecular networks controlling ectopic lipid deposition, glucose tolerance, and insulin sensitivity is essential to identifying new pharmacological approaches to treat type 2 diabetes. We recently identified serine/threonine protein kinase 25 (STK25) as a negative regulator...... to the metabolic phenotype of Stk25 transgenic mice, reinforcing the validity of the results. The findings suggest that STK25 deficiency protects against the metabolic consequences of chronic exposure to dietary lipids and highlight the potential of STK25 antagonists for the treatment of type 2 diabetes....

  5. Hepatic metabolism of 11C-methionine and secretion of 11C-protein measured by PET in pigs

    DEFF Research Database (Denmark)

    Horsager, Jacob; Lausten, Susanne Bach; Bender, Dirk

    2017-01-01

    Hepatic amino acid metabolism and protein secretion are essential liver functions that may be altered during metabolic stress, e.g. after surgery. We wished to develop a dynamic liver PET method using the radiolabeled amino acid 11C-methionine to examine this question. Eleven 40-kg pigs were...... allocated to either laparotomy or pneumoperitoneum. 24 hours after surgery a 70-min dynamic PET scanning of the liver with arterial blood sampling was performed immediately after intravenous injection of 11C-methionine. Time course of arterial plasma 11C-methionine concentration was used as input function...

  6. Huntingtin Protein is Essential for Mitochondrial Metabolism, Bioenergetics and Structure in Murine Embryonic Stem Cells

    Science.gov (United States)

    Ismailoglu, Ismail; Chen, Qiuying; Popowski, Melissa; Yang, Lili; Gross, Steven S.; Brivanlou, Ali H.

    2014-01-01

    Mutations in the Huntington locus (htt) have devastating consequences. Gain-of-poly-Q repeats in Htt protein causes Huntington's disease (HD), while htt-/- mutants display early embryonic lethality. Despite its importance, the function of Htt remains elusive. To address this, we compared more than 3,700 compounds in three syngeneic mouse embryonic stem cell (mESC) lines: htt-/-, extended poly-Q (Htt-Q140/7), and wildtype mESCs (Htt-Q7/7) using untargeted metabolite profiling. While Htt-Q140/7 cells, did not show major differences in cellular bioenergetics, we find extensive metabolic aberrations in htt-/- mESCs, including: (i) complete failure of ATP production despite preservation of the mitochondrial membrane potential; (ii) near-maximal glycolysis, with little or no glycolytic reserve; (iii) marked ketogenesis; (iv) depletion of intracellular NTPs; (v) accelerated purine biosynthesis and salvage; and (vi) loss of mitochondrial structural integrity. Together, our findings reveal that Htt is necessary for mitochondrial structure and function from the earliest stages of embryogenesis, providing a molecular explanation for htt-/- early embryonic lethality. PMID:24780625

  7. The role of thioredoxin h in protein metabolism during wheat (Triticum aestivum L.) seed germination.

    Science.gov (United States)

    Guo, Hongxiang; Wang, Shaoxin; Xu, Fangfang; Li, Yongchun; Ren, Jiangping; Wang, Xiang; Niu, Hongbin; Yin, Jun

    2013-06-01

    Thioredoxin h can regulate the redox environment in the cell and play an important role in the germination of cereals. In the present study, the thioredoxin s antisense transgenic wheat with down-regulation of thioredoxin h was used to study the role of thioredoxin h in protein metabolism during germination of wheat seeds, and to explore the mechanism of the thioredoxin s antisense transgenic wheat seeds having high resistance to pre-harvest sprouting. The qRT-PCR results showed that the expression of protein disulfide isomerase in the thioredoxin s antisense transgenic wheat was up-regulated, which induced easily forming glutenin macropolymers and the resistance of storage proteins to degradation. The expression of serine protease inhibitor was also up-regulated in transgenic wheat, which might be responsible for the decreased activity of thiocalsin during the germination. The expression of WRKY6 in transgenic wheat was down-regulated, which was consistent with the decreased activity of glutamine oxoglutarate aminotransferase. In transgenic wheat, the activities of glutamate dehydrogenase, glutamic pyruvic transaminase and glutamic oxaloacetic transaminase were down-regulated, indicating that the metabolism of amino acid was lower than that in wild-type wheat during seed germination. A putative model for the role of thioredoxin h in protein metabolism during wheat seed germination was proposed and discussed. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  8. BET Bromodomain Proteins Brd2, Brd3 and Brd4 Selectively Regulate Metabolic Pathways in the Pancreatic β-Cell.

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    Jude T Deeney

    Full Text Available Displacement of Bromodomain and Extra-Terminal (BET proteins from chromatin has promise for cancer and inflammatory disease treatments, but roles of BET proteins in metabolic disease remain unexplored. Small molecule BET inhibitors, such as JQ1, block BET protein binding to acetylated lysines, but lack selectivity within the BET family (Brd2, Brd3, Brd4, Brdt, making it difficult to disentangle contributions of each family member to transcriptional and cellular outcomes. Here, we demonstrate multiple improvements in pancreatic β-cells upon BET inhibition with JQ1 or BET-specific siRNAs. JQ1 (50-400 nM increases insulin secretion from INS-1 cells in a concentration dependent manner. JQ1 increases insulin content in INS-1 cells, accounting for increased secretion, in both rat and human islets. Higher concentrations of JQ1 decrease intracellular triglyceride stores in INS-1 cells, a result of increased fatty acid oxidation. Specific inhibition of both Brd2 and Brd4 enhances insulin transcription, leading to increased insulin content. Inhibition of Brd2 alone increases fatty acid oxidation. Overlapping yet discrete roles for individual BET proteins in metabolic regulation suggest new isoform-selective BET inhibitors may be useful to treat insulin resistant/diabetic patients. Results imply that cancer and diseases of chronic inflammation or disordered metabolism are related through shared chromatin regulatory mechanisms.

  9. BET Bromodomain Proteins Brd2, Brd3 and Brd4 Selectively Regulate Metabolic Pathways in the Pancreatic β-Cell.

    Science.gov (United States)

    Deeney, Jude T; Belkina, Anna C; Shirihai, Orian S; Corkey, Barbara E; Denis, Gerald V

    2016-01-01

    Displacement of Bromodomain and Extra-Terminal (BET) proteins from chromatin has promise for cancer and inflammatory disease treatments, but roles of BET proteins in metabolic disease remain unexplored. Small molecule BET inhibitors, such as JQ1, block BET protein binding to acetylated lysines, but lack selectivity within the BET family (Brd2, Brd3, Brd4, Brdt), making it difficult to disentangle contributions of each family member to transcriptional and cellular outcomes. Here, we demonstrate multiple improvements in pancreatic β-cells upon BET inhibition with JQ1 or BET-specific siRNAs. JQ1 (50-400 nM) increases insulin secretion from INS-1 cells in a concentration dependent manner. JQ1 increases insulin content in INS-1 cells, accounting for increased secretion, in both rat and human islets. Higher concentrations of JQ1 decrease intracellular triglyceride stores in INS-1 cells, a result of increased fatty acid oxidation. Specific inhibition of both Brd2 and Brd4 enhances insulin transcription, leading to increased insulin content. Inhibition of Brd2 alone increases fatty acid oxidation. Overlapping yet discrete roles for individual BET proteins in metabolic regulation suggest new isoform-selective BET inhibitors may be useful to treat insulin resistant/diabetic patients. Results imply that cancer and diseases of chronic inflammation or disordered metabolism are related through shared chromatin regulatory mechanisms.

  10. High-sensitivity C-reactive protein predicts target organ damage in Chinese patients with metabolic syndrome

    DEFF Research Database (Denmark)

    Zhao, Zhigang; Nie, Hai; He, Hongbo

    2007-01-01

    with metabolic syndrome. A total of 1082 consecutive patients of Chinese origin were screened for the presence of metabolic syndrome according to the National Cholesterol Education Program's Adult Treatment Panel III. High-sensitivity C-reactive protein and target organ damage, including cardiac hypertrophy......Observational studies established high-sensitivity C-reactive protein as a risk factor for cardiovascular events in the general population. The goal of this study was to determine the relationship between target organ damage and high-sensitivity C-reactive protein in a cohort of Chinese patients......, carotid intima-media thickness, and renal impairment, were investigated. The median (25th and 75th percentiles) of high-sensitivity C-reactive protein in 619 patients with metabolic syndrome was 2.42 mg/L (0.75 and 3.66 mg/L) compared with 1.13 mg/L (0.51 and 2.46 mg/L) among 463 control subjects (P

  11. Treatment of metabolic syndrome by combination of physical activity and diet needs an optimal protein intake: a randomized controlled trial.

    Science.gov (United States)

    Dutheil, Frédéric; Lac, Gérard; Courteix, Daniel; Doré, Eric; Chapier, Robert; Roszyk, Laurence; Sapin, Vincent; Lesourd, Bruno

    2012-09-17

    The recommended dietary allowance (RDA) for protein intake has been set at 1.0-1.3 g/kg/day for senior. To date, no consensus exists on the lower threshold intake (LTI = RDA/1.3) for the protein intake (PI) needed in senior patients ongoing both combined caloric restriction and physical activity treatment for metabolic syndrome. Considering that age, caloric restriction and exercise are three increasing factors of protein need, this study was dedicated to determine the minimal PI in this situation, through the determination of albuminemia that is the blood marker of protein homeostasis. Twenty eight subjects (19 M, 9 F, 61.8 ± 6.5 years, BMI 33.4 ± 4.1 kg/m²) with metabolic syndrome completed a three-week residential programme (Day 0 to Day 21) controlled for nutrition (energy balance of -500 kcal/day) and physical activity (3.5 hours/day). Patients were randomly assigned in two groups: Normal-PI (NPI: 1.0 g/kg/day) and High-PI (HPI: 1.2 g/kg/day). Then, patients returned home and were followed for six months. Albuminemia was measured at D0, D21, D90 and D180. At baseline, PI was spontaneously 1.0 g/kg/day for both groups. Albuminemia was 40.6 g/l for NPI and 40.8 g/l for HPI. A marginal protein under-nutrition appeared in NPI with a decreased albuminemia at D90 below 35 g/l (34.3 versus 41.5 g/l for HPI, p treatment based on restricted diet and exercise in senior people with metabolic syndrome, the lower threshold intake for protein must be set at 1.2 g/kg/day to maintain blood protein homeostasis.

  12. Role of Decorin Core Protein in Collagen Organisation in Congenital Stromal Corneal Dystrophy (CSCD.

    Directory of Open Access Journals (Sweden)

    Christina S Kamma-Lorger

    Full Text Available The role of Decorin in organising the extracellular matrix was examined in normal human corneas and in corneas from patients with Congenital Stromal Corneal Dystrophy (CSCD. In CSCD, corneal clouding occurs due to a truncating mutation (c.967delT in the decorin (DCN gene. Normal human Decorin protein and the truncated one were reconstructed in silico using homology modelling techniques to explore structural changes in the diseased protein. Corneal CSCD specimens were also examined using 3-D electron tomography and Small Angle X-ray diffraction (SAXS, to image the collagen-proteoglycan arrangement and to quantify fibrillar diameters, respectively. Homology modelling showed that truncated Decorin had a different spatial geometry to the normal one, with the truncation removing a major part of the site that interacts with collagen, compromising its ability to bind effectively. Electron tomography showed regions of abnormal stroma, where collagen fibrils came together to form thicker fibrillar structures, showing that Decorin plays a key role in the maintenance of the order in the normal corneal extracellular matrix. Average diameter of individual fibrils throughout the thickness of the cornea however remained normal.

  13. The Effect of Vegan Protein-Based Diets on Metabolic Parameters, Expressions of Adiponectin and Its Receptors in Wistar Rats.

    Science.gov (United States)

    Chen, Jie-Hua; Song, Jia; Chen, Yan; Ding, Qiang; Peng, Anfang; Mao, Limei

    2016-10-18

    Vegan protein-based diet has attracted increasing interest in the prevention of metabolic syndrome (MetS). Meanwhile, adiponectin has become a highly potential molecular target in the prevention of MetS. Our study will identify a potential vegan protein diet for the prevention of MetS using rat models. Thirty-six Wistar rats were randomly assigned into three groups and given diets containing one of the following proteins for 12 weeks: casein (CAS, control diet), soy protein (SOY), and gluten-soy mixed protein (GSM). Changes in metabolic parameters as well as the expressions of adiponectin and its receptors were identified. Compared to CAS diet, both SOY and GSM diets led to decreases in blood total cholesterol and triglycerides, but only GSM diet led to an increase in HDL-cholesterol; no marked difference was observed in blood glucose in all three groups; HOMA-IR was found lower only in SOY group. Among groups, the order of serum adiponectin level was found as GSM > SOY > CAS. Similar order pattern was also observed in expression of adiponectin in adipose tissue and AdipoR1 mRNA in skeletal muscle. Our results suggested for the first time that, besides SOY diet, GSM diet could also be a possible substitute of animal protein to prevent MetS.

  14. The Effect of Vegan Protein-Based Diets on Metabolic Parameters, Expressions of Adiponectin and Its Receptors in Wistar Rats

    Directory of Open Access Journals (Sweden)

    Jie-Hua Chen

    2016-10-01

    Full Text Available Vegan protein-based diet has attracted increasing interest in the prevention of metabolic syndrome (MetS. Meanwhile, adiponectin has become a highly potential molecular target in the prevention of MetS. Our study will identify a potential vegan protein diet for the prevention of MetS using rat models. Thirty-six Wistar rats were randomly assigned into three groups and given diets containing one of the following proteins for 12 weeks: casein (CAS, control diet, soy protein (SOY, and gluten-soy mixed protein (GSM. Changes in metabolic parameters as well as the expressions of adiponectin and its receptors were identified. Compared to CAS diet, both SOY and GSM diets led to decreases in blood total cholesterol and triglycerides, but only GSM diet led to an increase in HDL-cholesterol; no marked difference was observed in blood glucose in all three groups; HOMA-IR was found lower only in SOY group. Among groups, the order of serum adiponectin level was found as GSM > SOY > CAS. Similar order pattern was also observed in expression of adiponectin in adipose tissue and AdipoR1 mRNA in skeletal muscle. Our results suggested for the first time that, besides SOY diet, GSM diet could also be a possible substitute of animal protein to prevent MetS.

  15. The diverse functions of the hepatitis B core/capsid protein (HBc) in the viral life cycle: Implications for the development of HBc-targeting antivirals.

    Science.gov (United States)

    Diab, Ahmed; Foca, Adrien; Zoulim, Fabien; Durantel, David; Andrisani, Ourania

    2018-01-01

    Virally encoded proteins have evolved to perform multiple functions, and the core protein (HBc) of the hepatitis B virus (HBV) is a perfect example. While HBc is the structural component of the viral nucleocapsid, additional novel functions for the nucleus-localized HBc have recently been described. These results extend for HBc, beyond its structural role, a regulatory function in the viral life cycle and potentially a role in pathogenesis. In this article, we review the diverse roles of HBc in HBV replication and pathogenesis, emphasizing how the unique structure of this protein is key to its various functions. We focus in particular on recent advances in understanding the significance of HBc phosphorylations, its interaction with host proteins and the role of HBc in regulating the transcription of host genes. We also briefly allude to the emerging niche for new direct-acting antivirals targeting HBc, known as Core (protein) Allosteric Modulators (CAMs). Copyright © 2017 Elsevier B.V. All rights reserved.

  16. A single amino acid substitution in the core protein of West Nile virus increases resistance to acidotropic compounds.

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    Miguel A Martín-Acebes

    Full Text Available West Nile virus (WNV is a worldwide distributed mosquito-borne flavivirus that naturally cycles between birds and mosquitoes, although it can infect multiple vertebrate hosts including horses and humans. This virus is responsible for recurrent epidemics of febrile illness and encephalitis, and has recently become a global concern. WNV requires to transit through intracellular acidic compartments at two different steps to complete its infectious cycle. These include fusion between the viral envelope and the membrane of endosomes during viral entry, and virus maturation in the trans-Golgi network. In this study, we followed a genetic approach to study the connections between viral components and acidic pH. A WNV mutant with increased resistance to the acidotropic compound NH4Cl, which blocks organelle acidification and inhibits WNV infection, was selected. Nucleotide sequencing revealed that this mutant displayed a single amino acid substitution (Lys 3 to Glu on the highly basic internal capsid or core (C protein. The functional role of this replacement was confirmed by its introduction into a WNV infectious clone. This single amino acid substitution also increased resistance to other acidification inhibitor (concanamycin A and induced a reduction of the neurovirulence in mice. Interestingly, a naturally occurring accompanying mutation found on prM protein abolished the resistant phenotype, supporting the idea of a genetic crosstalk between the internal C protein and the external glycoproteins of the virion. The findings here reported unveil a non-previously assessed connection between the C viral protein and the acidic pH necessary for entry and proper exit of flaviviruses.

  17. A single amino acid substitution in the core protein of West Nile virus increases resistance to acidotropic compounds.

    Science.gov (United States)

    Martín-Acebes, Miguel A; Blázquez, Ana-Belén; de Oya, Nereida Jiménez; Escribano-Romero, Estela; Shi, Pei-Yong; Saiz, Juan-Carlos

    2013-01-01

    West Nile virus (WNV) is a worldwide distributed mosquito-borne flavivirus that naturally cycles between birds and mosquitoes, although it can infect multiple vertebrate hosts including horses and humans. This virus is responsible for recurrent epidemics of febrile illness and encephalitis, and has recently become a global concern. WNV requires to transit through intracellular acidic compartments at two different steps to complete its infectious cycle. These include fusion between the viral envelope and the membrane of endosomes during viral entry, and virus maturation in the trans-Golgi network. In this study, we followed a genetic approach to study the connections between viral components and acidic pH. A WNV mutant with increased resistance to the acidotropic compound NH4Cl, which blocks organelle acidification and inhibits WNV infection, was selected. Nucleotide sequencing revealed that this mutant displayed a single amino acid substitution (Lys 3 to Glu) on the highly basic internal capsid or core (C) protein. The functional role of this replacement was confirmed by its introduction into a WNV infectious clone. This single amino acid substitution also increased resistance to other acidification inhibitor (concanamycin A) and induced a reduction of the neurovirulence in mice. Interestingly, a naturally occurring accompanying mutation found on prM protein abolished the resistant phenotype, supporting the idea of a genetic crosstalk between the internal C protein and the external glycoproteins of the virion. The findings here reported unveil a non-previously assessed connection between the C viral protein and the acidic pH necessary for entry and proper exit of flaviviruses.

  18. The dense core vesicle protein IA-2, but not IA-2β, is required for active avoidance learning.

    Science.gov (United States)

    Carmona, G N; Nishimura, T; Schindler, C W; Panlilio, L V; Notkins, A L

    2014-06-06

    The islet-antigens IA-2 and IA-2β are major autoantigens in type-1 diabetes and transmembrane proteins in dense core vesicles (DCV). Recently we showed that deletion of both IA-2 and IA-2β alters the secretion of hormones and neurotransmitters and impairs behavior and learning. The present study was designed to evaluate the contribution to learning of each of these genes by using single knockout (SKO) and double knockout (DKO) mice in an active avoidance test. After 5 days of training, wild-type (WT) mice showed 60-70% active avoidance responses, whereas the DKO mice showed only 10-15% active avoidance responses. The degree of active avoidance responses in the IA-2 SKO mice was similar to that of the DKO mice, but in contrast, the IA-2β SKO mice behaved like WT mice showing 60-70% active avoidance responses. Molecular studies revealed a marked decrease in the phosphorylation of the cAMP response element-binding protein (CREB) and Ca(2+)/calmodulin-dependent protein kinase II (CAMKII) in the striatum and hippocampus of the IA-2 SKO and DKO mice, but not in the IA-2β SKO mice. To evaluate the role of CREB and CAMKII in the SKO and DKO mice, GBR-12909, which selectively blocks the dopamine uptake transporter and increases CREB and CAMKII phosphorylation, was administered. GBR-12909 restored the phosphorylation of CREB and CAMKII and increased active avoidance learning in the DKO and IA-2 SKO to near the normal levels found in the WT and IA-2β SKO mice. We conclude that in the absence of the DCV protein IA-2, active avoidance learning is impaired. Published by Elsevier Ltd.

  19. Metabolic reprogramming through fatty acid transport protein 1 (FATP1 regulates macrophage inflammatory potential and adipose inflammation

    Directory of Open Access Journals (Sweden)

    Amy R. Johnson

    2016-07-01

    Full Text Available Objective: A novel approach to regulate obesity-associated adipose inflammation may be through metabolic reprogramming of macrophages (MΦs. Broadly speaking, MΦs dependent on glucose are pro-inflammatory, classically activated MΦs (CAM, which contribute to adipose inflammation and insulin resistance. In contrast, MΦs that primarily metabolize fatty acids are alternatively activated MΦs (AAM and maintain tissue insulin sensitivity. In actuality, there is much flexibility and overlap in the CAM-AAM spectrum in vivo dependent upon various stimuli in the microenvironment. We hypothesized that specific lipid trafficking proteins, e.g. fatty acid transport protein 1 (FATP1, would direct MΦ fatty acid transport and metabolism to limit inflammation and contribute to the maintenance of adipose tissue homeostasis. Methods: Bone marrow derived MΦs (BMDMs from Fatp1−/− and Fatp1+/+ mice were used to investigate FATP1-dependent substrate metabolism, bioenergetics, metabolomics, and inflammatory responses. We also generated C57BL/6J chimeric mice by bone marrow transplant specifically lacking hematopoetic FATP1 (Fatp1B−/− and controls Fatp1B+/+. Mice were challenged by high fat diet (HFD or low fat diet (LFD and analyses including MRI, glucose and insulin tolerance tests, flow cytometric, histologic, and protein quantification assays were conducted. Finally, an FATP1-overexpressing RAW 264.7 MΦ cell line (FATP1-OE and empty vector control (FATP1-EV were developed as a gain of function model to test effects on substrate metabolism, bioenergetics, metabolomics, and inflammatory responses. Results: Fatp1 is downregulated with pro-inflammatory stimulation of MΦs. Fatp1−/− BMDMs and FATP1-OE RAW 264.7 MΦs demonstrated that FATP1 reciprocally controled metabolic flexibility, i.e. lipid and glucose metabolism, which was associated with inflammatory response. Supporting our previous work demonstrating the positive relationship between glucose

  20. Sex difference in the association of metabolic syndrome with high sensitivity C-reactive protein in a Taiwanese population

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    Lin Wen-Yuan

    2010-07-01

    Full Text Available Abstract Background Although sex differences have been reported for associations between components of metabolic syndrome and inflammation, the question of whether there is an effect modification by sex in the association between inflammation and metabolic syndrome has not been investigated in detail. Therefore, the aim of this study was to compare associations of high sensitivity C-creative protein (hs-CRP with metabolic syndrome and its components between men and women. Methods A total of 1,305 subjects aged 40 years and over were recruited in 2004 in a metropolitan city in Taiwan. The biochemical indices, such as hs-CRP, fasting glucose levels, lipid profiles, urinary albumin, urinary creatinine and anthropometric indices, were measured. Metabolic syndrome was defined using the American Heart Association and the National Heart, lung and Blood Institute (AHA/NHLBI definition. The relationship between metabolic syndrome and hs-CRP was examined using multivariate logistic regression analysis. Results After adjustment for age and lifestyle factors including smoking, and alcohol intake, elevated concentrations of hs-CRP showed a stronger association with metabolic syndrome in women (odds ratio comparing tertile extremes 4.80 [95% CI: 3.31-6.97] than in men (2.30 [1.65-3.21]. The p value for the sex interaction was 0.002. All components were more strongly associated with metabolic syndrome in women than in men, and all sex interactions were significant except for hypertension. Conclusions Our data suggest that inflammatory processes may be of particular importance in the pathogenesis of metabolic syndrome in women.

  1. In vivo versus in vitro protein abundance analysis of Shigella dysenteriae type 1 reveals changes in the expression of proteins involved in virulence, stress and energy metabolism

    Directory of Open Access Journals (Sweden)

    Donohue-Rolfe Arthur

    2011-06-01

    Full Text Available Abstract Background Shigella dysenteriae serotype 1 (SD1 causes the most severe form of epidemic bacillary dysentery. Quantitative proteome profiling of Shigella dysenteriae serotype 1 (SD1 in vitro (derived from LB cell cultures and in vivo (derived from gnotobiotic piglets was performed by 2D-LC-MS/MS and APEX, a label-free computationally modified spectral counting methodology. Results Overall, 1761 proteins were quantitated at a 5% FDR (false discovery rate, including 1480 and 1505 from in vitro and in vivo samples, respectively. Identification of 350 cytoplasmic membrane and outer membrane (OM proteins (38% of in silico predicted SD1 membrane proteome contributed to the most extensive survey of the Shigella membrane proteome reported so far. Differential protein abundance analysis using statistical tests revealed that SD1 cells switched to an anaerobic energy metabolism under in vivo conditions, resulting in an increase in fermentative, propanoate, butanoate and nitrate metabolism. Abundance increases of transcription activators FNR and Nar supported the notion of a switch from aerobic to anaerobic respiration in the host gut environment. High in vivo abundances of proteins involved in acid resistance (GadB, AdiA and mixed acid fermentation (PflA/PflB indicated bacterial survival responses to acid stress, while increased abundance of oxidative stress proteins (YfiD/YfiF/SodB implied that defense mechanisms against oxygen radicals were mobilized. Proteins involved in peptidoglycan turnover (MurB were increased, while β-barrel OM proteins (OmpA, OM lipoproteins (NlpD, chaperones involved in OM protein folding pathways (YraP, NlpB and lipopolysaccharide biosynthesis (Imp were decreased, suggesting unexpected modulations of the outer membrane/peptidoglycan layers in vivo. Several virulence proteins of the Mxi-Spa type III secretion system and invasion plasmid antigens (Ipa proteins required for invasion of colonic epithelial cells, and release

  2. The SNARE protein vti1a functions in dense-core vesicle biogenesis

    DEFF Research Database (Denmark)

    Walter, Alexander M; Kurps, Julia; de Wit, Heidi

    2014-01-01

    overlapping with syntaxin-6. Exocytosis is impaired in vti1a null cells, partly due to fewer Ca(2+)-channels at the plasma membrane, partly due to fewer vesicles of reduced size and synaptobrevin-2 content. In contrast, release kinetics and Ca(2+)-sensitivity remain unchanged, indicating that the final fusion......The SNARE protein vti1a is proposed to drive fusion of intracellular organelles, but recent data also implicated vti1a in exocytosis. Here we show that vti1a is absent from mature secretory vesicles in adrenal chromaffin cells, but localizes to a compartment near the trans-Golgi network, partially...... reaction leading to transmitter release is unperturbed. Additional deletion of the closest related SNARE, vti1b, does not exacerbate the vti1a phenotype, and vti1b null cells show no secretion defects, indicating that vti1b does not participate in exocytosis. Long-term re-expression of vti1a (days...

  3. Systematization of the protein sequence diversity in enzymes related to secondary metabolic pathways in plants, in the context of big data biology inspired by the KNApSAcK motorcycle database.

    Science.gov (United States)

    Ikeda, Shun; Abe, Takashi; Nakamura, Yukiko; Kibinge, Nelson; Hirai Morita, Aki; Nakatani, Atsushi; Ono, Naoaki; Ikemura, Toshimichi; Nakamura, Kensuke; Altaf-Ul-Amin, Md; Kanaya, Shigehiko

    2013-05-01

    Biology is increasingly becoming a data-intensive science with the recent progress of the omics fields, e.g. genomics, transcriptomics, proteomics and metabolomics. The species-metabolite relationship database, KNApSAcK Core, has been widely utilized and cited in metabolomics research, and chronological analysis of that research work has helped to reveal recent trends in metabolomics research. To meet the needs of these trends, the KNApSAcK database has been extended by incorporating a secondary metabolic pathway database called Motorcycle DB. We examined the enzyme sequence diversity related to secondary metabolism by means of batch-learning self-organizing maps (BL-SOMs). Initially, we constructed a map by using a big data matrix consisting of the frequencies of all possible dipeptides in the protein sequence segments of plants and bacteria. The enzyme sequence diversity of the secondary metabolic pathways was examined by identifying clusters of segments associated with certain enzyme groups in the resulting map. The extent of diversity of 15 secondary metabolic enzyme groups is discussed. Data-intensive approaches such as BL-SOM applied to big data matrices are needed for systematizing protein sequences. Handling big data has become an inevitable part of biology.

  4. The effect of simvastatin treatment on the amyloid precursor protein and brain cholesterol metabolism in patients with Alzheimer's disease

    DEFF Research Database (Denmark)

    Hoglund, K; Thelen, K M; Syversen, S

    2005-01-01

    During the last years, several clinical studies have been published trying to elucidate the effect of statin treatment on amyloid precursor protein (APP) processing and metabolism of brain cholesterol in Alzheimer's disease (AD) in humans. We present an open biochemical study where 19 patients...... with AD have been treated with simvastatin (20 mg/day) for 12 months. The aim was to further investigate the effect of simvastatin treatment on cerebrospinal fluid (CSF) biomarkers of APP processing, AD biomarkers as total tau and tau phosphorylated at threonine 181, brain cholesterol metabolism as well...... as on cognitive decline in patients with AD. Despite biochemical data suggesting that treatment with 20 mg/day of simvastatin for 12 months does affect the brain cholesterol metabolism, we did not find any change in CSF or plasma levels of beta-amyloid (Abeta)(1-42). However, by analysis of APP isoforms, we found...

  5. Quantitative Metabolomics and Instationary 13C-Metabolic Flux Analysis Reveals Impact of Recombinant Protein Production on Trehalose and Energy Metabolism in Pichia pastoris

    Directory of Open Access Journals (Sweden)

    Joel Jordà

    2014-05-01

    Full Text Available Pichia pastoris has been recognized as an effective host for recombinant protein production. In this work, we combine metabolomics and instationary 13C metabolic flux analysis (INST 13C-MFA using GC-MS and LC-MS/MS to evaluate the potential impact of the production of a Rhizopus oryzae lipase (Rol on P. pastoris central carbon metabolism. Higher oxygen uptake and CO2 production rates and slightly reduced biomass yield suggest an increased energy demand for the producing strain. This observation is further confirmed by 13C-based metabolic flux analysis. In particular, the flux through the methanol oxidation pathway and the TCA cycle was increased in the Rol-producing strain compared to the reference strain. Next to changes in the flux distribution, significant variations in intracellular metabolite concentrations were observed. Most notably, the pools of trehalose, which is related to cellular stress response, and xylose, which is linked to methanol assimilation, were significantly increased in the recombinant strain.

  6. Reconstruction of the yeast protein-protein interaction network involved in nutrient sensing and global metabolic regulation

    DEFF Research Database (Denmark)

    Nandy, Subir Kumar; Jouhten, Paula; Nielsen, Jens

    2010-01-01

    proteins. Despite the value of BioGRID for studying protein-protein interactions, there is a need for manual curation of these interactions in order to remove false positives. RESULTS: Here we describe an annotated reconstruction of the protein-protein interactions around four key nutrient......) and for all the interactions between them (edges). The annotated information is readily available utilizing the functionalities of network modelling tools such as Cytoscape and CellDesigner. CONCLUSIONS: The reported fully annotated interaction model serves as a platform for integrated systems biology studies...

  7. Effect of ethanol in utero on higher nervous activity and protein and lipid metabolism in the rat brain

    International Nuclear Information System (INIS)

    Zabbudovskii, A.L.; Zhulin, V.V.

    1985-01-01

    The authors study parameters of protein phosphorylation and glycoprotein and phospholipid synthesis in the neocortex and hippocampus of adult rats and compare the findings with the results of an investigation of formation and preservation of defensive conditioned reflexes. The pattern of changes in these metabolic parameters are studied in response to stress. For the biochemical tests, the animals were lightly anesthetized with ether and injected with a mixture of (P 32)-orthophosphate and (H 3)-fucose. Phospholipids were identified with molybdate reagent and radioactivity of the protein digest and lipids was measured in Bray's scintillator. The study shows that the use of stress brought metabolic differences between the brain of the experimental and control rats more clearly to light

  8. Different intracellular distribution of avian reovirus core protein sigmaA in cells of avian and mammalian origin

    International Nuclear Information System (INIS)

    Vázquez-Iglesias, Lorena; Lostalé-Seijo, Irene; Martínez-Costas, José; Benavente, Javier

    2012-01-01

    A comparative analysis of the intracellular distribution of avian reovirus (ARV) core protein sigmaA in cells of avian and mammalian origin revealed that, whereas the viral protein accumulates in the cytoplasm and nucleolus of avian cells, most sigmaA concentrates in the nucleoplasm of mammalian cells in tight association with the insoluble nuclear matrix fraction. Our results further showed that sigmaA becomes arrested in the nucleoplasm of mammalian cells via association with mammalian cell-specific factors and that this association prevents nucleolar targeting. Inhibition of RNA polymerase II activity, but not of RNA polymerase I activity, in infected mammalian cells induces nucleus-to-cytoplasm sigmaA translocation through a CRM1- and RanGTP-dependent mechanism, yet a heterokaryon assay suggests that sigmaA does not shuttle between the nucleus and cytoplasm. The scarcity of sigmaA in cytoplasmic viral factories of infected mammalian cells could be one of the factors contributing to limited ARV replication in mammalian cells.

  9. Different intracellular distribution of avian reovirus core protein sigmaA in cells of avian and mammalian origin

    Energy Technology Data Exchange (ETDEWEB)

    Vazquez-Iglesias, Lorena; Lostale-Seijo, Irene; Martinez-Costas, Jose [Departamento de Bioquimica y Biologia Molecular, Facultad de Farmacia, y Centro Singular de Investigacion en Quimica Biologica y Materiales Moleculares (CIQUS), Universidad de Santiago de Compostela, 15782-Santiago de Compostela (Spain); Benavente, Javier, E-mail: franciscojavier.benavente@usc.es [Departamento de Bioquimica y Biologia Molecular, Facultad de Farmacia, y Centro Singular de Investigacion en Quimica Biologica y Materiales Moleculares (CIQUS), Universidad de Santiago de Compostela, 15782-Santiago de Compostela (Spain)

    2012-10-25

    A comparative analysis of the intracellular distribution of avian reovirus (ARV) core protein sigmaA in cells of avian and mammalian origin revealed that, whereas the viral protein accumulates in the cytoplasm and nucleolus of avian cells, most sigmaA concentrates in the nucleoplasm of mammalian cells in tight association with the insoluble nuclear matrix fraction. Our results further showed that sigmaA becomes arrested in the nucleoplasm of mammalian cells via association with mammalian cell-specific factors and that this association prevents nucleolar targeting. Inhibition of RNA polymerase II activity, but not of RNA polymerase I activity, in infected mammalian cells induces nucleus-to-cytoplasm sigmaA translocation through a CRM1- and RanGTP-dependent mechanism, yet a heterokaryon assay suggests that sigmaA does not shuttle between the nucleus and cytoplasm. The scarcity of sigmaA in cytoplasmic viral factories of infected mammalian cells could be one of the factors contributing to limited ARV replication in mammalian cells.

  10. Coevolved Mutations Reveal Distinct Architectures for Two Core Proteins in the Bacterial Flagellar Motor.

    Directory of Open Access Journals (Sweden)

    Alessandro Pandini

    Full Text Available Switching of bacterial flagellar rotation is caused by large domain movements of the FliG protein triggered by binding of the signal protein CheY to FliM. FliG and FliM form adjacent multi-subunit arrays within the basal body C-ring. The movements alter the interaction of the FliG C-terminal (FliGC "torque" helix with the stator complexes. Atomic models based on the Salmonella entrovar C-ring electron microscopy reconstruction have implications for switching, but lack consensus on the relative locations of the FliG armadillo (ARM domains (amino-terminal (FliGN, middle (FliGM and FliGC as well as changes during chemotaxis. The generality of the Salmonella model is challenged by the variation in motor morphology and response between species. We studied coevolved residue mutations to determine the unifying elements of switch architecture. Residue interactions, measured by their coevolution, were formalized as a network, guided by structural data. Our measurements reveal a common design with dedicated switch and motor modules. The FliM middle domain (FliMM has extensive connectivity most simply explained by conserved intra and inter-subunit contacts. In contrast, FliG has patchy, complex architecture. Conserved structural motifs form interacting nodes in the coevolution network that wire FliMM to the FliGC C-terminal, four-helix motor module (C3-6. FliG C3-6 coevolution is organized around the torque helix, differently from other ARM domains. The nodes form separated, surface-proximal patches that are targeted by deleterious mutations as in other allosteric systems. The dominant node is formed by the EHPQ motif at the FliMMFliGM contact interface and adjacent helix residues at a central location within FliGM. The node interacts with nodes in the N-terminal FliGc α-helix triad (ARM-C and FliGN. ARM-C, separated from C3-6 by the MFVF motif, has poor intra-network connectivity consistent with its variable orientation revealed by structural data. ARM

  11. Exploring abiotic stress on asynchronous protein metabolism in single kernels of wheat studied by NMR spectroscopy and chemometrics

    DEFF Research Database (Denmark)

    Winning, H.; Viereck, N.; Wollenweber, B.

    2009-01-01

    at the vegetative growth stage had little effect on the parameters investigated. For the first time, H-1 HR-MAS NMR spectra of grains taken during grain-filling were analysed by an advanced multiway model. In addition to the results from the chemical protein analysis and the H-1 HR-MAS NMR spectra of single kernels...... was to examine the implications of different drought treatments on the protein fractions in grains of winter wheat using H-1 nuclear magnetic resonance spectroscopy followed by chemometric analysis. Triticum aestivum L. cv. Vinjett was studied in a semi-field experiment and subjected to drought episodes either...... at terminal spikelet, during grain-filling or at both stages. Principal component trajectories of the total protein content and the protein fractions of flour as well as the H-1 NMR spectra of single wheat kernels, wheat flour, and wheat methanol extracts were analysed to elucidate the metabolic development...

  12. Disruption of the acyl-coa binding protein gene delays hepatic adaptation to metabolic changes at weaning

    DEFF Research Database (Denmark)

    Neess, Ditte; Bloksgaard, Maria; Sørensen, Signe Bek

    2011-01-01

    The acyl-CoA binding protein/diazepam binding inhibitor (ACBP/DBI) is an intracellular protein that binds C14-C22 acyl-CoA esters and is thought to act as an acyl-CoA transporter. In vitro analyses have indicated that ACBP can transport acyl-CoA esters between different enzymatic systems; however....... The delayed induction of SREBP target genes around weaning is caused by a compromised processing and decreased expression of SREBP precursors leading to reduced binding of SREBP to target sites in chromatin. In conclusion, lack of ACBP interferes with the normal metabolic adaptation to weaning and leads...

  13. Structural Insight into the Core of CAD, the Multifunctional Protein Leading De Novo Pyrimidine Biosynthesis.

    Science.gov (United States)

    Moreno-Morcillo, María; Grande-García, Araceli; Ruiz-Ramos, Alba; Del Caño-Ochoa, Francisco; Boskovic, Jasminka; Ramón-Maiques, Santiago

    2017-06-06

    CAD, the multifunctional protein initiating and controlling de novo biosynthesis of pyrimidines in animals, self-assembles into ∼1.5 MDa hexamers. The structures of the dihydroorotase (DHO) and aspartate transcarbamoylase (ATC) domains of human CAD have been previously determined, but we lack information on how these domains associate and interact with the rest of CAD forming a multienzymatic unit. Here, we prove that a construct covering human DHO and ATC oligomerizes as a dimer of trimers and that this arrangement is conserved in CAD-like from fungi, which holds an inactive DHO-like domain. The crystal structures of the ATC trimer and DHO-like dimer from the fungus Chaetomium thermophilum confirm the similarity with the human CAD homologs. These results demonstrate that, despite being inactive, the fungal DHO-like domain has a conserved structural function. We propose a model that sets the DHO and ATC complex as the central element in the architecture of CAD. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Incorporating Protein Biosynthesis into the Saccharomyces cerevisiae Genome-scale Metabolic Model

    DEFF Research Database (Denmark)

    Olivares Hernandez, Roberto

    Based on stoichiometric biochemical equations that occur into the cell, the genome-scale metabolic models can quantify the metabolic fluxes, which are regarded as the final representation of the physiological state of the cell. For Saccharomyces Cerevisiae the genome scale model has been construc......Based on stoichiometric biochemical equations that occur into the cell, the genome-scale metabolic models can quantify the metabolic fluxes, which are regarded as the final representation of the physiological state of the cell. For Saccharomyces Cerevisiae the genome scale model has been...

  15. The effect of passive heating on heat shock protein 70 and interleukin-6: a possible treatment tool for metabolic diseases?

    OpenAIRE

    Faulkner, SH; Jackson, S; Fatania, G; Leicht, CA

    2017-01-01

    Exercise and physical activity remain the gold standard methods of enhancing and maintaining health and wellbeing. However, in populations that benefit most from exercise, adherence is often poor and alternatives to exercise are important to bring about health improvements. Recent work suggests a role for passive heating (PH) and heat shock proteins (HSP) in improving cardio-metabolic health. The aim of this study was to investigate the expression of HSP70 and IL-6 in response to either exerc...

  16. Effect of 21-day head down bed rest on urine proteins related to endothelium: Correlations with changes in carbohydrate metabolism

    Science.gov (United States)

    Kashirina, D.; Pastushkova, L.; Custaud, M. A.; Dobrokhotov, I.; Brzhozovsky, A.; Navasiolava, N.; Nosovsky, A.; Kononikhin, A.; Nikolaev, E.; Larina, I.

    2017-08-01

    We performed liquid chromatography-mass spectrometric study of the urine proteome in 8 healthy volunteers aged between 20 and 44 y.o. who have completed 21-day head-down bed rest. ANDSystem software which builds associative networks was used to identify the urinary proteins functionally related to the endothelium. We identified 7 endothelium-related biological processes, directly linked to 13 urine proteins. We performed manual annotation of the proteins which were the most important in terms of endothelial functions. Analysis of the correlations with biochemical variables revealed a positive correlation between fasting blood glucose and the following urine proteins: albumin, CD44 antigen, endothelial protein C receptor, mucin-1, osteopontin, receptor tyrosine kinase. As well, we found a positive correlation between HOMA-insulin resistance index and the following urine proteins: endothelial protein C receptor and syndecan-4. These results might suggest the involvement of above-mentioned proteins in glucose metabolism and their participation in the response to changes in blood glucose level.

  17. Aroclor 1254, a developmental neurotoxicant, alters energy metabolism- and intracellular signaling-associated protein networks in rat cerebellum and hippocampus

    Energy Technology Data Exchange (ETDEWEB)

    Kodavanti, Prasada Rao S., E-mail: kodavanti.prasada@epa.gov [Neurotoxicology Branch, NHEERL, ORD, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina (United States); Osorio, Cristina [Systems Proteomics Center, University of North Carolina at Chapel Hill, North Carolina (United States); Program on Molecular Biology and Biotechnology, University of North Carolina at Chapel Hill, North Carolina (United States); Royland, Joyce E.; Ramabhadran, Ram [Genetic and Cellular Toxicology Branch, NHEERL, ORD, U.S. Environmental Protection Agency, Research Triangle Park, North Carolina (United States); Alzate, Oscar [Department of Cellular and Developmental Biology, University of North Carolina at Chapel Hill, North Carolina (United States); Systems Proteomics Center, University of North Carolina at Chapel Hill, North Carolina (United States); Program on Molecular Biology and Biotechnology, University of North Carolina at Chapel Hill, North Carolina (United States)

    2011-11-15

    The vast literature on the mode of action of polychlorinated biphenyls (PCBs) indicates that PCBs are a unique model for understanding the mechanisms of toxicity of environmental mixtures of persistent chemicals. PCBs have been shown to adversely affect psychomotor function and learning and memory in humans. Although the molecular mechanisms for PCB effects are unclear, several studies indicate that the disruption of Ca{sup 2+}-mediated signal transduction plays significant roles in PCB-induced developmental neurotoxicity. Culminating events in signal transduction pathways include the regulation of gene and protein expression, which affects the growth and function of the nervous system. Our previous studies showed changes in gene expression related to signal transduction and neuronal growth. In this study, protein expression following developmental exposure to PCB is examined. Pregnant rats (Long Evans) were dosed with 0.0 or 6.0 mg/kg/day of Aroclor-1254 from gestation day 6 through postnatal day (PND) 21, and the cerebellum and hippocampus from PND14 animals were analyzed to determine Aroclor 1254-induced differential protein expression. Two proteins were found to be differentially expressed in the cerebellum following PCB exposure while 18 proteins were differentially expressed in the hippocampus. These proteins are related to energy metabolism in mitochondria (ATP synthase, sub unit {beta} (ATP5B), creatine kinase, and malate dehydrogenase), calcium signaling (voltage-dependent anion-selective channel protein 1 (VDAC1) and ryanodine receptor type II (RyR2)), and growth of the nervous system (dihydropyrimidinase-related protein 4 (DPYSL4), valosin-containing protein (VCP)). Results suggest that Aroclor 1254-like persistent chemicals may alter energy metabolism and intracellular signaling, which might result in developmental neurotoxicity. -- Highlights: Black-Right-Pointing-Pointer We performed brain proteomic analysis of rats exposed to the neurotoxicant

  18. Toxic influence of organophosphate, carbamate, and organochlorine pesticides on cellular metabolism of lipids, proteins, and carbohydrates: a systematic review.

    Science.gov (United States)

    Karami-Mohajeri, Somayyeh; Abdollahi, Mohammad

    2011-09-01

    Pesticides, including organophosphate (OP), organochlorine (OC), and carbamate (CB) compounds, are widely used in agricultural and indoor purposes. OP and CB act as acetyl cholinesterase (AChE) inhibitors that affect lots of organs such as peripheral and central nervous systems, muscles, liver, pancreas, and brain, whereas OC are neurotoxic involved in alteration of ion channels. There are several reports about metabolic disorders, hyperglycemia, and also oxidative stress in acute and chronic exposures to pesticides that are linked with diabetes and other metabolic disorders. In this respect, there are several in vitro and in vivo but few clinical studies about mechanism underlying these effects. Bibliographic databases were searched for the years 1963-2010 and resulted in 1652 articles. After elimination of duplicates or irrelevant papers, 204 papers were included and reviewed. Results indicated that OP and CB impair the enzymatic pathways involved in metabolism of carbohydrates, fats and protein within cytoplasm, mitochondria, and proxisomes. It is believed that OP and CB show this effect through inhibition of AChE or affecting target organs directly. OC mostly affect lipid metabolism in the adipose tissues and change glucose pathway in other cells. As a shared mechanism, all OP, CB and OC induce cellular oxidative stress via affecting mitochondrial function and therefore disrupt neuronal and hormonal status of the body. Establishing proper epidemiological studies to explore exact relationships between exposure levels to these pesticides and rate of resulted metabolic disorders in human will be helpful.

  19. Acetic acid activates the AMP-activated protein kinase signaling pathway to regulate lipid metabolism in bovine hepatocytes.

    Directory of Open Access Journals (Sweden)

    Xinwei Li

    Full Text Available The effect of acetic acid on hepatic lipid metabolism in ruminants differs significantly from that in monogastric animals. Therefore, the aim of this study was to investigate the regulation mechanism of acetic acid on the hepatic lipid metabolism in dairy cows. The AMP-activated protein kinase (AMPK signaling pathway plays a key role in regulating hepatic lipid metabolism. In vitro, bovine hepatocytes were cultured and treated with different concentrations of sodium acetate (neutralized acetic acid and BML-275 (an AMPKα inhibitor. Acetic acid consumed a large amount of ATP, resulting in an increase in AMPKα phosphorylation. The increase in AMPKα phosphorylation increased the expression and transcriptional activity of peroxisome proliferator-activated receptor α, which upregulated the expression of lipid oxidation genes, thereby increasing lipid oxidation in bovine hepatocytes. Furthermore, elevated AMPKα phosphorylation reduced the expression and transcriptional activity of the sterol regulatory element-binding protein 1c and the carbohydrate responsive element-binding protein, which reduced the expression of lipogenic genes, thereby decreasing lipid biosynthesis in bovine hepatocytes. In addition, activated AMPKα inhibited the activity of acetyl-CoA carboxylase. Consequently, the triglyceride content in the acetate-treated hepatocytes was significantly decreased. These results indicate that acetic acid activates the AMPKα signaling pathway to increase lipid oxidation and decrease lipid synthesis in bovine hepatocytes, thereby reducing liver fat accumulation in dairy cows.

  20. The effects of wet cupping on serum high-sensitivity C-reactive protein and heat shock protein 27 antibody titers in patients with metabolic syndrome.

    Science.gov (United States)

    Farahmand, Seyed Kazem; Gang, Li Zhi; Saghebi, Seyed Ahmad; Mohammadi, Maryam; Mohammadi, Shabnam; Mohammadi, Ghazaleh; Ferns, Gordan A; Ghanbarzadeh, Majid; Razmgah, Gholamreza Ghayour; Ramazani, Zahra; Ghayour-Mobarhan, Majid; Esmaily, Habibollah; Bahrami Taghanaki, Hamidreza; Azizi, Hoda

    2014-08-01

    It has previously been reported that increased level of serum heat shock proteins (Hsps) antibody in patients with metabolic syndrome. It is possible that the expression of Hsp and inflammatory markers can be affected by cupping and traditional Chinese medicine. There is a little data investigating the effects of cupping on markers of inflammation and Hsp proteins, hence, the objective of this study was evaluation of the effects of wet cupping on serum high-sensitivity C-reactive protein (hs-CRP) and Hsp27 antibody titers in patients with metabolic syndrome. Serum Hs-CRP and Hsp27 antibody titers were assessed in samples from 126 patients with metabolic syndrome (18-65 years of age) at baseline, and after 6 and 12 weeks after treatment. One hundred and twenty-six patients were randomly divided into the experimental group treated with wet cupping combined with dietary advice, and the control group treated with dietary advice alone using a random number table. Eight patients in case group and five subjects in control groups were excluded from the study. Data were analyzed using SPSS 15.0 software and a repeated measure ANCOVA. Serum hs-CRP titers did not change significantly between groups (p>0.05) and times (p=0.27). The same result was found for Hsp27 titers (p>0.05). Wet-cupping on the interscapular region has no effect on serum hs-CRP and Hsp27 patients with metabolic syndrome. Copyright © 2014 Elsevier Ltd. All rights reserved.

  1. HCV core protein-induced down-regulation of microRNA-152 promoted aberrant proliferation by regulating Wnt1 in HepG2 cells.

    Directory of Open Access Journals (Sweden)

    Shifeng Huang

    Full Text Available Hepatitis C virus (HCV has been reported to regulate cellular microRNAs (miRNAs. The HCV core protein is considered to be a potential oncoprotein in HCV-related hepatocellular carcinoma (HCV-HCC, but HCV core-regulated miRNAs are largely unknown. Our preliminary experiments revealed significant down-regulation of microRNA-152 (miR-152 by HCV core protein in HepG2 cells. Through target gene prediction softwares, Wnt1 was predicted to be a potential target of miR-152. The present study was initiated to investigate whether miR-152 is aberrantly regulated by the HCV core protein, and involved in the regulation of the aberrant proliferation of HCV-HCC cells.MiR-152 levels were examined by stem-loop real-time RT-PCR (SLqRT-PCR. Cell proliferation was analyzed by MTT and colony formation assay. Cell cycle analysis was performed by flow cytometry. Luciferase reporter assay was conducted to confirm miRNA-target association. Wnt1 expression was determined by real-time qPCR and Western blotting.HCV core protein significantly suppressed miR-152 expression, and led to significant Wnt1 up-regulation with a concomitant aberrantly promoted proliferation. Moreover, we validated that miR-152 inhibition promoted, while miR-152 mimics inhibited cell proliferation. Using, qRT-PCR and western blot, Wnt1 was demonstrated to be regulated by miR-152. Luciferase activity assay showed that while miR-152 mimics significantly reduced the luciferase activity by 83.76% (P<0.0001, miR-152 inhibitor showed no effect on luciferase reporter. Most notably, salvage expression of miR-152 after Ad-HCV core infection for 24 h almost totally reversed the proliferation-promoting effect of the HCV core protein, and meanwhile, reduced the expression of both Wnt1 mRNA and protein to basal levels.These findings provide important evidence that the reduced miR-152 expression by HCV core protein can indirectly lose an inhibitory effect on Wnt1, which might, at least partially lead to cell

  2. Temporal patterns of cardiac performance and genes encoding heat shock proteins and metabolic sensors of an intertidal limpet Cellana toreuma during sublethal heat stress.

    Science.gov (United States)

    Zhang, Shu; Han, Guo-dong; Dong, Yun-wei

    2014-04-01

    Intertidal invertebrates develop effective physiological adaptations to cope with the rapidly changing thermal environment in the intertidal zone. In the present study, the temporal patterns of heart rate, protein carbonyl groups, and genes encoding heat shock proteins (hsp70 and hsp90) and metabolic sensors (ampkα, ampkβ and sirt1) were measured to study the effect of sublethal heat stress on the cardiac function, oxidative stress, heat shock response and cellular metabolism of an intertidal limpet Cellana toreuma. All the physiological parameters are sensitive to temperature and duration of heat stress. Spearman correlation analysis revealed that the correlations between heart rate and levels of heat shock proteins mRNA and metabolic sensors mRNA were statistically significant. These results further suggest that cardiac function plays crucial roles in cellular energy metabolism and heat shock responses. The significant increase of protein carbonyl groups at 34°C after 4h exposure indicated that the failure of cardiac function and the increase of anaerobic metabolism partly leads to the increase of protein carbonyl groups. Generally, the physiological responses to heat stress are sensitive to temperature and are energy-consumptive, as indicated by the upregulation of metabolic sensors mRNA. However, the upregulation of heat shock proteins and metabolic sensors at the post-transcriptional level and related functions need to be confirmed in further experiments. Copyright © 2014 Elsevier Ltd. All rights reserved.

  3. Methodical investigation of the protein metabolism and of the bioenergetics of protein retention in growing animals. 1

    International Nuclear Information System (INIS)

    Schiemann, R.; Bock, H.D.; Keller, J.; Hoffmann, L.; Krawielitzki, K.; Klein, M.

    1983-01-01

    The influence of different protein levels in the feed (group R1 20%, R2 38% crude protein) and of different energy levels (group J1 low, J2 high energy level) on the composition of the carcass and the apparent half-life periods of the body proteins were determined in 4 groups of 15 male broiler chickens labelled with 15 NH 4 acetate. In all slaughtering phases the higher protein level resulted in a higher weight of the feathers, breast and leg muscles, higher amounts of N in all parts of the body and a higher percentage of feathers, breast and leg muscles of the total carcass than the lower protein level. Between 13 and 19% of the N in the carcass contributed to the feathers, 24-31% to the breast and leg muscles and 50-63% to the rest of the carcass. The relative quotas of the sum of breast and leg muscles in the carcass were higher for the low energy level than for the high energy level. There were no remarkable differences as to the protein content of the muscles in dependence on the energy level, the quota of sarcoplasmatic proteins, however, was higher on the high level in contrast to the low energy level, that of the myofibrillar proteins was lower. The apparent half-life period of the total body protein after normal protein supply was 22 days (group R1) and 14 after high protein supply. The energy levels in groups J1 and J2 had no significant influence on the half-life period of the total body protein. In the body fractions examined the apparent half-life periods were highest in the breast muscle and lowest in the rest of the carcass. The protein stored in the feathers did not undergo decomposition. The protein fractions 'sarcoplasmatic protein' and 'myofibrillar protein' of breast and leg muscle neither differed from one another nor from the respective total muscle fractions as regards their half-life period. (author)

  4. Identification of differentially expressed reproductive and metabolic proteins in the female abalone (Haliotis laevigata) gonad following artificial induction of spawning.

    Science.gov (United States)

    Mendoza-Porras, Omar; Botwright, Natasha A; Reverter, Antonio; Cook, Mathew T; Harris, James O; Wijffels, Gene; Colgrave, Michelle L

    2017-12-01

    Inefficient control of temperate abalone spawning prevents pair-wise breeding and production of abalone with highly marketable traits. Traditionally, abalone farmers have used a combination of UV irradiation and application of temperature gradients to the tank water to artificially induce spawning. Proteins are known to regulate crucial processes such as respiration, muscle contraction, feeding, growth and reproduction. Spawning as a pre-requisite of abalone reproduction is likely to be regulated, in part, by endogenous proteins. A first step in elucidating the mechanisms that regulate spawning is to identify which proteins are directly involved during spawning. The present study examined protein expression following traditional spawning induction in the Haliotis laevigata female. Gonads were collected from abalone in the following physiological states: (1) spawning; (2) post-spawning; and (3) failed-to-spawn. Differential protein abundance was initially assessed using two-dimensional difference in-gel electrophoresis coupled with mass spectrometry for protein identification. A number of reproductive proteins such as vitellogenin, vitelline envelope zona pellucida domain 29 and prohibitin, and metabolic proteins such as thioredoxin peroxidase, superoxide dismutase and heat shock proteins were identified. Differences in protein abundance levels between physiological states were further assessed using scheduled multiple reaction monitoring mass spectrometry. Positive associations were observed between the abundance of specific proteins, such as heat shock cognate 70 and peroxiredoxin 6, and the propensity or failure to spawn in abalone. These findings have contributed to better understand both the effects of oxidative and heat stress over abalone physiology and their influence on abalone spawning. Crown Copyright © 2016. Published by Elsevier Inc. All rights reserved.

  5. Intrauterine growth retarded progeny of pregnant sows fed high protein:low carbohydrate diet is related to metabolic energy deficit.

    Directory of Open Access Journals (Sweden)

    Cornelia C Metges

    Full Text Available High and low protein diets fed to pregnant adolescent sows led to intrauterine growth retardation (IUGR. To explore underlying mechanisms, sow plasma metabolite and hormone concentrations were analyzed during different pregnancy stages and correlated with litter weight (LW at birth, sow body weight and back fat thickness. Sows were fed diets with low (6.5%, LP, adequate (12.1%, AP, and high (30%, HP protein levels, made isoenergetic by adjusted carbohydrate content. At -5, 24, 66, and 108 days post coitum (dpc fasted blood was collected. At 92 dpc, diurnal metabolic profiles were determined. Fasted serum urea and plasma glucagon were higher due to the HP diet. High density lipoprotein cholesterol (HDLC, %HDLC and cortisol were reduced in HP compared with AP sows. Lowest concentrations were observed for serum urea and protein, plasma insulin-like growth factor-I, low density lipoprotein cholesterol, and progesterone in LP compared with AP and HP sows. Fasted plasma glucose, insulin and leptin concentrations were unchanged. Diurnal metabolic profiles showed lower glucose in HP sows whereas non-esterified fatty acids (NEFA concentrations were higher in HP compared with AP and LP sows. In HP and LP sows, urea concentrations were 300% and 60% of AP sows, respectively. Plasma total cholesterol was higher in LP than in AP and HP sows. In AP sows, LW correlated positively with insulin and insulin/glucose and negatively with glucagon/insulin at 66 dpc, whereas in HP sows LW associated positively with NEFA. In conclusion, IUGR in sows fed high protein:low carbohydrate diet was probably due to glucose and energy deficit whereas in sows with low protein:high carbohydrate diet it was possibly a response to a deficit of indispensable amino acids which impaired lipoprotein metabolism and favored maternal lipid disposal.

  6. Quantitative Comparison of Dense-Core Amyloid Plaque Accumulation in Amyloid-β Precursor Protein Transgenic Mice

    Science.gov (United States)

    Liu, Peng; Reichl, John H.; Rao, Eshaan R.; McNellis, Brittany M.; Huang, Eric S.; Hemmy, Laura S.; Forster, Colleen L.; Kuskowski, Michael A.; Borchelt, David R.; Vassar, Robert; Ashe, Karen H.; Zahs, Kathleen R.

    2016-01-01

    There exist several dozen lines of transgenic mice that express human amyloid-β precursor protein (AβPP) with Alzheimer’s disease (AD)-linked mutations. AβPP transgenic mouse lines differ in the types and amounts of Aβ that they generate and in their spatiotemporal patterns of expression of Aβ assemblies, providing a toolkit to study Aβ amyloidosis and the influence of Aβ aggregation on brain function. More complete quantitative descriptions of the types of Aβ assemblies present in transgenic mice and in humans during disease progression should add to our understanding of how Aβ toxicity in mice relates to the pathogenesis of AD. Here, we provide a direct quantitative comparison of amyloid plaque burdens and plaque sizes in four lines of AβPP transgenic mice. We measured the fraction of cortex and hippocampus occupied by dense-core plaques, visualized by staining with Thioflavin S, in mice from young adulthood through advanced age. We found that the plaque burdens among the transgenic lines varied by an order of magnitude: at 15 months of age, the oldest age studied, the median cortical plaque burden in 5XFAD mice was already ~4.5 times that of 21-month Tg2576 mice and ~15 times that of 21–24-month rTg9191 mice. Plaque-size distributions changed across the lifespan in a line- and region-dependent manner. We also compared the dense-core plaque burdens in the mice to those measured in a set of pathologically-confirmed AD cases from the Nun Study. Cortical plaque burdens in Tg2576, APPSwePS1ΔE9, and 5XFAD mice eventually far exceeded those measured in the human cohort. PMID:28059792

  7. Quantitative Comparison of Dense-Core Amyloid Plaque Accumulation in Amyloid-β Protein Precursor Transgenic Mice.

    Science.gov (United States)

    Liu, Peng; Reichl, John H; Rao, Eshaan R; McNellis, Brittany M; Huang, Eric S; Hemmy, Laura S; Forster, Colleen L; Kuskowski, Michael A; Borchelt, David R; Vassar, Robert; Ashe, Karen H; Zahs, Kathleen R

    2017-01-01

    There exist several dozen lines of transgenic mice that express human amyloid-β protein precursor (AβPP) with Alzheimer's disease (AD)-linked mutations. AβPP transgenic mouse lines differ in the types and amounts of Aβ that they generate and in their spatiotemporal patterns of expression of Aβ assemblies, providing a toolkit to study Aβ amyloidosis and the influence of Aβ aggregation on brain function. More complete quantitative descriptions of the types of Aβ assemblies present in transgenic mice and in humans during disease progression should add to our understanding of how Aβ toxicity in mice relates to the pathogenesis of AD. Here, we provide a direct quantitative comparison of amyloid plaque burdens and plaque sizes in four lines of AβPP transgenic mice. We measured the fraction of cortex and hippocampus occupied by dense-core plaques, visualized by staining with Thioflavin S, in mice from young adulthood through advanced age. We found that the plaque burdens among the transgenic lines varied by an order of magnitude: at 15 months of age, the oldest age studied, the median cortical plaque burden in 5XFAD mice was already ∼4.5 times that of 21-month-old Tg2576 mice and ∼15 times that of 21-24-month-old rTg9191 mice. Plaque-size distributions changed across the lifespan in a line- and region-dependent manner. We also compared the dense-core plaque burdens in the mice to those measured in a set of pathologically-confirmed AD cases from the Nun Study. Cortical plaque burdens in Tg2576, APPSwePS1ΔE9, and 5XFAD mice eventually far exceeded those measured in the human cohort.

  8. Metabolic labeling of cellular glycoproteins with glucosamine: potential for erroneous interpretations due to nonenzymatic radiolabeling of proteins

    International Nuclear Information System (INIS)

    Briles, E.I.B.; Updyke, T.V.

    1986-01-01

    Proteins, including serum proteins of culture media, become nonenzymatically radiolabeled under conditions used for metabolic labeling of cultured cells with glucosamine. This occurs even under sterile conditions in the absence of cells. Various commercial lots of 3 H or 14 C glcN gave similar results: ∼ 0.7% of total label was incorporated into 20% serum (14 mg/ml protein) in 48 h at 37 0 C. By SDS-PAGE fluorography, labeled serum bands correspond to Coomassie stained bands. Incorporation is linear with protein concentration and label input, shows biphasic kinetics (initial rapid rate within first 3 hr, followed by slower linear rate with no sign of saturation through 120 hr), and is temperature-dependent (no reaction at 0 0 C; incorporation at 20 0 C is ∼ 45% of that at 37 0 C). Poly-D-lysine is a better acceptor than protein: 0.5 mg/ml PL accepts as much label as 7 mg/ml protein. Incorporation is inhibited by excess unlabeled glcN and ethanolamine, but not by man, gal or glucose. However, when proteins were incubated with 160 mM glcN, SDS-PAGE bands were yellow-brown, suggesting the occurrence of Maillard-type reactions. Although the chemical mechanism(s) responsible for nonmetabolic radiolabeling by glcN are not clear at this point, the fact that it occurs represents a serious artifact which may lead to erroneous interpretation of data

  9. Glucokinase regulatory protein genetic variant interacts with omega-3 PUFA to influence insulin resistance and inflammation in metabolic syndrome.

    Directory of Open Access Journals (Sweden)

    Pablo Perez-Martinez

    Full Text Available Glucokinase Regulatory Protein (GCKR plays a central role regulating both hepatic triglyceride and glucose metabolism. Fatty acids are key metabolic regulators, which interact with genetic factors and influence glucose metabolism and other metabolic traits. Omega-3 polyunsaturated fatty acids (n-3 PUFA have been of considerable interest, due to their potential to reduce metabolic syndrome (MetS risk.To examine whether genetic variability at the GCKR gene locus was associated with the degree of insulin resistance, plasma concentrations of C-reactive protein (CRP and n-3 PUFA in MetS subjects.Homeostasis model assessment of insulin resistance (HOMA-IR, HOMA-B, plasma concentrations of C-peptide, CRP, fatty acid composition and the GCKR rs1260326-P446L polymorphism, were determined in a cross-sectional analysis of 379 subjects with MetS participating in the LIPGENE dietary cohort.Among subjects with n-3 PUFA levels below the population median, carriers of the common C/C genotype had higher plasma concentrations of fasting insulin (P = 0.019, C-peptide (P = 0.004, HOMA-IR (P = 0.008 and CRP (P = 0.032 as compared with subjects carrying the minor T-allele (Leu446. In contrast, homozygous C/C carriers with n-3 PUFA levels above the median showed lower plasma concentrations of fasting insulin, peptide C, HOMA-IR and CRP, as compared with individuals with the T-allele.We have demonstrated a significant interaction between the GCKR rs1260326-P446L polymorphism and plasma n-3 PUFA levels modulating insulin resistance and inflammatory markers in MetS subjects. Further studies are needed to confirm this gene-diet interaction in the general population and whether targeted dietary recommendations can prevent MetS in genetically susceptible individuals.ClinicalTrials.gov NCT00429195.

  10. Growth Patterns in the Irish Pyridoxine Nonresponsive Homocystinuria Population and the Influence of Metabolic Control and Protein Intake.

    Science.gov (United States)

    Purcell, Orla; Coughlan, Aoife; Grant, Tim; McNulty, Jenny; Clark, Anne; Deverell, Deirdre; Mayne, Philip; Hughes, Joanne; Monavari, Ahmad; Knerr, Ina; Crushell, Ellen

    2017-01-01

    A low methionine diet is the mainstay of treatment for pyridoxine nonresponsive homocystinuria (HCU). There are various guidelines for recommended protein intakes for HCU and clinical practice varies. Poor growth has been associated with low cystine levels. This retrospective review of 48 Irish pyridoxine nonresponsive HCU patients assessed weight, height, body mass index (BMI), protein intake, and metabolic control up to 18 years at nine set time points. Patients diagnosed through newborn screening (NBS) were compared to late diagnosed (LD) patients. At 18 years the LD group ( n = 12, mean age at diagnosis 5.09 years) were heavier (estimated effect +4.97 Kg, P = 0.0058) and taller (estimated effect +7.97 cm P = 0.0204) than the NBS group ( n = 36). There was no difference in growth rate between the groups after 10 years of age. The HCU population were heavier and taller than the general population by one standard deviation with no difference in BMI. There was no association between intermittently low cystine levels and height. Three protein intake guidelines were compared; there was no difference in adult height between those who met the lowest of the guidelines (Genetic Metabolic Dietitians International) and those with a higher protein intake.

  11. Control of Secreted Protein Gene Expression and the Mammalian Secretome by the Metabolic Regulator PGC-1α.

    Science.gov (United States)

    Minsky, Neri; Roeder, Robert G

    2017-01-06

    Secreted proteins serve pivotal roles in the development of multicellular organisms, acting as structural matrix, extracellular enzymes, and signal molecules. However, how the secretome is regulated remains incompletely understood. Here we demonstrate, unexpectedly, that peroxisome proliferator-activated receptor γ coactivator 1-α (PGC-1α), a critical transcriptional co-activator of metabolic gene expression, functions to down-regulate the expression of diverse genes encoding secreted molecules and extracellular matrix components to modulate the secretome. Using cell lines, primary cells, and mice, we show that both endogenous and exogenous PGC-1α down-regulate the expression of numerous genes encoding secreted molecules. Mechanistically, results obtained using mRNA stability measurements as well as intronic RNA expression analysis are consistent with a transcriptional effect of PGC-1α on the expression of genes encoding secreted proteins. Interestingly, PGC-1α requires the central heat shock response regulator heat shock factor protein 1 (HSF1) to affect some of its targets, and both factors co-reside on several target genes encoding secreted molecules in cells. Finally, using a mass spectrometric analysis of secreted proteins, we demonstrate that PGC-1α modulates the secretome of mouse embryonic fibroblasts. Our results define a link between a key pathway controlling metabolic regulation and the regulation of the mammalian secretome. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  12. Growth Patterns in the Irish Pyridoxine Nonresponsive Homocystinuria Population and the Influence of Metabolic Control and Protein Intake

    Directory of Open Access Journals (Sweden)

    Orla Purcell

    2017-01-01

    Full Text Available A low methionine diet is the mainstay of treatment for pyridoxine nonresponsive homocystinuria (HCU. There are various guidelines for recommended protein intakes for HCU and clinical practice varies. Poor growth has been associated with low cystine levels. This retrospective review of 48 Irish pyridoxine nonresponsive HCU patients assessed weight, height, body mass index (BMI, protein intake, and metabolic control up to 18 years at nine set time points. Patients diagnosed through newborn screening (NBS were compared to late diagnosed (LD patients. At 18 years the LD group (n=12, mean age at diagnosis 5.09 years were heavier (estimated effect +4.97 Kg, P=0.0058 and taller (estimated effect +7.97 cm P=0.0204 than the NBS group (n=36. There was no difference in growth rate between the groups after 10 years of age. The HCU population were heavier and taller than the general population by one standard deviation with no difference in BMI. There was no association between intermittently low cystine levels and height. Three protein intake guidelines were compared; there was no difference in adult height between those who met the lowest of the guidelines (Genetic Metabolic Dietitians International and those with a higher protein intake.

  13. An amino-terminal segment of hantavirus nucleocapsid protein presented on hepatitis B virus core particles induces a strong and highly cross-reactive antibody response in mice

    International Nuclear Information System (INIS)

    Geldmacher, Astrid; Skrastina, Dace; Petrovskis, Ivars; Borisova, Galina; Berriman, John A.; Roseman, Alan M.; Crowther, R. Anthony; Fischer, Jan; Musema, Shamil; Gelderblom, Hans R.; Lundkvist, Aake; Renhofa, Regina; Ose, Velta; Krueger, Detlev H.; Pumpens, Paul; Ulrich, Rainer

    2004-01-01

    Previously, we have demonstrated that hepatitis B virus (HBV) core particles tolerate the insertion of the amino-terminal 120 amino acids (aa) of the Puumala hantavirus nucleocapsid (N) protein. Here, we demonstrate that the insertion of 120 amino-terminal aa of N proteins from highly virulent Dobrava and Hantaan hantaviruses allows the formation of chimeric core particles. These particles expose the inserted foreign protein segments, at least in part, on their surface. Analysis by electron cryomicroscopy of chimeric particles harbouring the Puumala virus (PUUV) N segment revealed 90% T = 3 and 10% T = 4 shells. A map computed from T = 3 shells shows additional density splaying out from the tips of the spikes producing the effect of an extra shell of density at an outer radius compared with wild-type shells. The inserted Puumala virus N protein segment is flexibly linked to the core spikes and only partially icosahedrally ordered. Immunisation of mice of two different haplotypes (BALB/c and C57BL/6) with chimeric core particles induces a high-titered and highly cross-reactive N-specific antibody response in both mice strains

  14. Liver cancer-derived hepatitis C virus core proteins shift TGF-beta responses from tumor suppression to epithelial-mesenchymal transition.

    Directory of Open Access Journals (Sweden)

    Serena Battaglia

    Full Text Available BACKGROUND: Chronic hepatitis C virus (HCV infection and associated liver cirrhosis represent a major risk factor for hepatocellular carcinoma (HCC development. TGF-beta is an important driver of liver fibrogenesis and cancer; however, its actual impact in human cancer progression is still poorly known. The aim of this study was to investigate the role of HCC-derived HCV core natural variants on cancer progression through their impact on TGF-beta signaling. PRINCIPAL FINDINGS: We provide evidence that HCC-derived core protein expression in primary human or mouse hepatocyte alleviates TGF-beta responses in terms or growth inhibition or apoptosis. Instead, in these hepatocytes TGF-beta was still able to induce an epithelial to mesenchymal transition (EMT, a process that contributes to the promotion of cell invasion and metastasis. Moreover, we demonstrate that different thresholds of Smad3 activation dictate the TGF-beta responses in hepatic cells and that HCV core protein, by decreasing Smad3 activation, may switch TGF-beta growth inhibitory effects to tumor promoting responses. CONCLUSION/SIGNIFICANCE: Our data illustrate the capacity of hepatocytes to develop EMT and plasticity under TGF-beta, emphasize the role of HCV core protein in the dynamic of these effects and provide evidence for a paradigm whereby a viral protein implicated in oncogenesis is capable to shift TGF-beta responses from cytostatic effects to EMT development.

  15. Room-temperature synthesis of core-shell structured magnetic covalent organic frameworks for efficient enrichment of peptides and simultaneous exclusion of proteins.

    Science.gov (United States)

    Lin, Guo; Gao, Chaohong; Zheng, Qiong; Lei, Zhixian; Geng, Huijuan; Lin, Zian; Yang, Huanghao; Cai, Zongwei

    2017-03-28

    Core-shell structured magnetic covalent organic frameworks (Fe 3 O 4 @COFs) were synthesized via a facile approach at room temperature. Combining the advantages of high porosity, magnetic responsiveness, chemical stability and selectivity, Fe 3 O 4 @COFs can serve as an ideal absorbent for the highly efficient enrichment of peptides and the simultaneous exclusion of proteins from complex biological samples.

  16. A 7-day high protein hypocaloric diet promotes cellular metabolic adaptations and attenuates lean mass loss in healthy males

    Directory of Open Access Journals (Sweden)

    Matthew Furber

    2017-08-01

    Full Text Available Mitochondrial quantity and density are associated with increased oxidative metabolism. It has been demonstrated that a hypocaloric high fat/low carbohydrate (HF/LC diet can up-regulate transcriptional markers of mitochondrial biogenesis; this was yet to be explored in vivo subsequent to a high protein/low carbohydrate (HP/LC diet. Thus the aims of the study were to explore such diets on transcriptional markers or mitochondrial biogenesis, body composition and resting metabolic rate (RMR. Forty-five healthy male participants were randomly assigned one of four intervention diets: eucaloric high protein low carbohydrate (PRO-EM, hypocaloric high protein low carbohydrate (PRO-ER, eucaloric high carbohydrate (CHO-EM or hypocaloric high carbohydrate (CHO-ER. The macronutrient ratio of the high protein diet and high carbohydrate diets was 40:30:30% and 10:60:30% (PRO:CHO:FAT respectively. Energy intake for the hypocaloric diets were calculated to match resting metabolic rate. Participants visited the laboratory on 3 occasions each separated by 7 days. On each visit body composition, resting metabolic rate and a muscle biopsy from the vastus lateralis was collected. Prior to visit 1 and 2 habitual diet was consumed which was used as a control, between visit 2 and 3 the intervention diet was consumed continuously for 7-days. No group × time effect was observed, however in the PRO-ER group a significant increase in AMPK, PGC-1α, SIRT1 and SIRT3 mRNA expression was observed post diet intervention groups (p < 0.05. No change was observed in any of the transcriptional markers in the other 3 groups. Despite ∼30% reduction in calorie intake no difference in lean mass (LM loss was observed between the PRO-ER and CHO-EM groups. The results from this study suggest that a 7-day a high protein low carbohydrate hypocaloric diet increased AMPK, SIRT1 and PGC-1 α mRNA expression at rest, and also suggest that increased dietary protein may attenuate LM mass

  17. Pulmonary heat shock protein expression after exposure to a metabolically activated Clara cell toxicant: relationship to protein adduct formation

    International Nuclear Information System (INIS)

    Williams, Kurt J.; Cruikshank, Michael K.; Plopper, Charles G.

    2003-01-01

    Heat shock proteins/stress proteins (Hsps) participate in regulation of protein synthesis and degradation and serve as general cytoprotectants, yet their role in lethal Clara cell injury is not clear. To define the pattern of Hsp expression in acute lethal Clara cell injury, mice were treated with the Clara cell-specific toxicant naphthalene (NA), and patterns of expression compared to electrophilic protein adduction and previously established organellar degradation and gluathione (GSH) depletion. In sites of lethal injury (distal bronchiole), prior to organellar degradation (1 h post-NA), protein adduction is detectable and ubiquitin, Hsp 25, Hsp 72, and heme-oxygenase 1 (HO-1) are increased. Maximal Hsp expression, protein adduction, and GSH depletion occur simultaneous (by 2-3 h) with early organelle disruption. Hsp expression is higher later (6-24 h), only in exfoliating cells. In airway sites (proximal bronchiole) with nonlethal Clara cell injury elevation of Hsp 25, 72, and HO-1 expression follows significant GSH depletion (greater than 50% 2 h post-NA). This data build upon our previous studies and we conclude that (1) in lethal (terminal bronchiole) and nonlethal (proximal bronchiole) Clara cell injury, Hsp induction is associated with the loss of GSH and increased protein adduction, and (2) in these same sites, organelle disruption is not a prerequisite for Hsp induction

  18. Metabolism of whole body protein in pregnant and non-pregnant gilts using 15N-glycine single-dose end-product method

    International Nuclear Information System (INIS)

    Wu De; Liu Huifang; Zhou Anguo; Wang Kangning; Yang Feng

    2007-01-01

    The metabolism of whole-body protein for pregnant and non-pregnant gilts was investigated using single-dose of 15 N-glycine end-product method. The results showed that there were no differences (P>0.05) in protein dynamic metabolism, amino acids utilization rate between pregnant and non-pregnant gilts at breeding. However, N flux, protein turnover rate, protein synthesis rate and breakdown rate of pregnant gilts were lower (P<0.05) than those of non-pregnant gilts at 30days after breeding, but the protein aggradiation's rate increased by 25% (P<0.05). During late gestation, N flux, protein turnover rate, protein synthesis rate and breakdown rate of pregnant sows were significantly increased (P<0.01), and protein aggradation's rate increased by 71.1%, compared with that of non-pregnant gilts. (authors)

  19. New method for the quality check of food proteins of the maintenance metabolism. 4. Investigation of isolated proteins as well as some protein sources of plant and animal origin

    Energy Technology Data Exchange (ETDEWEB)

    Simon, O; Hernandez, M; Bergner, H [Humboldt-Universitaet, Berlin (German Democratic Republic). Sektion Tierproduktion und Veterinaermedizin

    1981-01-01

    Male adult rats (370 g body weight) were fed on maintenance level (460 kJ ME/kgsup(0,75). In a 10 days preliminary period they received a casein/methionine (95/5) diet supplemented with 10 mg /sup 15/N excess per 0.178 kg metabolic body weight in form of ammonium acetate. Thereafter the animals were put on 8 isonitrogenous diets containing as protein sources casein, soya protein, gelatine, whole-egg, fish meal, pea, wheat and yeast. The /sup 15/N excretion via urine and feces was used to evaluate the dietary proteins for maintenance. /sup 15/N in urine was lowest in animals fed on wheat diet and highest after feeding whole-egg diet. From these data a so called '/sup 15/N excretion biological valence (BV)' was calculated, which indicated the highest quality for wheat and soy protein in meeting the needs of the intermediary maintenance metabolism. On the other hand, dietary protein sources influence the loss of endogenous nitrogen as metabolic fecal nitrogen (MFN). It was found to be lowest in animals fed on diets containing isolated proteins (6 mg MFN/100 g body weight) and highest after feeding protein sources of plant origin with a high content in crude fibre (10 mg MFN/100 g). Both, losses of /sup 15/N via urine and via feces were combined in a parameter called 'total BV'. According to this parameter the differences in quality for maintenance were only little between the protein sources tested (casein 100, soy protein 100, pea 99, wheat 99, whole egg 92, fish meal 89, gelatin 89). It was concluded that in the state of maintenance the supply with essential amino acids is not critical and that the supply with dispensable amino acids (or nonspecific nitrogen) is of great importance.

  20. Proteomic analysis of ACTN4-interacting proteins reveals it's a putative involvement in mRNA metabolism

    International Nuclear Information System (INIS)

    Khotin, Mikhail; Turoverova, Lidia; Aksenova, Vasilisa; Barlev, Nikolai; Borutinskaite, Veronika Viktorija; Vener, Alexander; Bajenova, Olga; Magnusson, Karl-Eric; Pinaev, George P.; Tentler, Dmitri

    2010-01-01

    Alpha-actinin 4 (ACTN4) is an actin-binding protein. In the cytoplasm, ACTN4 participates in structural organisation of the cytoskeleton via cross-linking of actin filaments. Nuclear localisation of ACTN4 has also been reported, but no clear role in the nucleus has been established. In this report, we describe the identification of proteins associated with ACTN4 in the nucleus. A combination of two-dimensional gel electrophoresis (2D-GE) and MALDI-TOF mass-spectrometry revealed a large number of ACTN4-bound proteins that are involved in various aspects of mRNA processing and transport. The association of ACTN4 with different ribonucleoproteins suggests that a major function of nuclear ACTN4 may be regulation of mRNA metabolism and signaling.

  1. Threonine supplementation reduces dietary protein and improves lipid metabolism in Pekin ducks.

    Science.gov (United States)

    Jiang, Y; Tang, J; Xie, M; Wen, Z G; Qiao, S Y; Hou, S S

    2017-12-01

    1. This study was conducted to investigate the efficiency of threonine (Thr) supplementation on reducing dietary crude protein (CP) content and the effects of Thr on lipid metabolism in Pekin ducks. The effects of dietary CP concentration (160, 190 and 220 g/kg) and Thr supplemental concentration (0, 0.7, 1.4, 2.1 and 2.8 g/kg) on growth performance, carcass, liver lipid and plasma profiles were determined in Pekin ducks from 1-21 d of age. 2. A total of 720-d-old male Pekin ducks were randomly allotted to 1 of 15 dietary treatments with 6 replicate cages of 8 birds per cage for each treatment according to average body weight. 3. Dietary Thr supplementation improved growth performance and breast muscle percentage at all CP diets, and ducks fed Thr-supplemented diets had higher plasma concentrations of some plasma amino acids. Thr supplementation reduced the concentrations of total lipid, triglyceride, cholesterol in liver, and plasma low density lipoprotein cholesterin concentration at 160 and 190 g/kg CP, whereas it increased triglyceride concentration at 160 g/kg CP. 4. Thr requirements based on quadratic broken-line model estimation were 6.6 and 7.0 g/kg for optimal average daily gain (ADG), and 6.7 and 7.3 g/kg for breast muscle percentage of Pekin ducks from 1-21 d of age at 190 and 220 g/kg CP, respectively. The dietary Thr requirements and estimated ADG (55.18 vs. 55.86 g/d/bird) and breast muscle percentage (2.79% vs. 2.75%) of Pekin ducks did not differ between 190 and 220 g/kg CP according to the t-test results. 5. Dietary CP level could be reduced to 190 g/kg in Pekin ducks from 1-21 d of age with Thr supplementation to balance dietary amino acids, and Thr supplementation prevented excess liver lipid deposition in this instance.

  2. Evaluation of metabolism, plasma protein binding and other biological parameters after administration of (−)-[18 F]Flubatine in humans

    International Nuclear Information System (INIS)

    Patt, Marianne; Becker, Georg A.; Grossmann, Udo; Habermann, Bernd; Schildan, Andreas; Wilke, Stephan; Deuther-Conrad, Winnie; Graef, Susanne; Fischer, Steffen; Smits, René; Hoepping, Alexander; Wagenknecht, Gudrun; Steinbach, Jörg; Gertz, Hermann-Josef; Hesse, Swen; Schönknecht, Peter

    2014-01-01

    Introduction: (−)-[ 18 F]Flubatine is a PET tracer with high affinity and selectivity for the nicotinic acetylcholine α 4 β 2 receptor subtype. A clinical trial assessing the availability of this subtype of nAChRs was performed. From a total participant number of 21 Alzheimer’s disease (AD) patients and 20 healthy controls (HCs), the following parameters were determined: plasma protein binding, metabolism and activity distribution between plasma and whole blood. Methods: Plasma protein binding and fraction of unchanged parent compound were assessed by ultracentrifugation and HPLC, respectively. The distribution of radioactivity (parent compound + metabolites) between plasma and whole blood was determined ex vivo at different time-points after injection by gamma counting after separation of whole blood by centrifugation into the cellular and non-cellular components. In additional experiments in vitro, tracer distribution between these blood components was assessed for up to 90 min. Results: A fraction of 15% ± 2% of (−)-[ 18 F]Flubatine was found to be bound to plasma proteins. Metabolic degradation of (−)-[ 18 F]Flubatine was very low, resulting in almost 90% unchanged parent compound at 90 min p.i. with no significant difference between AD and HC. The radioactivity distribution between plasma and whole blood changed in vivo only slightly over time from 0.82 ± 0.03 at 3 min p.i. to 0.87 ± 0.03 at 270 min p.i. indicating the contribution of only a small amount of metabolites. In vitro studies revealed that (−)-[ 18 F]Flubatine was instantaneously distributed between cellular and non-cellular blood parts. Discussion: (−)-[ 18 F]Flubatine exhibits very favourable characteristics for a PET radiotracer such as slow metabolic degradation and moderate plasma protein binding. Equilibrium of radioactivity distribution between plasma and whole blood is reached instantaneously and remains almost constant over time allowing both convenient sample handling and

  3. Effect of long-term refeeding on protein metabolism in patients with cirrhosis of the liver

    DEFF Research Database (Denmark)

    Kondrup, J; Nielsen, K; Juul, A

    1997-01-01

    in protein synthesis was associated with significant increases in plasma concentrations of total amino acids (25%), leucine (58%), isoleucine (82%), valine (72%), proline (48%) and triiodothyronine (27%) while insulin, growth hormone, insulin-like growth factor (IGF)-I and IGF-binding protein-3 were...... a normal protein requirement with the patients who had an increased protein requirement suggests that the increased protein requirement is due to a primary increase in protein degradation. It is speculated that this is due to low levels of IGF-I secondary to impaired liver function, since initial plasma...

  4. Protein profiling of plastoglobules in chloroplasts and chromoplasts. A surprising site for differential accumulation of metabolic enzymes.

    Science.gov (United States)

    Ytterberg, A Jimmy; Peltier, Jean-Benoit; van Wijk, Klaas J

    2006-03-01

    Plastoglobules (PGs) are oval or tubular lipid-rich structures present in all plastid types, but their specific functions are unclear. PGs contain quinones, alpha-tocopherol, and lipids and, in chromoplasts, carotenoids as well. It is not known whether PGs contain any enzymes or regulatory proteins. Here, we determined the proteome of PGs from chloroplasts of stressed and unstressed leaves of Arabidopsis (Arabidopsis thaliana) as well as from pepper (Capsicum annuum) fruit chromoplasts using mass spectrometry. Together, this showed that the proteome of chloroplast PGs consists of seven fibrillins, providing a protein coat and preventing coalescence of the PGs, and an additional 25 proteins likely involved in metabolism of isoprenoid-derived molecules (quinines and tocochromanols), lipids, and carotenoid cleavage. Four unknown ABC1 kinases were identified, possibly involved in regulation of quinone monooxygenases. Most proteins have not been observed earlier but have predicted N-terminal chloroplast transit peptides and lack transmembrane domains, consistent with localization in the PG lipid monolayer particles. Quantitative differences in PG composition in response to high light stress and degreening were determined by differential stable-isotope labeling using formaldehyde. More than 20 proteins were identified in the PG proteome of pepper chromoplasts, including four enzymes of carotenoid biosynthesis and several homologs of proteins observed in the chloroplast PGs. Our data strongly suggest that PGs in chloroplasts form a functional metabolic link between the inner envelope and thylakoid membranes and play a role in breakdown of carotenoids and oxidative stress defense, whereas PGs in chromoplasts are also an active site for carotenoid conversions.

  5. Protein Profiling of Plastoglobules in Chloroplasts and Chromoplasts. A Surprising Site for Differential Accumulation of Metabolic Enzymes1[W

    Science.gov (United States)

    Ytterberg, A. Jimmy; Peltier, Jean-Benoit; van Wijk, Klaas J.

    2006-01-01

    Plastoglobules (PGs) are oval or tubular lipid-rich structures present in all plastid types, but their specific functions are unclear. PGs contain quinones, α-tocopherol, and lipids and, in chromoplasts, carotenoids as well. It is not known whether PGs contain any enzymes or regulatory proteins. Here, we determined the proteome of PGs from chloroplasts of stressed and unstressed leaves of Arabidopsis (Arabidopsis thaliana) as well as from pepper (Capsicum annuum) fruit chromoplasts using mass spectrometry. Together, this showed that the proteome of chloroplast PGs consists of seven fibrillins, providing a protein coat and preventing coalescence of the PGs, and an additional 25 proteins likely involved in metabolism of isoprenoid-derived molecules (quinines and tocochromanols), lipids, and carotenoid cleavage. Four unknown ABC1 kinases were identified, possibly involved in regulation of quinone monooxygenases. Most proteins have not been observed earlier but have predicted N-terminal chloroplast transit peptides and lack transmembrane domains, consistent with localization in the PG lipid monolayer particles. Quantitative differences in PG composition in response to high light stress and degreening were determined by differential stable-isotope labeling using formaldehyde. More than 20 proteins were identified in the PG proteome of pepper chromoplasts, including four enzymes of carotenoid biosynthesis and several homologs of proteins observed in the chloroplast PGs. Our data strongly suggest that PGs in chloroplasts form a functional metabolic link between the inner envelope and thylakoid membranes and play a role in breakdown of carotenoids and oxidative stress defense, whereas PGs in chromoplasts are also an active site for carotenoid conversions. PMID:16461379

  6. Krüppel-like factor 15: Regulator of BCAA metabolism and circadian protein rhythmicity.

    Science.gov (United States)

    Fan, Liyan; Hsieh, Paishiun N; Sweet, David R; Jain, Mukesh K

    2018-04-01

    Regulation of nutrient intake, utilization, and storage exhibits a circadian rhythmicity that allows organisms to anticipate and adequately respond to changes in the environment across day/night cycles. The branched-chain amino acids (BCAAs) leucine, isoleucine, and valine are important modulators of metabolism and metabolic health - for example, their catabolism yields carbon substrates for gluconeogenesis during periods of fasting. Krüppel-like factor 15 (KLF15) has recently emerged as a critical transcriptional regulator of BCAA metabolism, and the absence of this transcription factor contributes to severe pathologies such as Duchenne muscular dystrophy and heart failure. This review highlights KLF15's role as a central regulator of BCAA metabolism during periods of fasting, throughout day/night cycles, and in experimental models of muscle disease. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Biochemical studies of effects of alcohol consumption on fat and carbohydrate metabolism in rats fed different levels of proteins

    International Nuclear Information System (INIS)

    Shalan, M.G.M.

    1996-01-01

    Alcohol, ethanol and ethyl alcohol are synonymously used during the present dissertation. Alcohol probably was among the first psychoactive substances to be used by man (Winger et al., 1992). Ethanol is mainly oxidized to acetaldehyde in the liver (Ugarte and Peresa, 1978) by alcohol dehydrogenase (ADH). Alcohol is associated with many metabolic disorders inside the body (Thayer and Rubin, 1979; Forsander and Poso, 1988; Poso and Hirsimaki, 1991; Bernal, et al., 1992). The nutritional factors which received little attention have an important role in alcoholic metabolizing alterations. Morphologically and biochemically, an increase in hepatic lipid was demonstrated when ethanol was given either as a supplement or as an iso caloric substitute for carbohydrate together with an otherwise nutritionally adequate diet. Low-protein diets have been shown to diminish hepatic alcohol dehydrogenase (ADH) levels in rats and to slow down the metabolism of ethanol considerably (Wilson et al., 1986). Hepatic steatosis was produced, even with a high-protein, vitamin-supplemented diet and was accompanied by major ultrastructural liver changes and by elevations of hepatic transaminases in blood (Lieber et al., 1963 and 1965 and Lane and Lieber, 1966). If dietary fat was reduced from 35 to 25% of total calories, hepatic triglyceride accumulation greatly decreased (Lieber and DeCarli, 970)

  8. Protein and lipid metabolism adjustments in silver catfish (Rhamdia quelen during different periods of fasting and refeeding

    Directory of Open Access Journals (Sweden)

    A. Marqueze

    2017-10-01

    Full Text Available Abstract The fish may experience periods of food deprivation or starvation which produce metabolic changes. In this study, adult Rhamdia quelen males were subjected to fasting periods of 1, 7, 14, and 21 days and of refeeding 2, 4, 6, and 12 days. The results demonstrated that liver protein was depleted after 1 day of fasting, but recovered after 6 days of refeeding. After 14 days of fasting, mobilization in the lipids of the muscular tissue took place, and these reserves began to re-establish themselves after 4 days of refeeding. Plasmatic triglycerides increased after 1 day of fasting, and decreased following 2 days of refeeding. The glycerol in the plasma oscillated constantly during the different periods of fasting and refeeding. Changes in the metabolism of both protein and lipids during these periods can be considered as survival strategies used by R. quelen. The difference in the metabolic profile of the tissues, the influence of the period of fasting, and the type of reserves mobilized were all in evidence.

  9. In vitro assembly into virus-like particles is an intrinsic quality of Pichia pastoris derived HCV core protein

    International Nuclear Information System (INIS)

    Acosta-Rivero, Nelson; Rodriguez, Armando; Musacchio, Alexis; Falcon, Viviana; Suarez, Viana M.; Martinez, Gillian; Guerra, Ivis; Paz-Lago, Dalila; Morera, Yanelys; Rosa, Maria C. de la; Morales-Grillo, Juan; Duenas-Carrera, Santiago

    2004-01-01

    Different variants of hepatitis C virus core protein (HCcAg) have proved to self-assemble in vitro into virus-like particles (VLPs). However, difficulties in obtaining purified mature HCcAg have limited these studies. In this study, a high degree of monomeric HCcAg purification was accomplished using chromatographic procedures under denaturing conditions. Size exclusion chromatography and sucrose density gradient centrifugation of renatured HCcAg (in the absence of structured RNA) under reducing conditions suggested that it assembled into empty capsids. The electron microscopy analysis of renatured HCcAg showed the presence of spherical VLPs with irregular shapes and an average diameter of 35 nm. Data indicated that HCcAg monomers assembled in vitro into VLPs in the absence of structured RNA, suggesting that recombinant HCcAg used in this work contains all the information necessary for the assembly process. However, they also suggest that some cellular factors might be required for the proper in vitro assembly of capsids

  10. Urinary deoxypyridinoline (DPD), serum bone glia protein (BGP) and bone metabolism change in hyperthyroidism

    International Nuclear Information System (INIS)

    Zeng Yun; Ding Jianzhong; Xiang Hong

    2002-01-01

    Objective: To study the effect of thyroid function on bone metabolism. Methods: Urinary DPD, Serum FT 3 , FT 4 and BGP levels were determined with chemiluminescence assay and RIA in 41 patients with hyperthyroidism and 47 healthy controls. Results: Urinary DPD and serum FT 3 , FT 4 , BGP levels were significantly higher in patients with hyperthyroidism than those in healthy controls (p < 0.01). Conclusion: The data showed that hyperthyroidism was correlated with bone metabolism

  11. The Biased G-Protein-Coupled Receptor Agonism Bridges the Gap between the Insulin Receptor and the Metabolic Syndrome

    Science.gov (United States)

    Liauchonak, Iryna; Dawoud, Fady; Riat, Yatin; Sambi, Manpreet; Jain, Justin; Kalaydina, Regina-Veronicka; Mendonza, Nicole; Bajwa, Komal

    2018-01-01

    Insulin signaling, as mediated through the insulin receptor (IR), plays a critical role in metabolism. Aberrations in this signaling cascade lead to several pathologies, the majority of which are classified under the umbrella term “metabolic syndrome”. Although many of these pathologies are associated with insulin resistance, the exact mechanisms are not well understood. One area of current interest is the possibility of G-protein-coupled receptors (GPCRs) influencing or regulating IR signaling. This concept is particularly significant, because GPCRs have been shown to participate in cross-talk with the IR. More importantly, GPCR signaling has also been shown to preferentially regulate specific downstream signaling targets through GPCR agonist bias. A novel study recently demonstrated that this GPCR-biased agonism influences the activity of the IR without the presence of insulin. Although GPCR-IR cross-talk has previously been established, the notion that GPCRs can regulate the activation of the IR is particularly significant in relation to metabolic syndrome and other pathologies that develop as a result of alterations in IR signaling. As such, we aim to provide an overview of the physiological and pathophysiological roles of the IR within metabolic syndrome and its related pathologies, including cardiovascular health, gut microflora composition, gastrointestinal tract functioning, polycystic ovarian syndrome, pancreatic cancer, and neurodegenerative disorders. Furthermore, we propose that the GPCR-biased agonism may perhaps mediate some of the downstream signaling effects that further exacerbate these diseases for which the mechanisms are currently not well understood. PMID:29462993

  12. Cholesteryl ester transfer protein alters liver and plasma triglyceride metabolism through two liver networks in female mice[S

    Science.gov (United States)

    Palmisano, Brian T.; Le, Thao D.; Zhu, Lin; Lee, Yoon Kwang; Stafford, John M.

    2016-01-01

    Elevated plasma TGs increase risk of cardiovascular disease in women. Estrogen treatment raises plasma TGs in women, but molecular mechanisms remain poorly understood. Here we explore the role of cholesteryl ester transfer protein (CETP) in the regulation of TG metabolism in female mice, which naturally lack CETP. In transgenic CETP females, acute estrogen treatment raised plasma TGs 50%, increased TG production, and increased expression of genes involved in VLDL synthesis, but not in nontransgenic littermate females. In CETP females, estrogen enhanced expression of small heterodimer partner (SHP), a nuclear receptor regulating VLDL production. Deletion of liver SHP prevented increases in TG production and expression of genes involved in VLDL synthesis in CETP mice with estrogen treatment. We also examined whether CETP expression had effects on TG metabolism independent of estrogen treatment. CETP increased liver β-oxidation and reduced liver TG content by 60%. Liver estrogen receptor α (ERα) was required for CETP expression to enhance β-oxidation and reduce liver TG content. Thus, CETP alters at least two networks governing TG metabolism, one involving SHP to increase VLDL-TG production in response to estrogen, and another involving ERα to enhance β-oxidation and lower liver TG content. These findings demonstrate a novel role for CETP in estrogen-mediated increases in TG production and a broader role for CETP in TG metabolism. PMID:27354419

  13. Cholesteryl ester transfer protein alters liver and plasma triglyceride metabolism through two liver networks in female mice.

    Science.gov (United States)

    Palmisano, Brian T; Le, Thao D; Zhu, Lin; Lee, Yoon Kwang; Stafford, John M

    2016-08-01

    Elevated plasma TGs increase risk of cardiovascular disease in women. Estrogen treatment raises plasma TGs in women, but molecular mechanisms remain poorly understood. Here we explore the role of cholesteryl ester transfer protein (CETP) in the regulation of TG metabolism in female mice, which naturally lack CETP. In transgenic CETP females, acute estrogen treatment raised plasma TGs 50%, increased TG production, and increased expression of genes involved in VLDL synthesis, but not in nontransgenic littermate females. In CETP females, estrogen enhanced expression of small heterodimer partner (SHP), a nuclear receptor regulating VLDL production. Deletion of liver SHP prevented increases in TG production and expression of genes involved in VLDL synthesis in CETP mice with estrogen treatment. We also examined whether CETP expression had effects on TG metabolism independent of estrogen treatment. CETP increased liver β-oxidation and reduced liver TG content by 60%. Liver estrogen receptor α (ERα) was required for CETP expression to enhance β-oxidation and reduce liver TG content. Thus, CETP alters at least two networks governing TG metabolism, one involving SHP to increase VLDL-TG production in response to estrogen, and another involving ERα to enhance β-oxidation and lower liver TG content. These findings demonstrate a novel role for CETP in estrogen-mediated increases in TG production and a broader role for CETP in TG metabolism. Copyright © 2016 by the American Society for Biochemistry and Molecular Biology, Inc.

  14. CCAAT/Enhancer Binding Protein β in relation to ER Stress, Inflammation, and Metabolic Disturbances

    Directory of Open Access Journals (Sweden)

    Sophie E. van der Krieken

    2015-01-01

    Full Text Available The prevalence of the metabolic syndrome and underlying metabolic disturbances increase rapidly in developed countries. Various molecular targets are currently under investigation to unravel the molecular mechanisms that cause these disturbances. This is done in attempt to counter or prevent the negative health consequences of the metabolic disturbances. Here, we reviewed the current knowledge on the role of C/EBP-β in these metabolic disturbances. C/EBP-β deletion in mice resulted in downregulation of hepatic lipogenic genes and increased expression of β-oxidation genes in brown adipose tissue. Furthermore, C/EBP-β is important in the differentiation and maturation of adipocytes and is increased during ER stress and proinflammatory conditions. So far, studies were only conducted in animals and in cell systems. The results found that C/EBP-β is an important transcription factor within the metabolic disturbances of the metabolic system. Therefore, it is interesting to examine the potential role of C/EBP-β at molecular and physiological level in humans.

  15. [Effect of protein intervention on amino acid metabolism spectrum of Qi and Yin deficiency type 2 diabetic rats].

    Science.gov (United States)

    Ma, Li-Na; Mao, Xin-Min; Ma, Xiao-Li; Li, Lin-Lin; Wang, Ye; Tao, Yi-Cun; Wang, Jing-Wei; Guo, Jia-Jia; Lan, Yi

    2016-11-01

    To study the effect of plant protein and animal protein on amino acid metabolism spectrum of Qi and Yin deficiency type 2 diabetic rats. 110 male SD rats were randomly divided into blank group (n=10), diabetic model group (n=20), disease-symptoms group (n=80). The rats of blank group received ordinary feeding, while other groups were fed with high sugar and fat diets. During the whole process of feeding, rats of disease-symptoms group were given with Qingpi-Fuzi (15.75 g•kg⁻¹) once a day through oral administration. Five weeks later, the rats were given with a low dose of STZ (40 mg•kg⁻¹) by intraperitoneal injection to establish experimental diabetic models. Then the models were randomly divided into disease-symptoms group 1 (Qi and Yin deficiency diabetic group, 15.75 g•kg⁻¹), disease-symptoms group 2 (plant protein group, 0.5 g•kg⁻¹), disease-symptoms group 3 (animal protein group, 0.5 g•kg⁻¹), disease-symptoms group 4 (berberine group, 0.1 g•kg⁻¹). The drugs were given for 4 weeks by gavage administration. After 4 weeks of protein intervention, the abdominal aortic blood was collected and serum was isolated to analyze its free amino acid by using AQC pre-column derivatization HPLC and fluorescence detector. Four weeks after the protein intervention, plant protein, animal protein and berberine had no obvious effect on body weight and blood sugar in type 2 diabetic rats. As compared with animal protein group, histidine and proline(PYin deficiency type 2 diabetic SD rats. Symbolic differential compounds could be found through metabonomics technology, providing experimental basis for early warning of type 2 diabetes and diagnosis of Qi and Yin deficiency syndrome. Copyright© by the Chinese Pharmaceutical Association.

  16. Differential regulation of lipid and protein metabolism in obese vs. lean subjects before and after