• 155. Effect of methanol and DMSO exposure on mitochondrial activity and distribution in stage III ovarian follicles of zebrafish Danio rerio

      Zampolla, Tiziana; Spikings, Emma; Zhang, Tiantian; Rawson, David M.; University of Bedfordshire (Elsevier, 2009-12)
    • 160. Effect of cryoprotectant treatment and chilling on oxidative stress in zebrafish (Danio rerio) early ovarian follicles

      Ghafari, Fataneh; Spikings, Emma; Rawson, David M.; Zhang, Tiantian; University of Bedfordshire (Elsevier, 2009-12)
    • Amino acid cycling by Rhizobium leguminosarum in pea nodules

      White, James P.; Bourdes, Alex; Hosie, Arthur H.F.; Kinghorn, Seonag; Poole, Philip S. (Springer, 2005)
    • Amino-acid cycling drives nitrogen fixation in the legume–Rhizobium symbiosis

      Lodwig, Emma M.; Hosie, Arthur H.F.; Bourdes, Alex; Findlay, K.; Allaway, D.; Karunakaran, R.; Downie, J. A.; Poole, Philip S. (Nature Publishing Group, 2003)
      The biological reduction of atmospheric N2 to ammonium (nitrogen fixation) provides about 65% of the biosphere's available nitrogen. Most of this ammonium is contributed by legume–rhizobia symbioses1, which are initiated by the infection of legume hosts by bacteria (rhizobia), resulting in formation of root nodules. Within the nodules, rhizobia are found as bacteroids, which perform the nitrogen fixation: to do this, they obtain sources of carbon and energy from the plant, in the form of dicarboxylic acids2, 3. It has been thought that, in return, bacteroids simply provide the plant with ammonium. But here the authors show that a more complex amino-acid cycle is essential for symbiotic nitrogen fixation by Rhizobium in pea nodules. The plant provides amino acids to the bacteroids, enabling them to shut down their ammonium assimilation. In return, bacteroids act like plant organelles to cycle amino acids back to the plant for asparagine synthesis. The mutual dependence of this exchange prevents the symbiosis being dominated by the plant, and provides a selective pressure for the evolution of mutualism.
    • The analysis of mitochondria and mitochondrial DNA in human embryonic stem cells.

      St John, Justin C.; Amaral, Alexandra; Bowles, Emma J.; Oliveira, João Facucho; Lloyd, Rhiannon E.I.; Freitas, Mariana; Gray, Heather L.; Navara, Christopher S.; Oliveira, Gisela; Schatten, Gerald P.; et al. (Humana Press, 2006)
      As human embryonic stem cells (hESCs) undergo differentiation, they express genes characteristic of the lineage for which they are destined. However, fully differentiated individual cell types can be characterized by the number of mitochondria they possess and the copies of the mitochondrial genome per mitochondrion. These characteristics are indicative of a specific cell's requirement for adenosine triphosphate (ATP) and therefore cellular viability and function. Consequently, failure for an ESC to possess the full complement of mitochondria and mitochondrial DNA (mtDNA) could limit its final commitment to a particular fate. We describe a series of protocols that analyze the process of cellular mitochondrial and mtDNA differentiation during hESC differentiation. In addition, mtDNA transcription and replication are key events in cellular differentiation that require interaction between the nucleus and the mitochondrion. To this extent, we describe a series of protocols that analyze the initiation of these key events as hESCs progress from their undifferentiated state to the fully committed cell. Last, we describe real-time polymerase chain reaction protocols that allow both the identification of mtDNA copy number and determine whether mtDNA copy is uniform (homoplasmy) in its transmission or heterogeneous (heteroplasmy).
    • Antitumor effects of saponin extract from Patrinia villosa (Thunb.) Juss on mice bearing U14 cervical cancer

      Zhang, Tao; Li, Qingwang; Li, Kun; Li, Yurong; Li, Jian; Wang, Guijie; Zhou, Shaobo (John Wiley and Sons, 2008-03)
    • Bacterial ABC transporters of amino acids

      Hosie, Arthur H.F.; Poole, Philip S. (Elsevier, 2001)
      There are two subfamilies of ABC uptake systems for amino acids in bacteria, the polar amino acid transport family and the hydrophobic amino acid transport family. We consider the general properties of these families and we examine the specific transporters. Focusing on some of the best-studied ATP binding cassette transporters we also examine the mechanism of amino acid uptake, paying particular attention to the question of bidirectionality of solute movement.
    • Binding modes of diketo-acid inhibitors of HIV-1 integrase: a comparative molecular dynamics simulation study.

      Huang, Meilan; Grant, Guy H.; Richards, W. Graham; Queens University Belfast (Elsevier, 2011-06)
      HIV-1 integrase (IN) has become an attractive target since drug resistance against HIV-1 reverse transcriptase (RT) and protease (PR) has appeared. Diketo acid (DKA) inhibitors are potent and selective inhibitors of HIV-1 IN: however the action mechanism is not well understood. Here, to study the inhibition mechanism of DKAs we performed 10 ns comparative molecular dynamics simulations on HIV-1 IN bound with three most representative DKA inhibitors: Shionogi inhibitor, S-1360 and two Merck inhibitors L-731,988 and L-708,906. Our simulations show that the acidic part of S-1360 formed salt bridge and cation-π interactions with Lys159. In addition, the catalytic Glu152 in S-1360 was pushed away from the active site to form an ion-pair interaction with Arg199. The Merck inhibitors can maintain either one or both of these ion-pair interaction features. The difference in potencies of the DKA inhibitors is thus attributed to the different binding modes at the catalytic site. Such structural information at atomic level, not only demonstrates the action modes of DKA inhibitors but also provides a novel starting point for structural-based design of HIV-1 IN inhibitors.
    • The C-4 stereochemistry of leucocyanidin substrates for anthocyanidin synthase affects product selectivity

      Turnbull, Jonathan J.; Nagle, Michael J.; Seibel, Jürgen F.; Welford, Richard W.D.; Grant, Guy H.; Schofield, Christopher J. (Elsevier, 2003)
      Anthocyanidin synthase (ANS), an iron(II) and 2-oxoglutarate (2OG) dependent oxygenase, catalyses the penultimate step in anthocyanin biosynthesis by oxidation of the 2R,3S,4S-cis-leucoanthocyanidins. It has been believed that in vivo the products of ANS are the anthocyanidins. However, in vitro studies on ANS using optically active cis- and trans-leucocyanidin substrates identified cyanidin as only a minor product; instead both quercetin and dihydroquercetin are products with the distribution being dependent on the C-4 stereochemistry of the leucocyanidin substrates.
    • Ca2+-regulated pool of phosphatidylinositol-3-phosphate produced by phosphatidylinositol 3-Kinase C2  on neurosecretory vesicles

      Wen, P.J.; Osborne, S.L.; Morrow, I.C.; Parton, R.G.; Domin, Jan; Meunier, F.A. (American Society for Cell Biology, 2008-12)
    • Calculation of protein domain structural similarity using two-dimensional representations

      Allen, B.C.P.; Grant, Guy H.; Richards, W. Graham (American Chemical Society, 2003)
      By reducing protein structures to two-dimensional representations, it is possible to speed up the alignment of the structures and hence calculate similarity indices faster that using three-dimensional representations. Using amino acid based representations gives much better discrimination between proteins and faster calculations. Taking into account the relative similarity of the amino acids involved allowed improved accuracy at very little time cost.
    • Caspase-2 and caspase-8 trigger caspase-3 activation following 6-OHDA-induced stress in human dopaminergic neurons differentiated from ReNVM stem cells

      Chaudhry, Zahara Latif; Ahmed, Bushra Y. (Maney Publishing, 2012)
      The purpose of the study was to establish a suitable model to study Parkinson’s disease (PD) pathogenesis in differentiated dopaminergic neurons (dDCN). The specific aim was to demonstrate the involvement of the caspase family and to identify specific caspases which are activated by 6-hydroxydopamine (6OHD) treatment leading to death of dDCN.
    • Caspase-mediated cleavage and functional changes of hematopoietic progenitor kinase 1 (HPK1)

      Chen, Yi-Rong; Meyer, Christian F; Ahmed, Bushra Y.; Yao, Zhengbin; Tan, Tse-Hua (Nature Publishing Group, 1999)
      Activation of c-Jun N-terminal kinase (JNK) by Fas ligation is caspase-dependent, suggesting that caspases may regulate activators of the JNK pathway. Here, we report that an upstream activator of JNK, hematopoietic progenitor kinase 1 (HPK1), was cleaved during apoptosis. Cleavage of HPK1 was blocked by peptide inhibitors for caspases. HPK1 was efficiently processed by recombinant caspase 3 in vitro. A conserved caspase recognition site, DDVD (amino acids 382 - 385), was found in the HPK1 protein sequence. By testing HPK1 proteins with in vivo and in vitro cleavage assays, we showed that aspartic acid residue 385 is the target for caspases. HPK1 cleavage separated the amino N-terminal kinase domain from the carboxyl C-terminal regulatory domain, and enhanced HPK1 kinase activity. Unlike the full-length HPK1, the N-terminal cleaved product failed to bind adaptor molecules Grb2 (growth factor receptor-bound protein 2) and Crk (CT10 regulator of kinase). The C-terminal fragment, although having three proline-rich domains, bound to Grb2 and Crk less efficiently than the full-length HPK1 protein. Taken together, the cleavage of HPK1 by caspase profoundly changed its biochemical properties.
    • Catalytic enantioselective [3 + 2]-cycloadditions of diazoketone-derived aryl-substituted carbonyl ylides

      Hodgson, David M.; Glen, Rebecca; Grant, Guy H.; Redgrave, Alison J.; University of Oxford; GlaxoSmithKline Medicines Research Centre (American Chemical Society, 2003)
      An evaluation of α-aryl-α-diazodiones in tandem carbonyl ylide formation−enantioselective [3 + 2]-cycloaddition reactions is described. Such substrates were designed to allow investigation of the electronic characteristics of the dipole upon asymmetric induction. Intramolecular cycloadditions (with a tethered alkene dipolarophile) were found to occur in good to quantitative yields, with a difference in ee exhibited by the two electronically different diazodiones 8 and 9. Intermolecular cycloadditions using diazodiones 12 and 13 with DMAD and arylacetylenes 16−18 again demonstrated that electronics play a key role in determining the outcome of the cycloaddition reactions. Enantioselectivities of up to 76% were observed.
    • Characterization and tissue-specific expression of two lepidopteran farnesyl diphosphate synthase homologs: Implications for the biosynthesis of ethyl-substituted juvenile hormones

      Cusson, Michel; Béliveau, Catherine; Sen, Stephanie E.; Vandermoten, Sophie; Rutledge, Robert G.; Stewart, Don; Francis, Frédéric; Haubruge, Éric; Rehse, Peter; Huggins, David J.; et al. (Wiley, 2006)
      The sesquiterpenoid juvenile hormone (JH) regulates insect development and reproduction. Most insects produce only one chemical form of JH, but the Lepidoptera produce four derivatives featuring ethyl branches. The biogenesis of these JHs requires the synthesis of ethyl-substituted farnesyl diphosphate (FPP) by FPP synthase (FPPS). To determine if there exist more than one lepidopteran FPPS, and whether one FPPS homolog is better adapted for binding the bulkier ethyl-branched substrates/products, we cloned three lepidopteran FPPS cDNAs, two from Choristoneura fumiferana and one from Pseudaletia unipuncta. Amino acid sequence comparisons among these and other eukaryotic FPPSs led to the recognition of two lepidopteran FPPS types.
    • Characterization of a novel sialic acid transporter of the sodium solute symporter (SSS) family and in vivo comparison with known bacterial sialic acid transporters.

      Severi, Emmanuele; Hosie, Arthur H.F.; Hawkhead, Judith A.; Thomas, Gavin H.; University of York (Blackwell Publishing, 2010-03)
      The function of sialic acids in the biology of bacterial pathogens is reflected by the diverse range of solute transporters that can recognize these sugar acids. Here, we use an Escherichia coliDeltananT strain to characterize the function of known and proposed bacterial sialic acid transporters. We discover that the STM1128 gene from Salmonella enterica serovar Typhimurium, which encodes a member of the sodium solute symporter family, is able to restore growth on sialic acid to the DeltananT strain and is able to transport [(14)C]-sialic acid. Using the DeltananT genetic background, we performed a direct in vivo comparison of the transport properties of the STM1128 protein with those of sialic acid transporters of the major facilitator superfamily and tripartite ATP-independent periplasmic families, E. coli NanT and Haemophilus influenzae SiaPQM, respectively. This revealed that both STM1128 and SiaPQM are sodium-dependent and, unlike SiaPQM, both STM1128 and NanT are reversible secondary carriers, demonstrating qualitative functional differences in the properties of sialic acid transporters used by bacteria that colonize humans.
    • A chronological study of the expression of glial fibrillary acidic protein and calbindin-D28 k by reactive astrocytes in the electrically lesioned rat brain

      Ahmed, Bushra Y.; Toyoshima, Tetsuhiko; Yamagami, Shinichi; Jin, Li; Itano, Toshifumi; Miyamoto, Osamu; Tokuda, Masaaki; Murakami, Tetsuhide H.; Hatase, Osamu (Elsevier, 1996)
      Immunoreactivity of neuronal and glial marker proteins of reactive astrocytes around the electrically damaged pyramidal layer and stratum radiatum of the hippocampal CA1 region and corpus callosum was chronologically studied in electrically lesioned rat brains. A monoclonal antibody against calbindin-D28 k (CD28-Ab) and a polyclonal antibody against glial fibrillary acidic protein (GFAP-Ab) were used for immunostaining
    • Classical molecular dynamics simulations of the complex between the RAD51 protein and the BRC hairpin loops of the BRCA2 protein

      Buis, Nick; Skylaris, Chris-Kriton; Grant, Guy H.; Rajendra, Eeson; Payne, Mike C.; Venkitaraman, Ashok R.; University of Southampton; University of Cambridge (Taylor & Francis, 2008-08)