Rozprawy doktorskie: „Carrier and Transport Proteins”

1

Covey, Scott D. Trigatti Bernardo L. "Carrier mediated lipid transport /". *McMaster only, 2003.

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Kwan, Miu-fan, i 關妙芬. "Characterization of TM4 of NRAMP1: implication for FEII transport". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2003. http://hub.hku.hk/bib/B29275143.

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Grabski, Robert. "Using RNA interference to study the function of the tethering protein p115 in ER-Golgi traffic". Thesis, Birmingham, Ala. : University of Alabama at Birmingham, 2008. https://www.mhsl.uab.edu/dt/2008p/grabski.pdf.

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Parry, Geraint. "Investigating the mechanisms of auxin transport". Thesis, University of Nottingham, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.391391.

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Christoffersen, Catherine Anne. "Molecular analysis of the ferric-enterobactin fepDGC transport permease complex in escherichia coli". free to MU campus, to others for purchase, 1997. http://wwwlib.umi.com/cr/mo/fullcit?p9842593.

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Bright, Alison R. "A Role for Intraflagellar Transport Proteins in Mitosis: A Dissertation". eScholarship@UMMS, 2013. https://escholarship.umassmed.edu/gsbs_diss/682.

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Disruption of cilia proteins results in a range of disorders called ciliopathies. However, the mechanism by which cilia dysfunction contributes to disease is not well understood. Intraflagellar transport (IFT) proteins are required for ciliogenesis. They carry ciliary cargo along the microtubule axoneme while riding microtubule motors. Interestingly, IFT proteins localize to spindle poles in non-ciliated, mitotic cells, suggesting a mitotic function for IFT proteins. Based on their role in cilia, we hypothesized that IFT proteins regulate microtubule-based transport during mitotic spindle assembly. Biochemical investigation revealed that in mitotic cells IFT88, IFT57, IFT52, and IFT20 interact with dynein1, a microtubule motor required for spindle pole maturation. Furthermore, IFT88 co-localizes with dynein1 and its mitotic cargo during spindle assembly, suggesting a role for IFT88 in regulating dynein-mediated transport to spindle poles. Based on these results we analyzed spindle poles after IFT protein depletion and found that IFT88 depletion disrupted EB1, γ-tubulin, and astral microtubule arrays at spindle poles. Unlike IFT88, depletion of IFT57, IFT52, or IFT20 did not disrupt spindle poles. Strikingly, the simultaneous depletion of IFT88 and IFT20 rescued the spindle pole disruption caused by IFT88 depletion alone, suggesting a model in which IFT88 negatively regulates IFT20, and IFT20 negatively regulates microtubulebased transport during mitosis. Our work demonstrates for the first time that IFT proteins function with dynein1 in mitosis, and it also raises the important possibility that mitotic defects caused by IFT protein disruption could contribute to the phenotypes associated with ciliopathies.

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Bright, Alison R. "A Role for Intraflagellar Transport Proteins in Mitosis: A Dissertation". eScholarship@UMMS, 2006. http://escholarship.umassmed.edu/gsbs_diss/682.

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Disruption of cilia proteins results in a range of disorders called ciliopathies. However, the mechanism by which cilia dysfunction contributes to disease is not well understood. Intraflagellar transport (IFT) proteins are required for ciliogenesis. They carry ciliary cargo along the microtubule axoneme while riding microtubule motors. Interestingly, IFT proteins localize to spindle poles in non-ciliated, mitotic cells, suggesting a mitotic function for IFT proteins. Based on their role in cilia, we hypothesized that IFT proteins regulate microtubule-based transport during mitotic spindle assembly. Biochemical investigation revealed that in mitotic cells IFT88, IFT57, IFT52, and IFT20 interact with dynein1, a microtubule motor required for spindle pole maturation. Furthermore, IFT88 co-localizes with dynein1 and its mitotic cargo during spindle assembly, suggesting a role for IFT88 in regulating dynein-mediated transport to spindle poles. Based on these results we analyzed spindle poles after IFT protein depletion and found that IFT88 depletion disrupted EB1, γ-tubulin, and astral microtubule arrays at spindle poles. Unlike IFT88, depletion of IFT57, IFT52, or IFT20 did not disrupt spindle poles. Strikingly, the simultaneous depletion of IFT88 and IFT20 rescued the spindle pole disruption caused by IFT88 depletion alone, suggesting a model in which IFT88 negatively regulates IFT20, and IFT20 negatively regulates microtubulebased transport during mitosis. Our work demonstrates for the first time that IFT proteins function with dynein1 in mitosis, and it also raises the important possibility that mitotic defects caused by IFT protein disruption could contribute to the phenotypes associated with ciliopathies.

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Nelson, Bryn D. "Examining the role of MalG in the assembly and function of the maltose transport complex in Escherichia coli : implications for the study of integral membrane proteins /". Thesis, Connect to this title online; UW restricted, 1998. http://hdl.handle.net/1773/11508.

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Bradley, Shannon. "Polymorphisms in the promoter region of the dopamine transporter : a candidate locus for alcohol abuse". Thesis, National Library of Canada = Bibliothèque nationale du Canada, 2000. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape2/PQDD_0029/MQ64326.pdf.

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Millman, Jonathan Scott Andrews David. "Characterization of membrane-binding by FtsY, the prokaryote SRP receptor /". *McMaster only, 2002.

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Ahlin, Gustav. "In vitro and in silico prediction of drug-drug interactions with transport proteins". Doctoral thesis, Uppsala : Acta Universitatis Upsaliensis, 2009. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-107492.

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Kennedy, Kathleen Anne. "Assembly of the maltose transport complex of Escherichia coli and the dimerization, localization, and functional domain structure of its ATP-binding subunit, MalK /". Thesis, Connect to this title online; UW restricted, 1999. http://hdl.handle.net/1773/11504.

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Yang, Ying. "Mechanism of metal delivery and binding to transport sites of Cu+-transporting ATPases". Link to electronic thesis, 2005. http://www.wpi.edu/Pubs/ETD/Available/etd-042905-112044/.

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Hansson, Stefan R. "The serotonin transporter and vesicular monoamine transporters during development". Lund : Lund University, 1998. http://catalog.hathitrust.org/api/volumes/oclc/68945023.html.

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Thomas, Dustin G. "Zinc transport protein-1 (ZnT1) contributes to Zn2⁺ efflux in primary cultures of rat cortical neurons". Ohio : Ohio University, 2006. http://www.ohiolink.edu/etd/view.cgi?ohiou1163544193.

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Mahato, Deependra. "Mutation of Polaris, an Intraflagellar Transport Protein, Shortens Neuronal Cilia". Thesis, University of North Texas, 2005. https://digital.library.unt.edu/ark:/67531/metadc4856/.

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Primary cilia are non-motile organelles having 9+0 microtubules that project from the basal body of the cell. While the main purpose of motile cilia in mammalian cells is to move fluid or mucus over the cell surface, the purpose of primary cilia has remained elusive for the most part. Primary cilia are shortened in the kidney tubules of Tg737orpk mice, which have polycystic kidney disease due to ciliary defects. The product of the Tg737 gene is polaris, which is directly involved in a microtubule-dependent transport process called intraflagellar transport (IFT). In order to determine the importance of polaris in the development of neuronal cilia, cilium length and numerical density of cilia were quantitatively assessed in six different brain regions on postnatal days 14 and 31 in Tg737orpk mutant and wildtype mice. Our results indicate that the polaris mutation leads to shortening of cilia as well as decreased percentage of ciliated neurons in all brain regions that were quantitatively assessed. Maintainance of cilia was especially affected in the ventromedial nucleus of the hypothalamus. Furthermore, the polaris mutation curtailed cilium length more severely on postnatal day 31 than postnatal day 14. These data suggests that even after ciliogenesis, intraflagellar transport is necessary in order to maintain neuronal cilia. Regional heterogeneity in the effect of this gene mutation on neuronal cilia suggests that the functions of some brain regions might be more compromised than others.

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Gabriel, Luke R. "Dynamic Regulation at the Neuronal Plasma Membrane: Novel Endocytic Mechanisms Control Anesthetic-Activated Potassium Channels and Amphetamine-Sensitive Dopamine Transporters: A Dissertation". eScholarship@UMMS, 2013. http://escholarship.umassmed.edu/gsbs_diss/725.

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Endocytic trafficking dynamically regulates neuronal plasma membrane protein presentation and activity, and plays a central role in excitability and plasticity. Over the course of my dissertation research I investigated endocytic mechanisms regulating two neuronal membrane proteins: the anesthetic-activated potassium leak channel, KCNK3, as well as the psychostimulant-sensitive dopamine transporter (DAT). My results indicate that KCNK3 internalizes in response to Protein Kinase C (PKC) activation, using a novel pathway that requires the phosphoserine binding protein, 14-3-3β, and demonstrates for the first time regulated KCNK3 channel trafficking in neurons. Additionally, PKC-mediated KCNK3 trafficking requires a non-canonical endocytic motif, which is shared exclusively between KCNK3 and sodium-dependent neurotransmitter transporters, such as DAT. DAT trafficking studies in intact ex vivo adult striatal slices indicate that DAT endocytic trafficking has both dynamin-dependent and –independent components. Moreover, DAT segregates into two populations at the neuronal plasma membrane: trafficking-competent and -incompetent. Taken together, these results demonstrate that novel, non-classical endocytic mechanisms dynamically control the plasma membrane presentation of these two important neuronal proteins.

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18

Lin, Xiaohui, i 林晓晖. "Molecular analysis of an iron transporter gene of Burkholderia speciesMBA4". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2009. http://hub.hku.hk/bib/B4218194X.

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Bravo, Silvina Alejandra. "The di/tri-peptide transporters PEPT1 and PEPT2 : expression and regulation in the intestinal Caco-2 and renal SKPT0193 cl.2 cell lines /". Cph. : Department of Pharmaceutics, The Danish University of Pharmaceutical Sciences, 2004. http://www.dfh.dk/phd/defences/silvinabravo.htm.

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Brault, Jeffrey J. "Creatine uptake and creatine transporter expression among rat skeletal muscle fiber types". free to MU Campus, others may purchase, 2003. http://wwwlib.umi.com/cr/mo/fullcit?p3091902.

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21

Madsen, Jacob. "Development of new PET and SPECT ligands for visualization of serotonin transporter in the brain /". Cph. : PET & Cyclotron Unit, Rigshospitalet, Copenhagen University Hospital : The Danish University Hospital, 2003. http://www.dfh.dk/phd/defences/jacobmadsen.htm.

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Woźnicka-Misăilă, Aleksandra. "An investigation and characterization of different ADP/ATP Carrier homologs". Thesis, Université Grenoble Alpes (ComUE), 2016. http://www.theses.fr/2016GREAV011/document.

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L'objectif principal de ce projet de thèse était d'obtenir de nouvelles données structurales sur les transporteurs ADP/ATP mitochondriaux et de développer des outils pour les approches de micro- et nano-cristallographie appliquées à la biologie structurale des protéines membranaires.Le rôle principal du transporteur ADP/ATP (AAC) est d'importer et d'exporter respectivement de l’ADP3- et l’ATP4- à travers la membrane mitochondriale interne, entre l'espace intermembranaire et la matrice. AAC est le transporteur mitochondrial le mieux caractérisé de toute cette famille de protéines. De nombreuses études ont été menées pour caractériser sa fonction et sa structure. Toutefois, les données structurales n’étant disponibles que pour une conformation de la protéine, de nombreuses questions fondamentales notamment sur les différents états conformationnels adoptés par la protéine au cours du processus de transport restent encore posées. Dans cette thèse, nous avons étudié les 4 isoformes humaines d’AAC. Elles sont impliquées dans diverses maladies génétiques, mais jouent également un rôle dans la cancérogenèse. Cette thèse décrit ainsi en détail la caractérisation structurale et fonctionnelle de ces protéines et leur comparaison. C’est est une étape essentielle pour définir leurs propriétés, et constitue un point de départ précieux dans le développement de nouvelles thérapies.Le domaine de la biologie structurale ne cesse de connaître de nouveaux développements, comme c’est le cas par exemple avec l’avènement de la cristallographie sérielle. Il y a donc un besoin constant de nouvelles approches notamment pour la préparation des échantillons, leur montage sur les lignes de lumière et les collectes de données afin de continuer à améliorer la qualité des données collectées au synchrotron. Ainsi, notre objectif était d'utiliser différents échantillons de protéines membranaires pour développer de nouvelles techniques de cristallisation et de montage d’échantillons sur les lignes de lumière afin de préserver au mieux la qualité des échantillons tout en permettant des collectes de données plus rapides, plus efficaces et plus simples
The main objective of this PhD project was to gain new structural data on the mitochondrial ADP/ATP carriers and develop tools for micro- and nano-crystallography approaches applied to membrane protein structural biology.The main role of the ADP/ATP carrier (AAC) is to import and export ADP3- and ATP4- respectively between the intermembrane space and the matrix through the inner mitochondrial membrane. AAC is the best characterized among all mitochondrial carriers. Much has been done to investigate its function and structure. However, since structural data are only available for one conformation of the protein some fundamental questions about the different conformational states adopted during the transport process still need to be answered.In this thesis we considered 4 human AAC homologs as a main target. They are involved in different genetic diseases but play also a role in cancerogenesis. This thesis describes and compares in detail the functional and structural characterization of the human AAC isoforms. It was an essential step to give insight into their native properties and is a precious starting point for the drug development field.Since the structural biology field is rapidly developing especially in serial crystallography techniques, there are more and more new applications for samples preparation, mounting and measurements in order to improve the quality of the data collected at the synchrotrons. Hence, our second objective was to use different membrane protein samples to develop new crystal-friendly crystallization set up combined with different sample environment on the beamline toward faster, more efficient and simpler data collection

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Blodgett, David M. "Human Erythrocyte Glucose Transporter (GLUT1) Structure, Function, and Regulation: A Dissertation". eScholarship@UMMS, 2007. https://escholarship.umassmed.edu/gsbs_diss/326.

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The structure-function relationship explains how the human erythrocyte glucose transport protein (GLUT1) catalyzes sugar transport across the plasma membrane. This work investigates the glucose transport mechanism, the structural arrangement and dynamics of GLUT1 membrane-spanning α-helices, the molecular basis for glucose transport regulation by ATP, and how cysteine accessibility contributes to GLUT1 structure. A rapid kinetics approach was applied to examine the conformational changes GLUT1 undergoes during the transport cycle. To transition from a global to molecular focus, a novel mass spectrometry technique was developed to resolve GLUT1 sequence that is associated either with membrane embedded GLUT1 subdomains or with water exposed domains. By studying accessibility changes of specific amino acids to covalent modification by a Sulfo-NHS-LC-Biotin probe, specific protein regions associated with glucose transport modulation by ATP were identified. Finally, mass spectrometry was applied to examine cysteine residue accessibility under native and reducing conditions. This thesis presents data supporting the isolation of an intermediate, occluded GLUT1 conformational state that temporally bridges import and export configurations during glucose translocation. Our results confirm that amphipathic α-helices line the translocation pathway and promote interactions with the aqueous environment and substrate. In addition, we show that GLUT1 is conformationally dynamic, undergoes reorganization in the cytoplasmic region in response to ATP modulation, and that GLUT1 contains differentially exposed cysteine residues that affect its folding.

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Follit, John A. "Building the Cell's Antenna: Protein Targeting to the Ciliary Membrane: A Dissertation". eScholarship@UMMS, 2012. https://escholarship.umassmed.edu/gsbs_diss/594.

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Protruding from the apical surface of nearly every cell in our body lies a specialized sensory organelle—the primary cilium. Eukaryotic cells use these ubiquitous structures to monitor the extracellular environment, defects in which result in an ever-growing list of human maladies termed ciliopathies including obesity, retinal degeneration and polycystic kidney disease. The sensory functions of primary cilia rely on the unique complement of receptors concentrated within the ciliary membrane. Vital to the proper functioning of the cilium is the cell's ability to target specific proteins to the ciliary membrane yet little is known how a cell achieves this highly polarized distribution. IFT20, a subunit of the intraflagellar transport particle is localized to the Golgi complex that is hypothesized to sort proteins to the ciliary membrane. We show that IFT20 is anchored to the Golgi complex by the golgin protein GMAP-210 and mice lacking GMAP210 die at birth with a pleiotropic phenotype that includes growth restriction and heart defects. Cilia on GMAP210 mutant cells have reduced amounts of the membrane protein polycystin-2 localized to them suggesting IFT20 and GMAP-210 function together in the sorting or transport of proteins to the ciliary membrane. To better understand the mechanism of ciliary protein trafficking, we identify a ciliary targeting sequence (CTS) contained within fibrocystin, the gene mutated in autosomal recessive polycystic kidney disease, and investigate a series of proteins required for the delivery of this sequence to the primary cilium. We demonstrate the small G protein Rab8 interacts with the CTS of fibrocystin and controls the ciliary levels of the CTS. Arf4 is another small G protein deemed a key regulator of ciliary protein trafficking. We show Arf4 binds the CTS of fibrocystin but is not absolutely required for trafficking of the fibrocystin CTS to cilia. Arf4 mutant mice are embryonic lethal and die at mid-gestation likely due to defects in the non-ciliated visceral endoderm, where the lack of Arf4 caused defects in cell structure and apical protein localization. This suggests Arf4 is not only important for the efficient transport of fibrocystin to cilia, but also plays critical roles in non-ciliary processes. Together this work aims to elucidate the mechanisms of protein targeting to the ciliary membrane.

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25

Lin, Xiaohui. "Molecular analysis of an iron transporter gene of Burkholderia species MBA4". Click to view the E-thesis via HKUTO, 2009. http://sunzi.lib.hku.hk/hkuto/record/B4218194X.

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Reynolds, Annie 1978. "Over-expression of the potassium-chloride co-transporter KCC2 in developing zebrafish". Thesis, McGill University, 2006. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=98778.

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In embryonic neurons, the intracellular chloride concentration is elevated, making GABA and glycine depolarizing. Later in development, coincident with neuronal maturation, the extruding potassium-chloride co-transporter KCC2 is expressed. It reverses the chloride gradient, rendering it hyperpolarizing. Early depolarization is assumed to play trophic roles during nervous system development. I thus decided to investigate the effects of the depolarizing chloride gradient on development in vivo in the zebrafish embryo. I first determined the temporal pattern of KCC2 expression in zebrafish and found it was absent in the embryo. I then over-expressed wild-type, gain-of-function and loss-of-function variants of human KCC2, using GFP-tagged constructs for detection purposes. Over-expression of functional hKCC2 perturbed the morphology and motor behaviours of the embryos. At the cellular level, KCC2 impaired axonal growth and affected the neuronal populations in the brain, hindbrain and spinal cord. This suggests the depolarizing effects of glycine are critical for neurogenesis.

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Stange, Christoph. "Combining artificial Membrane Systems and Cell Biology Studies: New Insights on Membrane Coats and post-Golgi Carrier Formation". Doctoral thesis, Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2013. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-102218.

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In mammalian cells, homeostasis and fate during development relies on the proper transport of membrane-bound cargoes to their designated cellular locations. The hetero-tetrameric adaptor protein complexes (APs) are required for sorting and concentration of cargo at donor membranes, a crucial step during targeted transport. AP2, which functions at the plasma membrane during clathrin-mediated endocytosis, is well characterized. In contrast, AP1 a clathrin adaptor mediating the delivery of lysosomal hydrolases via mannose 6-phosphate receptors (MPRs) and AP3 an adaptor ensuring the proper targeting of lysosomal membrane protein are difficult to study by classic cell biology tools. To gain new insights on these APs, our lab has previously designed an in vitro system. Reconstituted liposomes were modified with small peptides mimicking the cytosolic domains of bona fide cargoes for AP1 and AP3 respectively and thereby enabling the selective recruitment of these APs and the identification of the interacting protein network. In the study at hand we utilize above-described liposomes to generate supported lipid bilayers and Giant Unilamellar Vesicles (GUVs), large-scale membrane systems suited for analysis by fluorescence microscopy. By using cytosol containing fluorescently-tagged subunits, we visualized clathrin coats on artificial membranes under near physiological conditions for the first time. Moreover, we demonstrated clathrin-independent recruitment of AP3 coats on respective GUVs. Presence of active ARF1 was sufficient for the selective assembly of AP1-dependent clathrin coats and AP3 coats on GUVs. By using dye-conjugated ARF1, we show that ARF1 colocalized with AP3 coats on GUVs and that increased association of ARF1 with GUVs coincided with AP1-dependent clathrin coats. Our previous study identified members of the septin family together with AP3 coats on liposomes. Here we show on GUVs, that active ARF1 stimulated the assembly of septin7 filaments, which may constrain the size and mobility of AP3 coats on the surface. Subsequent cell biology studies in HeLa cells linked septins to actin fibers on which they may control mobility of AP3-coated endosomes and thus their maturation. An actin nucleation complex, based on CYFIP1 was identified together with AP1 on liposomes before. Here we show on GUVs, that CYFIP1 is recruited on the surface surrounding clathrin coats. Upon supply of ATP, sustained actin polymerization generated a thick shell of actin on the GUV surface. The force generated by actin assembly lead to formation of long tubular protrusions, which projected from the GUV surface and were decorated with clathrin coats. Thereby the GUV model illustrated a possible mechanism for tubular carriers formation. The importance of CYFIP1-reliant actin polymerization for the generation of MPR-positive tubules at the trans-Golgi network (TGN) of HeLa cells was subsequently demonstrated in our lab. The notion that tubulation of artificial membranes could be triggered by actin polymerization allowed us to perform a comparative mass spectrometry screen. By comparing the abundance of proteins on liposomes under conditions promoting or inhibiting actin polymerization, candidates possibly involved in stabilization, elongation or fission of membrane tubules could be identified. Among the proteins enriched under conditions promoting tubulation, we identified type I phosphatidylinositol-4-phosphate 5-kinases. Their presence suggested an involvement of phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) in tubule formation. By cell biology studies in HeLa we show, that down regulation of these enzymes altered the dynamics of fluorescently-tagged MPRs, illustrating the importance of locally confined PI(4,5)P2 synthesis during formation of coated carriers at the TGN. Bin–Amphiphysin–Rvs (BAR) domains are known to sense membrane curvature and induce membrane tubulation. Among various BAR domain proteins, Arfaptin2 was enriched under conditions allowing tubulation of liposomes. By microscopy studies on HeLa cells we show, that Arfaptin2 as well as its close paralog Arfaptin1 were present on AP1-coated MPR tubules emerging from the TGN. We further show, that tubule fission occurred at regions were Arfaptin1 is concentrated and that simultaneous down regulation of both Arfaptins lead to increased number and length of MPR tubules. Since fission of coated transport intermediates at the TGN is poorly understood, our findings contribute a valuable component towards a model describing the entire biogenesis of coated post-Golgi carriers. In conclusion, combining artificial membrane systems and cell biology studies allowed us to propose new models for formation as wall as for fission of AP1-coated transport intermediates at the TGN. Further we gained new insights on AP3 coats and the possible involvement of septin filaments in AP3-dependent endosomal maturation.

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Brohée, Sylvain. "Etude bioinformatique du réseau d'interactions entre protéines de transport ches les Fungi". Doctoral thesis, Universite Libre de Bruxelles, 2008. http://hdl.handle.net/2013/ULB-DIPOT:oai:dipot.ulb.ac.be:2013/210432.

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Les protéines associées aux membranes sont d'une importance cruciale pour la cellule. Cependant, en raison d'une plus grande difficulté de manipulation, les données biochimiques les concernant sont très lacunaires, notamment au point de vue de la formation de complexes entre ces protéines.

L'objectif global de notre travail consiste à combler ces lacunes et à préciser les interactions entre protéines membranaires chez la levure Saccharomyces cerevisiae et plus précisément, entre les transporteurs. Nous avons commencé notre travail par l'étude d'un jeu de données d'interactions à grande échelle entre toutes les perméases détectées par une méthode de double hybride spécialement adaptée aux protéines insolubles (split ubiquitin). Premièrement, la qualité des données a été estimée en étudiant le comportement global des données et des témoins négatifs et positifs. Les données ont ensuite été standardisées et filtrées de façon à ne conserver que les plus significatives. Ces interactions ont ensuite été étudiées en les modélisant dans un réseau d'interactions que nous avons étudié par des techniques issues de la théorie des graphes. Après une évaluation systématique de différentes méthodes de clustering, nous avons notamment recherché au sein du réseau des groupes de protéines densément interconnectées et de fonctions similaires qui correspondraient éventuellement à des complexes protéiques. Les résultats révélés par l'étude du réseau expérimental se sont révélés assez décevants. En effet, même si nous avons pu retrouver certaines interactions déjà décrites, un bon nombre des interactions filtrées semblait n'avoir aucune réalité biologique et nous n'avons pu retrouver que très peu de modules de protéines de fonction semblable hautement inter-connectées. Parmi ceux-ci, il est apparu que les transporteurs d'acides aminés semblaient interagir entre eux.

L'approche expérimentale n'ayant eu que peu de succès, nous l'avons contournée en utilisant des méthodes de génomique comparative d'inférence d'interactions fonctionnelles. Dans un premier temps, malgré une évaluation rigoureuse, l'étude des profils phylogénétiques (la prédiction d'interactions fonctionnelles en étudiant la corrééélation des profils de présence - absence des gènes dans un ensemble de génomes), n'a produit que des résultats mitigés car les perméases semblent très peu conservées dès lors que l'on considère d'autres organismes que les \
Doctorat en Sciences
info:eu-repo/semantics/nonPublished

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Hagan, G. Nana. "Adipocyte Insulin-Mediated Glucose Transport: The Role of Myosin 1c, and a Method for in vivo Investigation: A Dissertation". eScholarship@UMMS, 2008. https://escholarship.umassmed.edu/gsbs_diss/403.

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The importance of insulin delivery and action is best characterized in Type 2 Diabetes, a disease that is becoming a pandemic both nationally and globally. Obesity is a principal risk factor for Type 2 Diabetes, and adipocyte function abnormalities due to adipose hypertrophy and hyperplasia, have been linked to obesity. Numerous reports suggest that the intracellular and systemic consequences of adipocyte function abnormalities include adipocyte insulin resistance, enhanced production of free fatty acids, and production of inflammatory mediators. A hallmark of adipocyte insulin sensitivity is the stimulation of glucose transporter isoform 4 (GLUT4) trafficking events to promote glucose uptake. In the Type 2 diabetic and insulin resistant states the mechanism behind insulin-stimulated GLUT4 trafficking is compromised. Therefore, understanding the role of factors involved in glucose-uptake in adipose tissue is of great importance. Studies from our laboratory suggest an important role for the unconventional myosin, Myo1c, in promoting insulin-mediated glucose uptake in cultured adipocytes. Our observations suggest that depletion of Myo1c in cultured adipocytes results in a significant reduction in the ability of adipocytes to take up glucose following insulin treatment, suggesting Myo1c is required for insulin-mediated glucose uptake. A plausible mechanism by which Myo1c promotes glucose uptake in adipocytes has been suggested by further work from our laboratory in which expression of fluorescently-tagged Myo1c in cultured adipocytes induces significant membrane ruffling at the cell periphery, insulin-independent GLUT4 translocation to the cell periphery, and accumulation of GLUT4 in membrane ruffling regions. Taken together Myo1c seems to facilitate glucose uptake through remodeling of cortical actin. In the first part of this thesis I, in collaboration with others, uncovered a possible mechanism through which Myo1c regulates adipocyte membrane ruffling. Here we identified a novel protein complex in cultured adipocytes, comprising Myo1c and the mTOR binding partner, Rictor. Interestingly our studies in cultured adipocytes suggest that the Rictor-Myo1c complex is biochemically distinct from the Rictor-mTOR complex of mTORC2. Functionally, only depletion of Rictor but not Myo1c results in decreased Akt phosphorylation at serine 473, but depletion of either Rictor or Myo1c results in compromised cortical actin dynamic events. Furthermore we observed that whereas the overexpression of Myo1c in cultured adipocytes causes remarkable membrane ruffling, Rictor depletion in cells overexpressing Myo1c significantly reduces these ruffling events. Taken together our findings suggest that Myo1c, in conjunction with Rictor, modulates cortical actin remodeling events in cultured adipocytes. These findings have implications for GLUT4 trafficking as GLUT4 has been previously observed to accumulate in Myo1c-induced membrane ruffles prior to fusion with the plasma membrane. During our studies of adipocyte function we noticed that current siRNA electroporation methods present numerous limitations. To silence genes more effectively we employed a lentivirus-mediated shRNA delivery system, and to standardize this technology in cultured adipocytes we targeted Myo1c and MAP4K4. Using this technology we were able to achieve clear advantages over siRNA oligonucleotide electroporation techniques in stability and permanence of gene silencing. Furthermore we showed that the use of lentiviral vectors in cultured adipocytes did not affect insulin signaling or insulin-mediated glucose uptake events. Despite our inability to use lentiviral vectors to achieve gene silencing in mice we were able to achieve adipose tissue-specific gene silencing effects in mice following manipulation of the lentiviral conditional silencing vector, and then crossing resulting founders with aP2-Cre mice. Interestingly however, only founders from the MAP4K4 conditional shRNA vector, but not founders from the Myo1c conditional shRNA vector, showed gene knockdown, possibly due to position-effect variegation. Taken together, findings from these studies are important because they present an alternative means of achieving gene silencing in cultured adipocytes, with numerous advantages not offered by siRNA oligonucleotide electroporation methods. Furthermore, the in vivo, adipose tissue-specific RNAi studies offer a quick, inexpensive, and less technically challenging means of achieving adipose tissue-specific gene ablations relative to traditional gene knockout approaches.

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Sales, Maria Carmen Oliveira Pinho de. "Regulação da expressão do transportador de aminoácidos de Leishmania (Leishmania) amazonensis". Universidade de São Paulo, 2014. http://www.teses.usp.br/teses/disponiveis/99/99131/tde-23022015-081238/.

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Leishmania caracteriza-se por apresentar duas formas morfologicamente distintas em seu ciclo de vida: promastigotas e amastigotas. As formas promastigotas vivem tubo digestório do vetor flebotomíneo, sob as condições de pH 7,0 e temperatura ambiente, ao redor de 25ºC. As formas amastigotas são encontradas no interior dos fagolisossomos de macrófagos infectados onde encontram um ambiente de pH ácido e temperatura ao redor de 34ºC. Leishmania utiliza arginina para a síntese de poliaminas, que desempenham papel fundamental no crescimento, diferenciação celular e sucesso da infecção. A tomada de arginina em L. (L.) amazonensis é feita pela proteína transportadora de aminoácidos - amino acid transporter-like 3 (AAP3), codificada por duas cópias do gene (5.1 aap3 e 4.7 aap3) dispostas em tandem no genoma. Os transcritos de 5.1 aap3 e de 4.7 aap3 apresentam 98% de identidade entre as ORFs, mas diferem nas 5\' e 3\' UTR. O objetivo do presente trabalho foi avaliar se os sinais de temperatura, pH e privação de arginina disparam a regulação da expressão de aap3 em formas promastigotas e amastigotas. Para isso avaliamos o nível dos transcritos e realizamos ensaios de tomada de arginina em células submetidas à privação ou suplementadas com arginina, nas temperaturas de 25°C ou 34°C em pH 7,0 ou 5,0. Constatou-se em formas promastigotas que o transcrito 5.1 aap3 apresentou maior abundância em relação a 4.7 aap3, e que a privação promoveu o aumento da tomada do aminoácido quando os parasitos eram mantidos em pH 7,0 a 25°C, corroborando dados anteriores do nosso grupo. Demonstramos que a mudança de temperatura foi um fator importante para o aumento do número de cópias de 5.1 aap3 em promastigotas privadas, principalmente quando associadas com o pH 5,0. Além disso, o aumento da temperatura favoreceu a tomada de arginina, corroborando com a elevação do número de cópias observada para o transcrito 5.1 aap3. Em amastigotas-like, mantidas a 25°C e pH 7,0 a privação reverteu a expressão de 5.1 aap3 para o mesmo perfil observado para promastigotas. Contudo, não observamos um favorecimento na tomada de arginina. Ainda em amastigotas, o tratamento a 34°C e pH 7,0 favoreceu a tomada de arginina, porém não observamos um aumento correspondente na quantificação do transcrito. O transcrito 4.7 aap3 não apresentou alteração significativa em qualquer tratamento em promastigotas e amastigotas. Os nossos resultados indicam que a variação de temperatura e do pH, além da privação de arginina, podem ser sinais importantes para regulação da expressão diferencial de aap3, principalmente a cópia 5.1 aap3, de forma a assegurar a oferta de arginina em formas promastigotas previamente à entrada no hospedeiro mamífero e em formas amastigotas, na passagem para o vetor, assegurando o sucesso da infecção
Leishmania presents two morphologically distinct forms in its life cycle: promastigote and amastigote. The promastigotes live in the midgut of the sand fly vector under the conditions of pH 7.0 and room temperature, around 25°C. The amastigote forms are found inside the phagolysosomes of infected macrophages where they encounter an environment of acidic pH and temperature around 34°C. Leishmania uses the arginine to synthesize polyamines which play an important role in cell growth, differentiation and in the successful of infection. The arginine uptake in Leishmania (L.) amazonensis is made by an amino acid porter 3-like protein (AAP3), coded by a two copies gene (5.1 aap3 e 4.7 aap3) arranged in tandem in the genome. The transcripts, 5.1 aap3 and 4.7 aap3, present 98% of ORFs identity, but differ in the 5\' and 3\' UTR. The aim of this work was to evaluate if canges in temperature, pH and arginine deprivation represent signals to trigger the regulation expression of aap3 in promastigotes and amastigotes. For this, we evaluated the transcripts level and performed assays of arginine uptake in parasites subjected to arginine starvation or supplemented with arginine, at temperatures of 25°C or 34°C and pH 7.0 or 5.0. In promastigotes we verified that the transcript 5.1 aap3 showed higher abundance in relation to 4.7 aap3, and that the arginine starvation promoted an increase in the amino acid uptake when the parasites were maintained at pH 7.0 and 25°C, confirming previous data from our group. The change of temperature was an important factor to the increase of 5.1 aap3 transcripts - in starved promastigotes, particularly when associated with pH 5.0. In addition, the increase of the temperature led to an increase of the arginine uptake, correlated to the increase of 5.1 aap3 transcript. The amastigotes-like maintained at 25°C and pH 7.0 and submitted to the amino acid starvation reverted 5.1 aap3 expression profile to the same observed for promastigotes. However, those condictions did not favor an increase in arginine uptake. The treatment of amastigotes at 34°C and pH 7.0 facilitated the increased of arginine uptake, but did not correlated with the transcripts level. The 4.7 aap3 transcript did not change significantly in any promastigotes and amastigotes treatments. Our results indicate that variation in temperature and pH, in addition to arginine starvation may be important signals for regulating the aap3 expression, mainly the copy 5.1 aap3, in order to ensure the correct supply of arginine in the previous period in relation to the entry of the promastigotes into the mammalian host or to the amastigotes transition in the vector, ensuring the success of the infection.

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31

Abbadi, Mehdi. "Synthèse de marqueurs de la protéine de transport du D-glucose". Université Joseph Fourier (Grenoble), 1999. http://www.theses.fr/1999GRE10001.

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De nombreuses pathologies comme le diabete ou la maladie d'alzheimer s'accompagnent de variations significatives de la consommation en d-glucose et vraisemblablement de variations de la densite membranaire des transporteurs du glucose (glut) correspondants. Il apparait ainsi souhaitable de pouvoir disposer d'outils permettant de detecter ces variations par denombrement des transporteurs du glucose, si possible in vivo. Dans un premier temps, des analogues du glucose et de l'acide-l-ascorbique (qui emprunte egalement les proteines glut) ont ete synthetises afin de mieux comprendre la nature des modifications pouvant etre apportees a ces composes sans que leur reconnaissance par les transporteurs glut ne soit affectee. La synthese de sondes pouvant permettre d'evaluer la densite membranaire des proteines glut a ensuite ete effectuee. Un premier type associe une detection externe (radiomarquage a l'iode) a un marquage d'affinite (fonction isothiocyanate) tandis que chez le second, deux glucoses sont relies entre eux par une chaine a partir de la face non-necessaire a la reconnaissance de glut ; le schema de synthese retenu permet d'acceder rapidement a des sondes pour un marquage radioactif d'une part ou par photo-affinite d'autre part, des proteines glut. L'ensemble des composes synthetises au cours de ce travail a ete evalue sur modeles biologiques (collaboration avec le service de medecine nucleaire du chu de grenoble et avec le departement de biochimie de l'universite de bath (grande-bretagne)). Parmi ces derives, une sonde de type bis-glucose s'est averee interessante puisqu'elle presente une affinite pour les proteines glut dix fois meilleure que celle du glucose lui-meme.

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Harnevik, Lotta. "Molecular genetic studies on cystinuria". Doctoral thesis, Linköping : Univ, 2007. http://www.bibl.liu.se/liupubl/disp/disp2007/med1034s.pdf.

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Phelps, Mitch A. "Novel approaches for characterizing the riboflavin transport and trafficking mechanism and its potential as a target in breast cancer". Columbus, Ohio : Ohio State University, 2005. http://rave.ohiolink.edu/etdc/view?acc%5Fnum=osu1133261831.

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França, Moana Rodrigues. "Perfis endócrinos peri-ovulatórios influenciam o transporte, metabolismo e disponibilidade de aminoácidos no lúmen uterino de vacas de corte durante o diestro inicial". Universidade de São Paulo, 2016. http://www.teses.usp.br/teses/disponiveis/10/10131/tde-21032017-151159/.

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Em vacas de corte, folículos pré-ovulatórios (FPO) maiores, maiores concentrações de estradiol (E2) no proestro/estro e progesterona (P4) no diestro favorecem o crescimento do concepto e a fertilidade. Porém, os mecanismos mediados pelos esteroides femininos que influenciam a receptividade uterina ao embrião precisam ser esclarecidos. Os aminoácidos (AA) são componentes das secreções uterinas que são cruciais para a sobrevivência do embrião antes da implantação. A hipótese deste trabalho é que o tamanho do FPO e as concentrações de E2 e P4 modulam a abundância de transcritos relacionados ao transporte e metabolismo de AA no endométrio e afetam a concentração luminal de AA. Para isso, o crescimento folicular de vacas Nelore foi manipulado com o objetivo de formar dois grupos: FPO grande e CL grande (FG-CLG) e FPO pequeno e CL pequeno (FP-CLP). No Dia 4 (D4; Exp 1) e Dia 7 (D7; Exp 2) após a injeção de GnRH para indução da ovulação, foram coletados tecido endometrial e lavado uterino post-mortem. A abundância de transcritos foi avaliada por qRT-PCR e as concentrações de AA nos lavados foram quantificadas por HPLC. No Exp 1, o tamanho do FPO, concentrações plasmáticas de E2 no D-1 e de P4 no D4 foram 15,70mm±0,43 vs. 11,31mm±0,23 (p<0,01), 2,44pg/ml±0,19 vs. 0,65pg/ml (p<0,01) e 1,40ng/ml±0,23 vs. 0,80ng/ml±0,10 (p<0,01) para os grupos FG-CLG vs. FP-CLP, respectivamente. No Exp 2, o tamanho do FPO, concentrações plasmáticas de E2 no D-1 e de P4 no D7 foram 13,18mm±0,44 vs. 10,63mm±0,30 (p<0,01), 2,30pg/ml±0,57 vs. 0,50pg/ml±0,13 (p<0,01) e 3,68ng/ml±0,38 vs. 2,49ng/ml±0,43 (p=0,04) para os grupos FG-CLG vs. FP-CLP, respectivamente. No D4 a abundância de SLC1A4, SLC38A1, SLC6A6, SLC7A4 e SLCY e no D7, SLC1A4, SLC6A1, SLC6A14, SLC7A4, SLC7A8, SLC38A1, SLC38A7, SLC43A2 e DDO foi maior no endométrio dos animais do grupo FG-CLG (p<0,05). No D4, maiores concentrações de taurina, alanina e ácido α-aminobutírico foram observadas no grupo FP-CLP (p<0,05). Em contraste, menores concentrações de valina e cistationina foram encontradas nos lavados uterinos do D7 dos animais do grupo FP-CLP (p<0,05). No D4, os animais do grupo FG-CLG, associado a maior fertilidade, apresentaram menor quantidade de AA nas secreções uterinas, porém, a abundância dos transportadores de AA foi compatível com maior transporte em comparação aos animais do grupo FP-CLP. Esses resultados sugerem que antes do embrião se mover do oviduto ao útero, o transporte e metabolismo dos AA prioriza a preparação das células endometriais para receber o embrião e não o acúmulo nas secreções uterinas. Porém, no D7, quando o embrião está em contato direto com as secreções uterinas, os genes relacionados ao transporte de AA no endométrio e a concentração de AA no histotrofo são estimulados nas vacas do grupo FG-CLG. Portanto o metabolismo e transporte de AA no sentido das células endometriais ou das secreções uterinas pode ser um mecanismo importante para a receptividade materna.
In beef cattle, a large size of the pre-ovulatory follicle (POF) and resulting elevated proestrus/estrus estradiol (E2) and diestrus progesterone (P4) concentrations positively affect conceptus growth and fertility. However, sex-steroid-mediated mechanisms that influence uterine receptivity to the embryo need to be elucidated. Amino acids are important components of maternally-derived secretions that are crucial for embryo survival before implantation. The hypothesis is that the size of the POF, E2 and P4 concentrations modulate endometrial abundance of solute carrier proteins (SLC) transcripts related to AA transport and metabolism and subsequently affect lumenal amino acids concentrations. Therefore, follicle growth of Nelore cows was manipulated to produce two experimental groups: large POF and CL (LF-LCL group) and small POF and CL (SF-SCL group). On Day 4 (D4; Experiment 1) and Day 7 (D7; Experiment 2) post GnRH injection to induce ovulation, endometrial tissue and uterine washings were collected post-mortem. Transcript abundance was evaluated by qRT-PCR and amino acid concentrations were quantified in washings by HPLC. On Experiment 1, POF size, plasma E2 concentration on D-1, and plasma concentration of P4 on D4 were 15.70mm±0.43 vs. 11.31mm±0.23 (p<0.01), 2.44pg/ml±0.19 vs. 0.65pg/ml (p<0.01) and 1.40ng/ml±0.23 vs. 0.80ng/ml±0.10 (p<0.01) for the LF-LCL vs. SF-SCL groups, respectively. For Experiment 2, POF size, plasma E2 concentration on D-1 and plasma P4 concentration on D7 were 13.18mm±0.44 vs. 10.63mm±0.30 (p<0.01), 2.30pg/ml±0.57 vs. 0.50pg/ml±0.13 (p<0.01) and 3.68ng/ml±0.38 vs. 2.49ng/ml±0.43 (p=0.04) for the LF-LCL vs. SF-SCL groups, respectively. On D4, abundance of SLC6A6, SLC7A4, SLC17A5, SLC38A1, SLC38A7 and SLCY and on D7, SLC1A4, SLC6A1, SLC6A14, SLC7A4, SLC7A7, SLC7A8, SLC17A5, SLC38A1, SLC38A7, SLC43A2 and DDO was greater in the endometrium of cows from the LF-LCL group (p<0.05). On D4, higher concentrations of taurine, alanine and α-aminobutiric acid were observed in SF-SCL (p<0.05). In contrast, lower concentrations of valine and cystathionine were quantified in D7 uterine washings from SF-SCL cows (p<0.05). On D4, animals from LF-LCL group, associated with greater fertility, presented less amino acid content in uterine secretion but abundance of transporters was compatible to greater transport in comparison to animals from SF-SCL group. This suggests that before embryo moves from oviduct to uterus, amino acids transport and metabolism pathways prioritizes endometrium cells preparation for receiving the embryo but not accumulation in uterine secretions. However, on D7, when the embryo is in direct contact with uterine secretions, genes related to amino acids transport in endometrium and amino acids concentration in histotroph are up-regulated in LF-LCL cows. The latter insights indicate that amino acids metabolism and transport, towards endometrial cells or uterine secretions, might be mechanisms contributing to maternal receptivity.

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Benba, Jamila. "Contribution à l'étude du système de transport des dicarboxylates des mitochondries; purification, caractérisation". Rouen, 1993. http://www.theses.fr/1993ROUES053.

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Le travail presenté dans ce mémoire a été entrepris dans le but de purifier et de caractériser le transporteur des dicarboxylates des mitochondries, qui catalyse l'échange un contre un des dicarboxylates entre eux (malate, malonate, succinate) ou contre le phosphate. Cette étude a été effectuée simultanément sur les mitochondries de foie de rat et sur les mitochondries d'une souche de levure: Saccharomyces cerevisiae. Les mitochondries de levure ont été solubilisées par le Triton X-100 et l'extrait a été chromatographié sur hydroxyapatite. Le filtrat, traité par électrophorèse en milieu dénaturant, a révélé par coloration à l'argent la présence d'environ 5 protéines de Mr compris entre 28000 et 35000. Après reconstitution de l'activité de transport dans des liposomes, un accroissement de 10 fois, de l'activité spécifique d'échange a été observé. Le passage des protéines du filtrat d'hydroxyapatite sur une colonne de malate deshydrogénase mitochondriale (EC 1. 1. 1. 37) immobilisée sur Sépharose, a conduit à la purification complète du transporteur. Cette protéine, purifiée à partir des deux types de mitochondries, possède toutes les propriétés caractéristiques du transporteur in situ et présente, en milieu dénaturant, un poids moléculaire de 28000. L'activité du transporteur purifié à partir des mitochondries de levure, reconstituée dans des liposomes, montre un Km pour le succinate de 2 mM et un Vmax de 1,5 lmol. Min-1. Mg-1 protéines tandis que l'activité spécifique augmente de 300 fois par rapport à l'extrait de départ. L'activité d'échange du transporteur reconstituée des deux types de mitochondries est inhibée par un réactif des groupements SH, le p-chloromercuriphénylsulfonate et par un réactif des acides aminés, le phosphate de pyridoxal, suggérant l'implication d'un ou plusieurs groupements SH et d'un ou plusieurs résidus lysine dans le mécanisme catalytique du transporteur. L'analyse de la composition en acides aminés de la protéine de transport des mitochondries de foie de rat indique qu'il s'agit d'une protéine hydrophobe, légèrement acide, avec la partie N-terminale bloquée

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Duvic, Bernard. "Purification et etude de la proteine de transport de la 20-hydroxyecdysone chez un insecte : locusta migratoria". Université Louis Pasteur (Strasbourg) (1971-2008), 1988. http://www.theses.fr/1988STR13160.

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La purification du transporteur est realisee a partir d'hemolymphe par differentes techniques (coagulation a temperature ambiante; chromatographie sur colonne echangeuse d'anions; chromatographie sur colonne de gel permeation; hplc sur phase echangeuse d'anions). Le transporteur est une glycoproteine acide (pi=5,6) de 280 kda. C'est un homodimere a 2 sous-unites de 140 kda dont la sequence n-terminale a ete determinee. Un polyserum de lapin et 2 anticorps monoclonaux de souris ont ete obtenus et permettent un dosage elisa du transporteur. Des dosages chez l. M. Montrent que le transporteur et l'ecdysone varient dans le meme sens. L'injection massive d'ecdysone suggere une synthese du transporteur sous le controle des ecdysteroides

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Fernandes, Carla Freire Celedonio. "Molecular characterization and expression of two new members of the SLC10 transporter family: SLC10A4 and SLC10A5". Giessen VVB Laufersweiler, 2007. http://d-nb.info/987398539/04.

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Desruet, Marie-Dominique. "Étude de traceurs radioactifs pour l'évaluation in vivo de la densité des transporteurs membranaires du glucose". Université Joseph Fourier (Grenoble), 1996. http://www.theses.fr/1996GRE10251.

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Le glucose, principale source d'energie des cellules, penetre a l'interieur de la cellule par l'intermediaire d'une proteine membranaire appelee transporteur du glucose. Or des variations de la densite membranaire des transporteurs du glucose ont ete observees dans certaines pathologies du metabolisme du glucose, en endocrinologie, neurologie et cancerologie. Un radioligand des transporteurs du glucose, marque a l'iode 123, emetteur gamma, permettrait de mettre en evidence des variations quantitatives des transporteurs du glucose in vivo. Dans le but de mettre au point un tel traceur radioactif, trois composes ont ete synthetises: 4,6-oegi, 4,6-obzgi et mi. Une fois synthetises, ces derives ont ete radiomarques et testes sur differents modeles biologiques: le 4,6-oegi et le 4,6-obzgi penetrent dans la cellule par diffusion passive sans interaction evidente avec la proteine de transport ; de plus, le 4,6-obzgi s'est avere cytotoxique. Quant a mi, son affinite pour le transporteur du glucose est trop faible pour permettre des etudes de denombrement. Aucun marqueur de reference des transporteurs du glucose in vivo n'est actuellement disponible ; et bien qu'aucun des composes ne puisse etre utilise comme marqueur des transporteurs du glucose, une premiere approche du developpement d'un radioligand marqueur des transporteurs du glucose et utilisable in vivo a ete abordee dans ce travail.

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Adrait, Annie. "Etudes spectroscopiques et magnétiques de la protéine Fur (Ferric Uptake Regulation) d'Escherichia coli". Université Joseph Fourier (Grenoble), 1998. http://www.theses.fr/1998GRE10212.

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La proteine fur (ferric uptake regulation), homodimere de 2 fois 17 kda, est impliquee dans le controle de la concentration intracellulaire du fer chez les bacteries gram negative telle que escherichia coli. Ce controle est essentiel car, bien que necessaire a la croissance de pratiquement toutes les cellules, le fer est aussi un element potentiellement toxique puisqu'il peut catalyser la formation de radicaux libres. Un mecanisme est propose selon lequel fur lie l'ion fe(ii) et, ainsi activee, peut se fixer sur la region promotrice de genes impliques dans le transport du fer, inhibant de ce fait la transcription. La presence du metal est essentielle a l'activite. In vitro, plusieurs cations divalents comme mn(ii) ou co(ii) peuvent remplacer fe(ii) pour conduire a la fixation specifique de fur sur l'adn. Toutefois, la stoechiometrie metal/monomere et l'environnement du metal n'ont pas ete jusque la clairement identifies. Notre objectif est de caracteriser les sites metalliques de la proteine et d'identifier les ligands du metal. Pour cela les proteines fur substituees au manganese et au cobalt ont ete preparees et etudiees par plusieurs techniques spectroscopiques et magnetiques, chaque metal representant une sonde structurale interessante grace a ses proprietes spectroscopiques. L'incorporation du metal, suivie par spectroscopie d'absorption electronique pour le co(ii) et rpe a temperature ambiante pour le mn(ii), a permis de montrer la presence d'un site metallique principal par monomere. La rpe et les mesures d'aimantation montrent que l'ion metallique est present a l'etat haut spin dans un environnement hexacoordonne deforme pour l'ion co(ii) (s = 3/2) et peu ou pas deforme dans le cas de l'ion mn(ii) (s = 5/2). Un site secondaire a zinc de type structural a egalement ete caracterise par ailleurs. Les experiences rmn de la forme co(ii)-fur ont permis d'identifier les protons n-h echangeables d'histidines et egalement des signaux correspondant a des protons ch#2 de ligands carboxylates. Au moins deux histidines (peut-etre trois) et au moins un aspartate ou glutamate sont ligands, la sphere de coordination est complete par des atomes n/o mais il n'y a pas de cysteine ligand du cobalt.

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Iden, Janne. "Assoziation der 2319-A-Variante des Dopamin-Transporter-Gens (DAT1) mit geringer Alkoholtoleranz /". Berlin : Lehmanns Media, 2008. http://d-nb.info/991659619/04.

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Goetz, David Henry. "Neutrophil gelatinase-associated lipocalin : a protein involved in immune defense against microbial pathogens /". Thesis, Connect to this title online; UW restricted, 2002. http://hdl.handle.net/1773/4991.

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Matsui, Yuto. "Visualization of procollagen IV reveals ER-to-Golgi transport by ERGIC-independent carriers". Kyoto University, 2020. http://hdl.handle.net/2433/259728.

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Roux, Pierre. "Le transporteur mitochondrial des adénine-nucléotides de la levure Saccharomyces cerevisiae : relations structure-fonction". Université Joseph Fourier (Grenoble), 1997. http://www.theses.fr/1997GRE10228.

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Le transporteur des adenine-nucleotides est une proteine de la membrane mitochondriale interne qui assure l'import d'adp cytosolique en echange d'atp neosynthetise dans la matrice mitochondriale. Ce systeme est inhibe de maniere tres specifique par le carboxyatractyloside (catr) et l'acide bongkrekique (ba). Les travaux presentes dans ce memoire se rapportent a la dynamique structurale du transporteur mitochondrial des adenine-nucleotides (ancp). Pour cette etude, nous avons mis en uvre le transporteur anc2p de la levure saccharomyces cerevisiae modifie pour porter une extension poly-histidine a son extremite c-terminale. Cette proteine et ses formes mutees par substitution sequentielle des tryptophanes par des tyrosines ont ete purifiees en presence de detergent par chromatographie d'affinite sur une resine portant des ions metalliques immobilises. L'innocuite des mutations et de l'extension poly-histidine a ete controlee au prealable. Nous avons etudie l'accessibilite des trois tryptophanes du transporteur isole en presence de detergent, en appliquant la technique d'extinction de fluorescence. Les resultats obtenus nous ont permis de mettre en evidence la coexistence de deux familles de monomeres du transporteur de structures differentes et de surcroit modifiees par la presence de catr ou de ba. Grace a des modifications chimiques selectives des differents tryptophanes, nous avons pu determiner le degre d'exposition au milieu aqueux de chacun de ces residus au sein des monomeres et en mesurer les variations lors des changements de conformation du transporteur. Enfin, nous avons mis en uvre une approche fluorimetrique par mesure de transfert d'energie d'activation pour evaluer les distances separant les residus de tryptophane du transporteur. A partir de l'ensemble de ces donnees, nous proposons un modele dynamique d'arrangement du transporteur dans la membrane mitochondriale.

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Duplay, Pascale. "Approche genetique de la topologie fonctionnelle de la proteine affine du maltose chez escherichia coli". Paris 7, 1987. http://www.theses.fr/1987PA077001.

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DeWit, Ralph. "Multimodal transport : carrier liability and documentation /". London [u.a.] : Lloyd's of London Press, 1995. http://www.gbv.de/dms/spk/sbb/recht/toc/278603084.pdf.

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Watling, Jeremy Richards. "Carrier transport in quantum well structures". Thesis, University of East Anglia, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.267463.

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Böhm, Andreas. "Imaging of light induced carrier transport". [S.l. : s.n.], 2002. http://www.bsz-bw.de/cgi-bin/xvms.cgi?SWB9820898.

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Dalbon, Pascal. "La protéine mitochondriale de transport des adénine-nucléotides : localisation des sites nucléotidiques par photomarquage, étude de la topographie membranaire de la chaîne polypeptidique à l'aide d'anticorps anti-peptides synthétiques". Grenoble 1, 1987. http://www.theses.fr/1987GRE10121.

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Robinson, Robert Charles. "Horse plasma vitamin D-binding protein : isolation and structural investigation". Thesis, University of British Columbia, 1990. http://hdl.handle.net/2429/30292.

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Vitamin D-binding protein (DBP) is an abundant serum protein, secreted by the liver, which transports vitamin D sterols and is part of an actin scavenging system. In this study, DBP was isolated from horse plasma in a highly reproducible, four step procedure: Affi-gel Blue affinity chromatography, gel filtration, hydroxy1apatite chromatography and anion exchange HPLC. 6-7 mg of DBP were obtained from 80 ml of plasma with a yield of 21-25%. The secondary structure of DBP was calculated from circular dichroism measurements to be 39% α-helix, 42% β-sheet and 19% random coil. A molecular mass of 53,000 ± 3,000 daltons was calculated from electrophoretic gels. Circular dichroism and fluorescence studies revealed that the disulphide bonds of DBP contribute substantial structural stabilization to the molecule with respect to thermal denaturation. Finally, acrylodan-labeled DBP was prepared. The fluorescence of this adduct was sensitive to the binding of actin and to the presence of dithiothreitol.
Science, Faculty of
Chemistry, Department of
Graduate

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Naudon, Laurent. "Recherche d'une participation du transporteur neuronal de la dopamine et du transporteur vésiculaire à l'adaptation neuronale". Rouen, 1994. http://www.theses.fr/1994ROUES066.

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L'ensemble des résultats que nous avons obtenus semblent indiquer que le transporteur neuronal de la dopamine et le transporteur vésiculaire des monoamines, malgré leurs rôles essentiels dans la transmission synaptique, ne participent que faiblement à l'adaptation des neurones dopaminergiques

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