The Experts below are selected from a list of 222 Experts worldwide ranked by ideXlab platform
Harvey B Pollard - One of the best experts on this subject based on the ideXlab platform.
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calcium channel activity of purified human Synexin and structure of
2016Co-Authors: Lee A Burns, K Magendzo, A Shirvan, Meera Srivastava, Eduardo Rojas, R Mohammed, Harvey B PollardAbstract:Synexin is a calcium-dependent membrane binding protein that not only fuses membranes but also acts as a voltage-dependent calcium channel. We have isolated and se- quenced a set of overlapping cDNA clones for human Synexin. The derived amino acid sequence of Synexin reveals strong homology in the C-terminal domain with a previously identified class of calcium-dependent membrane binding proteins. These include endonexin 11, lipocortin I, calpactin I heavy chain (p36), protein II, and calelectrin 67K. The Mr 51,000 Synexin molecule can be divided into a unique, highly hydrophobic N-terminal domain of 167 amino acids and a conserved C-terminal region of 299 amino acids. The latter domain is composed of alternating hydrophobic and hydrophilic segments. Analysis of the entire structure reveals possible insights into such diverse properties as voltage-sensitive calcium channel activity, ion selectivity, affmity for phospholip- ids, and membrane fusion.
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Synexin (annexin VII) hypothesis for Ca2+/GTP-regulated exocytosis.
Advances in pharmacology, 2008Co-Authors: Harvey B Pollard, Hung Caohuy, A P Minton, Meera SrivastavaAbstract:Publisher Summary The current fusion machine hypothesis envisions a core complex formed between plasma membrane syntaxin and SNAP-25 and the synaptic vesicle protein synaptobrevinNAMP (S), with vesicular synaptotagmin identified as a low-affinity calcium sensor that interacts with regulatory syntaxin 1. The site of GTP action in exocytosis has been thought to be closely associated with the site of calcium action at some common site. The affinity of this site for Ca 2+ has been shown to be in the 50- to 200- μ M range, as estimated from electrophysiological studies with caged calcium in chromaffin cells, neurons, and digitonin-permeabilized chromaffin cells. Therefore, a good experimental goal would seem to be to search for a embrane fusion protein that is activated by both calcium and GTP. Chromaffin and many other cell types contain a membrane fusion protein, Synexin (annexin VII), associated with secretory vesicles and plasma membranes, which have an appropriate intrinsic K d for Ca 2+ of approximately 200 μ M. In addition to binding of GTP and Ca 2+ , a further identifying feature of the hypothetical “fusion scaffold” protein has been proposed to be self-association. Indeed, it has been shown by light scattering and electron microscopy studies that the Ca 2+ -activated form of Synexin has a polymeric structure. Consistently, the protein concentration dependence for many Synexindependent reactions has consistently been sigmoidal, with Hill coefficients (n H ). It is true that Synexin alone is able to mediate efficiently both membrane contact and fusion reactions in vitro . However, the exocytosis process is likely to be much more complicated.
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Chapter 20 Role of Ions and Intracellular Proteins in Exocytosis
Methods in Cell Biology, 2008Co-Authors: Harvey B Pollard, Carl E Creutz, Christopher J PazolesAbstract:Publisher Summary This chapter describes a new protein called Synexin that induces Ca 2+ -dependent formation of pentalaminar complexes among secretory granule membranes. Synexin has proved distinct from other more conventional candidates for Ca 2+ -effect mediators, such as calmodulin, actomyosin, and tubulin. The chapter also describes the process of membrane breakage or fission that finally results in the release of secretory vesicle contents. Studies comparing the granule lysis reaction with predicted behavior of secreting chromaffin and other cells are briefly reviewed in the chapter. The biochemical data supporting the exocytosis hypothesis are that the storage organelles from different cells can be isolated and therefore shown to be discrete objects and when secretion occurs, the entire organelle contents ranging from small molecules to large proteins can be found in the extracellular medium. Calcium plays a critical role in regulating exocytosis in a number of systems.
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Annexin hypothesis for exocytosis
Biomembranes: A Multi-Volume Treatise, 2007Co-Authors: A. Lee Burns, Harvey B PollardAbstract:Publisher Summary Membrane fusion processes in cells underlie the structural integrity of the cytoplasm and are crucial to directing synthesis and export of enzymes, hormones, and transmitters. Export is accomplished by a process of exocytosis in which the membranes of secretory vesicles fuse with the plasma membrane. The fusion mechanism may involve a class of calcium-binding proteins termed “annexins.” This chapter discusses the annexin hypothesis for exocytosis. It focuses on function and molecular biology Synexin (annexin VII) in secretory cells. Results from several laboratories have also shown that annexins I, II, and VII aggregate chromaffin granules and liposomes in vitro , and that annexin VII drives fusion of granule ghosts and liposomes. These data lend support to the hypothesis that these annexins may be secretory factors in vivo . The above mutagenesis experiments revealed the importance in annexin I of repeat one for aggregation activity and of repeats 2, 3, and 4 for calcium-binding sites. As all annexins bind to phospholipids in a calcium-dependent manner, membranes continue to be a focus for understanding annexin activity. Annexin II was detected in postpartum mammary epithelial cells, when calcium-dependent casein secretion starts and immunogold labeling of annexin VII in chromaffin cells was shown to decrease following treatment with a secretagogue. In addition, annexin II has been reported to insert into artificial membranes in the presence of calcium and to produce voltage-dependent channels.
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Development of a cell-free model for compound exocytosis using components of the chromaffin cell
Journal of The Autonomic Nervous System, 2003Co-Authors: Carl E Creutz, Harvey B PollardAbstract:Abstract A study of exocytosis and the possible role of chromaffin granules in adrenal medulla secretion is presented. This review describes a system that may recreate Ca 2+ -dependent membrane contact and fusion events that occur in exocytosis. The nature and role of Synexin and other compounds in this process are discussed.
Meera Srivastava - One of the best experts on this subject based on the ideXlab platform.
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calcium channel activity of purified human Synexin and structure of
2016Co-Authors: Lee A Burns, K Magendzo, A Shirvan, Meera Srivastava, Eduardo Rojas, R Mohammed, Harvey B PollardAbstract:Synexin is a calcium-dependent membrane binding protein that not only fuses membranes but also acts as a voltage-dependent calcium channel. We have isolated and se- quenced a set of overlapping cDNA clones for human Synexin. The derived amino acid sequence of Synexin reveals strong homology in the C-terminal domain with a previously identified class of calcium-dependent membrane binding proteins. These include endonexin 11, lipocortin I, calpactin I heavy chain (p36), protein II, and calelectrin 67K. The Mr 51,000 Synexin molecule can be divided into a unique, highly hydrophobic N-terminal domain of 167 amino acids and a conserved C-terminal region of 299 amino acids. The latter domain is composed of alternating hydrophobic and hydrophilic segments. Analysis of the entire structure reveals possible insights into such diverse properties as voltage-sensitive calcium channel activity, ion selectivity, affmity for phospholip- ids, and membrane fusion.
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Synexin (annexin VII) hypothesis for Ca2+/GTP-regulated exocytosis.
Advances in pharmacology, 2008Co-Authors: Harvey B Pollard, Hung Caohuy, A P Minton, Meera SrivastavaAbstract:Publisher Summary The current fusion machine hypothesis envisions a core complex formed between plasma membrane syntaxin and SNAP-25 and the synaptic vesicle protein synaptobrevinNAMP (S), with vesicular synaptotagmin identified as a low-affinity calcium sensor that interacts with regulatory syntaxin 1. The site of GTP action in exocytosis has been thought to be closely associated with the site of calcium action at some common site. The affinity of this site for Ca 2+ has been shown to be in the 50- to 200- μ M range, as estimated from electrophysiological studies with caged calcium in chromaffin cells, neurons, and digitonin-permeabilized chromaffin cells. Therefore, a good experimental goal would seem to be to search for a embrane fusion protein that is activated by both calcium and GTP. Chromaffin and many other cell types contain a membrane fusion protein, Synexin (annexin VII), associated with secretory vesicles and plasma membranes, which have an appropriate intrinsic K d for Ca 2+ of approximately 200 μ M. In addition to binding of GTP and Ca 2+ , a further identifying feature of the hypothetical “fusion scaffold” protein has been proposed to be self-association. Indeed, it has been shown by light scattering and electron microscopy studies that the Ca 2+ -activated form of Synexin has a polymeric structure. Consistently, the protein concentration dependence for many Synexindependent reactions has consistently been sigmoidal, with Hill coefficients (n H ). It is true that Synexin alone is able to mediate efficiently both membrane contact and fusion reactions in vitro . However, the exocytosis process is likely to be much more complicated.
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HIV-1 GAG SHARES A SIGNATURE MOTIF WITH ANNEXIN (ANX7), WHICH IS REQUIRED FOR VIRUS REPLICATION
Proceedings of the National Academy of Sciences of the United States of America, 1999Co-Authors: Meera Srivastava, H B Pollard, Maria Cartas, Tahir A. Rizvi, Satya P. Singh, D. Serio, Vaniambadi S. Kalyanaraman, Alagarsamy SrinivasanAbstract:Genetic and biochemical analyses of the Gag protein of HIV-1 indicate a crucial role for this protein in several functions related to viral replication, including viral assembly. It has been suggested that Gag may fulfill some of the functions by recruiting host cellular protein(s). In our effort to identify structural and functional homologies between Gag and cellular cytoskeletal and secretory proteins involved in transport, we observed that HIV-1 Gag contains a unique PGQM motif in the capsid region. This motif was initially noted in the regulatory domain of Synexin the membrane fusion protein of Xenopus laevis. To evaluate the functional significance of the highly conserved PGQM motif, we introduced alanine (A) in place of individual residues of the PGQM and deleted the motif altogether in a Gag expression plasmid and in an HIV-1 proviral DNA. The proviral DNA containing mutations in the PGQM motif showed altered expression, assembly, and release of viral particles in comparison to parental (NL4-3) DNA. When tested in multiple- and single-round replication assays, the mutant viruses exhibited distinct replication phenotypes; the viruses containing the A for the G and Q residues failed to replicate, whereas A in place of the P and M residues did not inhibit viral replication. Deletion of the tetrapeptide also resulted in the inhibition of replication. These results suggest that the PGQM motif may play an important role in the infection process of HIV-1 by facilitating protein–protein interactions between viral and/or viral and cellular proteins.
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Synexin annexin vii hypothesis for ca2 gtp regulated exocytosis
Advances in pharmacology, 1997Co-Authors: Harvey B Pollard, Meera Srivastava, Hung Caohuy, A P MintonAbstract:Publisher Summary The current fusion machine hypothesis envisions a core complex formed between plasma membrane syntaxin and SNAP-25 and the synaptic vesicle protein synaptobrevinNAMP (S), with vesicular synaptotagmin identified as a low-affinity calcium sensor that interacts with regulatory syntaxin 1. The site of GTP action in exocytosis has been thought to be closely associated with the site of calcium action at some common site. The affinity of this site for Ca 2+ has been shown to be in the 50- to 200- μ M range, as estimated from electrophysiological studies with caged calcium in chromaffin cells, neurons, and digitonin-permeabilized chromaffin cells. Therefore, a good experimental goal would seem to be to search for a embrane fusion protein that is activated by both calcium and GTP. Chromaffin and many other cell types contain a membrane fusion protein, Synexin (annexin VII), associated with secretory vesicles and plasma membranes, which have an appropriate intrinsic K d for Ca 2+ of approximately 200 μ M. In addition to binding of GTP and Ca 2+ , a further identifying feature of the hypothetical “fusion scaffold” protein has been proposed to be self-association. Indeed, it has been shown by light scattering and electron microscopy studies that the Ca 2+ -activated form of Synexin has a polymeric structure. Consistently, the protein concentration dependence for many Synexindependent reactions has consistently been sigmoidal, with Hill coefficients (n H ). It is true that Synexin alone is able to mediate efficiently both membrane contact and fusion reactions in vitro . However, the exocytosis process is likely to be much more complicated.
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detection and localization of Synexin annexin vii in xenopus adult and embryonic tissues using an antibody to the xenopus n terminal pgqm repeat domain
Experimental Cell Research, 1996Co-Authors: Meera Srivastava, Hung Caohuy, Peter Mcphie, Gertrude Goping, Harvey B PollardAbstract:Abstract Synexin (Annexin VII) is a widely distributed member of the annexin gene family which forms calcium channels and drives calcium-dependent membrane fusion. In Xenopus laevis, different Synexins contain two to six tandem repeats of the tetra amino acid sequence PGQM in the unique N-terminal, with a distribution specific to adult tissues and embryonic stages. Immunogold studies using the PGQM-specific polyclonal antibody showed that Synexin is localized in adult muscle to myosin-rich A-bands, Z-bands, and T-tubules, and in other adult tissues to nuclei and mitochondria and other formed elements. In oocytes, Synexin was also found associated with yolk granules. The PGQM tandem repeats could represent interaction sites for other proteins, and we therefore synthesized a synthetic peptide containing the maximum six tandem repeats [NH 2 -(PGQM) 6 -Y-COOH] to test this hypothesis. We found that the peptide alone could specifically bind and crosslink to different proteins in a tissue-specific manner. In liver, it bound to a single 35-kDa protein. In muscle, it bound to four proteins (35, 45, 48, and 116 kDa). Therefore, we conclude that the PGQM domain is accessible to specific antibodies and that the PGQM repeat is sufficiently ordered to unambiguously identify specific binding proteins in different Xenopus tissues.
Carl E Creutz - One of the best experts on this subject based on the ideXlab platform.
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Novel protein ligands of the annexin A7 N-terminal region suggest pro-beta helices engage one another with high specificity.
General physiology and biophysics, 2020Co-Authors: Carl E CreutzAbstract:The N-terminal regions of annexins A7 (Synexin) and A11 consist of an extended series of short sequence repeats rich in tyrosine, proline, and glycine that provide binding sites for other proteins that may be recruited to membranes by the annexins and that may modulate the calcium and membrane binding activities of the annexin core domains. In this study two new ligands for the annexin A7 N terminal region were identified by yeast two hybrid screening: the TNFalpha receptor regulatory protein SODD (Suppressor Of Death Domains) and KIAA0280, a protein of unknown function. Strikingly, the sites of interaction of these proteins with the annexin also contain sequence repeats similar to those present in the N-termini of annexins A7 and A11. It was also found that the annexin A7 N-terminal region interacts with itself in the two hybrid assay. These results suggest that sequence repeats of this nature form novel structures, called YP pro-beta helices, that are characterized by an ability to interact with one another. Specificity of interactions between the pro-beta helices in different proteins may be encoded by the variations of residues and lengths of the sequence repeats.
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Novel protein ligands of the annexin A7 N-terminal region suggest pro-β helices engage one another with high specificity
General Physiology and Biophysics, 2020Co-Authors: Carl E CreutzAbstract:The N-terminal regions of annexins A7 (Synexin) and A11 consist of an extended series of short sequence repeats rich in tyrosine, proline, and glycine that provide binding sites for other proteins that may be recruited to membranes by the annexins and that may modulate the calcium and membrane binding activities of the annexin core domains. In this study two new ligands for the annexin A7 N terminal region were identified by yeast two hybrid screening: the TNFα receptor regulatory protein SODD (Suppressor Of Death Domains) and KIAA0280, a protein of unknown function. Strikingly, the sites of interaction of these proteins with the annexin also contain sequence repeats similar to those present in the N-termini of annexins A7 and A11. It was also found that the annexin A7 N-terminal region interacts with itself in the two hybrid assay. These results suggest that sequence repeats of this nature form novel structures, called YP pro-β helices, that are characterized by an ability to interact with one another. Specificity of interactions between the pro-β helices in different proteins may be encoded by the variations of residues and lengths of the sequence repeats.
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Chapter 20 Role of Ions and Intracellular Proteins in Exocytosis
Methods in Cell Biology, 2008Co-Authors: Harvey B Pollard, Carl E Creutz, Christopher J PazolesAbstract:Publisher Summary This chapter describes a new protein called Synexin that induces Ca 2+ -dependent formation of pentalaminar complexes among secretory granule membranes. Synexin has proved distinct from other more conventional candidates for Ca 2+ -effect mediators, such as calmodulin, actomyosin, and tubulin. The chapter also describes the process of membrane breakage or fission that finally results in the release of secretory vesicle contents. Studies comparing the granule lysis reaction with predicted behavior of secreting chromaffin and other cells are briefly reviewed in the chapter. The biochemical data supporting the exocytosis hypothesis are that the storage organelles from different cells can be isolated and therefore shown to be discrete objects and when secretion occurs, the entire organelle contents ranging from small molecules to large proteins can be found in the extracellular medium. Calcium plays a critical role in regulating exocytosis in a number of systems.
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Polyproline, β‐turn helices. Novel secondary structures proposed for the tandem repeats within rhodopsin, synaptophysin, Synexin, gliadin, RNA polymerase II, hordein, and gluten
Proteins, 2004Co-Authors: Norio Matsushima, Carl E Creutz, Robert H KretsingerAbstract:Seven proteins each contain 8 to 52 tandem repeats of a unique class of oligopeptide. The consensus peptide for each is rhodopsin Tyr Pro Pro Gln Gly synapto-physin Tyr Gly Pro Gln Gly Synexin Tyr Pro Pro Pro Pro Gly gliadin Tyr Pro Pro Pro Gln Pro RNA polymerase II Tyr Ser Pro Thr Ser Pro Ser hordein Phe Pro Gln Gln Pro Gln Gln Pro gluten Tyr Pro Thr Ser Pro Gln Gn Gly Tyr Although there is obvious variations of sequence and of length, the penta-to nonapeptides share an initial Tyr(or Phe) and have high Pro contents and abundant Gly, Gln, and Ser. We have evaluated helical models that both recognize the uniqueness of these sequence repeats and accommodate variations on the basic theme. We have developed a group of related heical model for these proteins with about three oligopeptide repeats per turn of 10–20 A. These models share several common features: Most of the ϕ dihedral angels are −54°, to accommodate Pro at all positions expect the first (Tyr). Except for the β-turns, most ψ dihedral angles are near +140° as found in polyproline. Each oligopeptide has at least one β-turn; several have two. Some contains a cis-Tyr, Pro peptide bond; a few have a cis-bond plus one β-turn. Tyr side chains vary from totally exposed to buried within the helices and could mode to accommodate either external hydrophobic interactions or phosphorylation. The several related structures seem to be readily interconverted without major change in the overall helical parameters, and therein may lie the key to their functions.
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Development of a cell-free model for compound exocytosis using components of the chromaffin cell
Journal of The Autonomic Nervous System, 2003Co-Authors: Carl E Creutz, Harvey B PollardAbstract:Abstract A study of exocytosis and the possible role of chromaffin granules in adrenal medulla secretion is presented. This review describes a system that may recreate Ca 2+ -dependent membrane contact and fusion events that occur in exocytosis. The nature and role of Synexin and other compounds in this process are discussed.
Hung Caohuy - One of the best experts on this subject based on the ideXlab platform.
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Synexin (annexin VII) hypothesis for Ca2+/GTP-regulated exocytosis.
Advances in pharmacology, 2008Co-Authors: Harvey B Pollard, Hung Caohuy, A P Minton, Meera SrivastavaAbstract:Publisher Summary The current fusion machine hypothesis envisions a core complex formed between plasma membrane syntaxin and SNAP-25 and the synaptic vesicle protein synaptobrevinNAMP (S), with vesicular synaptotagmin identified as a low-affinity calcium sensor that interacts with regulatory syntaxin 1. The site of GTP action in exocytosis has been thought to be closely associated with the site of calcium action at some common site. The affinity of this site for Ca 2+ has been shown to be in the 50- to 200- μ M range, as estimated from electrophysiological studies with caged calcium in chromaffin cells, neurons, and digitonin-permeabilized chromaffin cells. Therefore, a good experimental goal would seem to be to search for a embrane fusion protein that is activated by both calcium and GTP. Chromaffin and many other cell types contain a membrane fusion protein, Synexin (annexin VII), associated with secretory vesicles and plasma membranes, which have an appropriate intrinsic K d for Ca 2+ of approximately 200 μ M. In addition to binding of GTP and Ca 2+ , a further identifying feature of the hypothetical “fusion scaffold” protein has been proposed to be self-association. Indeed, it has been shown by light scattering and electron microscopy studies that the Ca 2+ -activated form of Synexin has a polymeric structure. Consistently, the protein concentration dependence for many Synexindependent reactions has consistently been sigmoidal, with Hill coefficients (n H ). It is true that Synexin alone is able to mediate efficiently both membrane contact and fusion reactions in vitro . However, the exocytosis process is likely to be much more complicated.
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Synexin annexin vii hypothesis for ca2 gtp regulated exocytosis
Advances in pharmacology, 1997Co-Authors: Harvey B Pollard, Meera Srivastava, Hung Caohuy, A P MintonAbstract:Publisher Summary The current fusion machine hypothesis envisions a core complex formed between plasma membrane syntaxin and SNAP-25 and the synaptic vesicle protein synaptobrevinNAMP (S), with vesicular synaptotagmin identified as a low-affinity calcium sensor that interacts with regulatory syntaxin 1. The site of GTP action in exocytosis has been thought to be closely associated with the site of calcium action at some common site. The affinity of this site for Ca 2+ has been shown to be in the 50- to 200- μ M range, as estimated from electrophysiological studies with caged calcium in chromaffin cells, neurons, and digitonin-permeabilized chromaffin cells. Therefore, a good experimental goal would seem to be to search for a embrane fusion protein that is activated by both calcium and GTP. Chromaffin and many other cell types contain a membrane fusion protein, Synexin (annexin VII), associated with secretory vesicles and plasma membranes, which have an appropriate intrinsic K d for Ca 2+ of approximately 200 μ M. In addition to binding of GTP and Ca 2+ , a further identifying feature of the hypothetical “fusion scaffold” protein has been proposed to be self-association. Indeed, it has been shown by light scattering and electron microscopy studies that the Ca 2+ -activated form of Synexin has a polymeric structure. Consistently, the protein concentration dependence for many Synexindependent reactions has consistently been sigmoidal, with Hill coefficients (n H ). It is true that Synexin alone is able to mediate efficiently both membrane contact and fusion reactions in vitro . However, the exocytosis process is likely to be much more complicated.
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detection and localization of Synexin annexin vii in xenopus adult and embryonic tissues using an antibody to the xenopus n terminal pgqm repeat domain
Experimental Cell Research, 1996Co-Authors: Meera Srivastava, Hung Caohuy, Peter Mcphie, Gertrude Goping, Harvey B PollardAbstract:Abstract Synexin (Annexin VII) is a widely distributed member of the annexin gene family which forms calcium channels and drives calcium-dependent membrane fusion. In Xenopus laevis, different Synexins contain two to six tandem repeats of the tetra amino acid sequence PGQM in the unique N-terminal, with a distribution specific to adult tissues and embryonic stages. Immunogold studies using the PGQM-specific polyclonal antibody showed that Synexin is localized in adult muscle to myosin-rich A-bands, Z-bands, and T-tubules, and in other adult tissues to nuclei and mitochondria and other formed elements. In oocytes, Synexin was also found associated with yolk granules. The PGQM tandem repeats could represent interaction sites for other proteins, and we therefore synthesized a synthetic peptide containing the maximum six tandem repeats [NH 2 -(PGQM) 6 -Y-COOH] to test this hypothesis. We found that the peptide alone could specifically bind and crosslink to different proteins in a tissue-specific manner. In liver, it bound to a single 35-kDa protein. In muscle, it bound to four proteins (35, 45, 48, and 116 kDa). Therefore, we conclude that the PGQM domain is accessible to specific antibodies and that the PGQM repeat is sufficiently ordered to unambiguously identify specific binding proteins in different Xenopus tissues.
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membrane fusion protein Synexin annexin vii as a ca2 gtp sensor in exocytotic secretion
Proceedings of the National Academy of Sciences of the United States of America, 1996Co-Authors: Hung Caohuy, Meera Srivastava, Harvey B PollardAbstract:Abstract Exocytotic membrane fusion and secretion are promoted by the concerted action of GTP and Ca2+, although the precise site(s) of action in the process are not presently known. However, the calcium-dependent membrane fusion reaction driven by Synexin (annexin VII) is an in vitro model for this process, which we have now found to be further activated by GTP. The mechanism of fusion activation depends on the unique ability of Synexin to bind and hydrolyze GTP in a calcium-dependent manner, both in vitro and in vivo in streptolysin O-permeabilized chromaffin cells. The required [Ca2+] for GTP binding by Synexin is in the range of 50-200 microM, which is known to occur at exocytotic sites in chromaffin cells, neurons, and other cell types. Previous immunolocalization studies place Synexin at exocytotic sites in chromaffin cells, and we conclude that Synexin is an atypical G protein that may be responsible for both detecting and mediating the Ca2+/GTP signal for exocytotic membrane fusion.
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Membrane fusion protein Synexin (annexin VII) as a Ca2+/GTP sensor in exocytotic secretion.
Proceedings of the National Academy of Sciences of the United States of America, 1996Co-Authors: Hung Caohuy, Meera Srivastava, Harvey B PollardAbstract:Abstract Exocytotic membrane fusion and secretion are promoted by the concerted action of GTP and Ca2+, although the precise site(s) of action in the process are not presently known. However, the calcium-dependent membrane fusion reaction driven by Synexin (annexin VII) is an in vitro model for this process, which we have now found to be further activated by GTP. The mechanism of fusion activation depends on the unique ability of Synexin to bind and hydrolyze GTP in a calcium-dependent manner, both in vitro and in vivo in streptolysin O-permeabilized chromaffin cells. The required [Ca2+] for GTP binding by Synexin is in the range of 50-200 microM, which is known to occur at exocytotic sites in chromaffin cells, neurons, and other cell types. Previous immunolocalization studies place Synexin at exocytotic sites in chromaffin cells, and we conclude that Synexin is an atypical G protein that may be responsible for both detecting and mediating the Ca2+/GTP signal for exocytotic membrane fusion.
Angelika A. Noegel - One of the best experts on this subject based on the ideXlab platform.
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The annexins of Dictyostelium.
European journal of cell biology, 2006Co-Authors: Marija Marko, Michael Schleicher, Yogikala Prabhu, Rolf Müller, Rosemarie Blau-wasser, Angelika A. NoegelAbstract:Annexins are a highly conserved ubiquitous family of Ca2+- and phospholipid-binding proteins present in nearly all eukaryotic cells. Analysis of the Dictyostelium genome revealed the presence of two annexin genes, the annexin C1 gene (nxnA) giving rise to two isoforms of 47 and 51 kDa (previously Synexin), and the annexin C2 gene (nxnB) coding for a 56-kDa protein with 33% sequence identity to annexin C1. Annexin C2 is expressed at very low and constant levels throughout development. Quantification by real-time PCR indicated that it is present in about 35-fold lower amounts compared to annexin C1. We have used a GFP-tagged annexin C2 to study its cellular distribution and dynamics. In cell fractionation studies, annexin C2 cofractionates with annexin C1 and is enriched in the 100,000 g pellet. Like annexin C1, GFP-AnxC2 stains the plasma membrane. In addition it is present in the perinuclear region and overlaps to some degree with the Golgi apparatus, whereas annexin C1 is present on intracellular membranes resembling endosomal membranes and in the nucleus. Annexin C2 is not observed in the nucleus. An annexin C1 mutant (SYN-) which shows a defect during multicellular development can be rescued by full-length annexin C1, whereas overexpression of GFP-AnxC2 did not rescue the developmental defect The data support the concept that annexins, although having a highly conserved structure, participate in different functions in a cell.
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Annexin VII: an astroglial protein exhibiting a Ca2+-dependent subcellular distribution.
Neuroreport, 2001Co-Authors: Christoph S. Clemen, Angelika A. Noegel, Claudia Herr, Rolf SchröderAbstract:A fundamental issue in neuronal and glial cells is how intracellular rises in Ca 2+ are coupled to signaling cascades and changes in subcellular morphology. We studied the expression and localization of annexin VII (Synexin), a Ca 2+ -/GTP-dependent membrane fusion protein, in the human CNS. Here, we demonstrate the presence of two annexin VII isoforms (47 and 51 kDa) in human brain tissue as well as its exclusive expression in astroglial cells. An in vitro study of astrocyte-derived C6 rat glioblastoma cells expressing a GFP tagged annexin VII fusion protein demonstrates a sequential redistribution of the fusion protein in response to rising intracellular Ca 2+ concentrations. Our findings indicate a role of annexin VII in the regulation of intracellular Ca 2+ -dependent processes in astroglial cells.
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The sorcin–annexin VII calcium-dependent interaction requires the sorcin N-terminal domain
FEBS Letters, 2000Co-Authors: Daniela Verzili, Carlotta Zamparelli, Benedetta Mattei, Angelika A. Noegel, Emilia ChianconeAbstract:Abstract Surface plasmon resonance experiments show that at neutral pH the stability of the complex between sorcin and annexin VII (Synexin) increases dramatically between 3 and 6 μM calcium; at the latter cation concentration the KD value is 0.63 μM. In turn, the lack of complex formation between the sorcin Ca2+ binding domain (33–198) and Synexin maps the annexin binding site to the N-terminal region of the sorcin polypeptide chain. Annexin VII likewise employs the N-terminal domain, more specifically the first 31 amino acids, to interact with sorcin [Brownawell, A.M. and Creutz, C.E. (1997) J. Biol. Chem. 272, 22182–22190]. The interaction may involve similar structural motifs in the two proteins, namely GGYY and GYGG in sorcin and GYPP in Synexin.
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The sorcin-annexin VII calcium-dependent interaction requires the sorcin N-terminal domain.
FEBS letters, 2000Co-Authors: Daniela Verzili, Carlotta Zamparelli, Benedetta Mattei, Angelika A. Noegel, Emilia ChianconeAbstract:Surface plasmon resonance experiments show that at neutral pH the stability of the complex between sorcin and annexin VII (Synexin) increases dramatically between 3 and 6 microM calcium; at the latter cation concentration the K(D) value is 0.63 microM. In turn, the lack of complex formation between the sorcin Ca(2+) binding domain (33-198) and Synexin maps the annexin binding site to the N-terminal region of the sorcin polypeptide chain. Annexin VII likewise employs the N-terminal domain, more specifically the first 31 amino acids, to interact with sorcin [Brownawell, A.M. and Creutz, C.E. (1997) J. Biol. Chem. 272, 22182-22190]. The interaction may involve similar structural motifs in the two proteins, namely GGYY and GYGG in sorcin and GYPP in Synexin.
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Severe developmental defects in Dictyostelium null mutants for actin-binding proteins.
Mechanisms of Development, 2000Co-Authors: Eleonora Ponte, Angelika A. Noegel, Francisco Rivero, Marcus Fechheimer, Salvatore BozzaroAbstract:The actin cytoskeleton is implicated in many cellular processes, such as cell adhesion, locomotion, contraction and cytokinesis, which are central to any development. The extent of polymerization, cross-linking, and bundling of actin is regulated by several actin-binding proteins. Knock-out mutations in these proteins have revealed in many cases only subtle, if any, defects in development, suggesting that the actin system is redundant, with multiple proteins sharing overlapping functions. The apparent redundancy may, however, reflect limitations of available laboratory assays in assessing the developmental role of a given protein. By using a novel assay, which reproduces conditions closer to the natural ones, we have re-examined the effects of disruption of many actin-binding proteins, and show here that deletion of α-actinin, interaptin, Synexin, 34-kDa actin-bundling protein, and gelation factor affect to varying degrees the efficiency of Dictyostelium cells to complete development and form viable spores. No phenotypic defects were found in hisactophilin or comitin null mutants.
