Duval N

General

Full name : Duval Nathalie

First name : Nathalie

Mail : Laboratoire de Neurobio!ogie, Ecole Normale Superieure. 46 rue d'Ulm, F-75015 Parts

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Country : France

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Phone : (33) 14329 1225

Fax : (33) 1432981 72

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References (9)

Title : Cloning and expression of acetylcholinesterase from Bungarus fasciatus venom. A new type of cooh-terminal domain\; involvement of a positively charged residue in the peripheral site - Cousin_1996_J.Biol.Chem_271_15099
Author(s) : Cousin X , Bon S , Duval N , Massoulie J , Bon C
Ref : Journal of Biological Chemistry , 271 :15099 , 1996
Abstract : As deduced from cDNA clones, the catalytic domain of Bungarus fasciatus venom acetylcholinesterase (AChE) is highly homologous to those of other AChEs. It is, however, associated with a short hydrophilic carboxyl-terminal region, containing no cysteine, that bears no resemblance to the alternative COOH-terminal peptides of the GPI-anchored molecules (H) or of other homomeric or heteromeric tailed molecules (T). Expression of complete and truncated AChE in COS cells showed that active hydrophilic monomers are produced and secreted in all cases, and that cleavage of a very basic 8-residue carboxyl-terminal fragment occurs upon secretion. The COS cells produced Bungarus AChE about 30 times more efficiently than an equivalent secreted monomeric rat AChE. The recombinant Bungarus AChE, like the natural venom enzyme, showed a distinctive ladder pattern in nondenaturing electrophoresis, probably reflecting a variation in the number of sialic acids. By mutagenesis, we showed that two differences (methionine instead of tyrosine at position 70; lysine instead of aspartate or glutamate at position 285) explain the low sensitivity of Bungarus AChE to peripheral site inhibitors, compared to the Torpedo or mammalian AChEs. These results illustrate the importance of both the aromatic and the charged residues, and the fact that peripheral site ligands (propidium, gallamine, D-tubocurarine, and fasciculin 2) interact with diverse subsets of residues.
ESTHER : Cousin_1996_J.Biol.Chem_271_15099
PubMedSearch : Cousin_1996_J.Biol.Chem_271_15099
PubMedID: 8662867
Gene_locus related to this paper: bunfa-ACHE

Title : H and T subunits of acetylcholinesterase from Torpedo, expressed in COS cells, generate all types of globular forms - Duval_1992_J.Cell.Biol_118_641
Author(s) : Duval N , Massoulie J , Bon S
Ref : Journal of Cell Biology , 118 :641 , 1992
Abstract : We analyzed the production of Torpedo marmorata acetylcholinesterase (AChE) in transfected COS cells. We report that the presence of an aspartic acid at position 397, homologous to that observed in other cholinesterases and related enzymes (Krejci, E., N. Duval, A. Chatonnet, P. Vincens, and J. Massouli. 1991. Proc. Natl. Acad. Sci. USA. 88:6647-6651), is necessary for catalytic activity. The presence of an asparagine in the previously reported cDNA sequence (Sikorav, J.L., E. Krejci, and J. Massouli. 1987. EMBO (Eur. Mol. Biol. Organ.) J. 6:1865-1873) was most likely due to a cloning error (codon AAC instead of GAC). We expressed the T and H subunits of Torpedo AChE, which differ in their COOH-terminal region and correspond respectively to the collagen-tailed asymmetric forms and to glycophosphatidylinositol-anchored dimers of Torpedo electric organs, as well as a truncated T subunit (T delta), lacking most of the COOH-terminal peptide. The transfected cells synthesized similar amounts of AChE immunoreactive protein at 37 degrees and 27 degrees C. However AChE activity was only produced at 27 degrees C and, even at this temperature, only a small proportion of the protein was active. We analyzed the molecular forms of active AChE produced at 27 degrees C. The H polypeptides generated glycophosphatidylinositol-anchored dimers, resembling the corresponding natural AChE form. The cells also released non-amphiphilic dimers G2na. The T polypeptides generated a series of active forms which are not produced in Torpedo electric organs: G1a, G2a, G4a, and G4na cellular forms and G2a and G4na secreted forms. The amphiphilic forms appeared to correspond to type II forms (Bon, S., J. P. Toutant, K. Mflah, and J. Massouli. 1988. J. Neurochem. 51:776-785; Bon, S., J. P. Toutant, K. Mflah, and J. Massouli. 1988. J. Neurochem. 51:786-794), which are abundant in the nervous tissue and muscles of higher vertebrates (Bon, S., T. L. Rosenberry, and J. Massouli. 1991. Cell. Mol. Neurobiol. 11:157-172). The H and T catalytic subunits are thus sufficient to account for all types of known AChE forms. The truncated T delta subunit yielded only non-amphiphilic monomers, demonstrating the importance of the T COOH-terminal peptide in the formation of oligomers, and in the hydrophobic character of type II forms.
ESTHER : Duval_1992_J.Cell.Biol_118_641
PubMedSearch : Duval_1992_J.Cell.Biol_118_641
PubMedID: 1639848

Title : Molecular architecture of acetylcholinesterase collagen-tailed forms\; construction of a glycolipid-tailed tetramer - Duval_1992_EMBO.J_11_3255
Author(s) : Duval N , Krejci E , Grassi J , Coussen F , Massoulie J , Bon S
Ref : EMBO Journal , 11 :3255 , 1992
Abstract : Asymmetric forms of Torpedo acetylcholinesterase (AChE) are produced in COS cells by the simultaneous expression of collagenic subunits (Q) and catalytic T subunits (AChET). Truncated AChET delta subunits, from which most of the C-terminal peptide (TC) had been deleted by mutagenesis, did not associate with Q subunits. The TC peptide is therefore necessary for the association of the AChET and Q subunits. In order to determine the orientation of the Q subunit in the collagen-tailed forms, we have developed an antiserum against its non-collagenic C-terminal domain, expressed as a fusion protein in Escherichia coli. This antiserum, which recognized the Q subunit in Western blots, was found to react with intact asymmetric forms, but not with collagenase-treated forms, from which the distal part of the tail had been cleaved, suggesting that the N-terminal non-collogenic domain (QN) is responsible for the interaction with the AChET subunits. This was confirmed by creating a chimeric subunit (QN/HC), in which QN was linked to the C-terminal peptide of the H subunit of Torpedo AChE, which contains the glycophosphatidylinositol (GPI) cleavage/attachment signal: co-expression of AChET and QN/NC produced GPI-anchored tetramers, which were sensitive to PI-PLC and largely exposed to the external surface of the cells. We thus demonstrate that: (i) the HC peptide is sufficient to determine the addition of a glycolipid anchor and (ii) the QN domain is sufficient to bind a catalytic AChET tetramer by interacting with the TC peptide.
ESTHER : Duval_1992_EMBO.J_11_3255
PubMedSearch : Duval_1992_EMBO.J_11_3255
PubMedID: 1380451

Title : Site-directed mutagenesis of active-site-related residues in Torpedo acetylcholinesterase. Presence of a glutamic acid in the catalytic triad - Duval_1992_FEBS.Lett_309_421
Author(s) : Duval N , Bon S , Silman I , Sussman JL , Massoulie J
Ref : FEBS Letters , 309 :421 , 1992
Abstract : Site-directed mutagenesis was used to investigate the role of acidic amino acid residues close to the active site of Torpedo acetylcholinesterase. The recently determined atomic structure of this enzyme shows the conserved Glu-327, together with His-440 and Ser-200 as forming a catalytic triad, while the adjacent conserved Asp-326 points away from the active site. Transfection of appropriately mutated DNA into COS cells showed that the mutation of Asp-326----Asn had little effect on catalytic activity or the molecular forms expressed, suggesting no crucial structural or functional role for this residue. Mutation of Glu-327 to Gln or to Asp led to an inactive product. These results support the conclusions of the structural analysis for the two acidic residues.
ESTHER : Duval_1992_FEBS.Lett_309_421
PubMedSearch : Duval_1992_FEBS.Lett_309_421
PubMedID: 1355448

Title : Site-Directed Mutagenesis of Functional Residues in Torpedo Acetylcholinesterase -
Author(s) : Silman I , Krejci E , Duval N
Ref : In Multidisciplinary approaches to cholinesterase functions - Proceedings of Fourth International Meeting on Cholinesterases , (Shafferman, A. and Velan, B., Eds) Plenum Press, New York :177 , 1992
PubMedID:

Title : Cholinesterase-like domains in enzymes and structural proteins: functional and evolutionary relationships and identification of a catalytically essential aspartic acid - Krejci_1991_Proc.Natl.Acad.Sci.U.S.A_88_6647
Author(s) : Krejci E , Duval N , Chatonnet A , Vincens P , Massoulie J
Ref : Proceedings of the National Academy of Sciences of the United States of America , 88 :6647 , 1991
Abstract : Primary sequences of cholinesterases and related proteins have been systematically compared. The cholinesterase-like domain of these proteins, about 500 amino acids, may fulfill a catalytic and a structural function. We identified an aspartic acid residue that is conserved among esterases and lipases (Asp-397 in Torpedo acetylcholinesterase) but that had not been considered to be involved in the catalytic mechanism. Site-directed mutagenesis demonstrated that this residue is necessary for activity. Analysis of evolutionary relationships shows that the noncatalytic members of the family do not constitute a separate subgroup, suggesting that loss of catalytic activity occurred independently on several occasions, probably from bifunctional molecules. Cholinesterases may thus be involved in cell-cell interactions in addition to the hydrolysis of acetylcholine. This would explain their specific expression in well-defined territories during embryogenesis before the formation of cholinergic synapses and their presence in noncholinergic tissues.
ESTHER : Krejci_1991_Proc.Natl.Acad.Sci.U.S.A_88_6647
PubMedSearch : Krejci_1991_Proc.Natl.Acad.Sci.U.S.A_88_6647
PubMedID: 1862088

Title : Primary structure of a collagenic tail peptide of Torpedo acetylcholinesterase: co-expression with catalytic subunit induces the production of collagen-tailed forms in transfected cells - Krejci_1991_EMBO.J_10_1285
Author(s) : Krejci E , Coussen F , Duval N , Chatel JM , Legay C , Puype M , Vandekerckhove J , Cartaud J , Bon S , Massoulie J
Ref : EMBO Journal , 10 :1285 , 1991
Abstract : The asymmetric forms of cholinesterases are synthesized only in differentiated muscular and neural cells of vertebrates. These complex oligomers are characterized by the presence of a collagen-like tail, associated with one, two or three tetramers of catalytic subunits. The collagenic tail is responsible for ionic interactions, explaining the insertion of these molecules in extracellular basal lamina, e.g. at neuromuscular endplates. We report the cloning of a collagenic subunit from Torpedo marmorata acetylcholinesterase (AChE). The predicted primary structure contains a putative signal peptide, a proline-rich domain, a collagenic domain, and a C-terminal domain composed of proline-rich and cysteine-rich regions. Several variants are generated by alternative splicing. Apart from the collagenic domain, the AChE tail subunit does not present any homology with previously known proteins. We show that co-expression of catalytic AChE subunits and collagenic subunits results in the production of asymmetric, collagen-tailed AChE forms in transfected COS cells. Thus, the assembly of these complex forms does not depend on a specific cellular processing, but rather on the expression of the collagenic subunits.
ESTHER : Krejci_1991_EMBO.J_10_1285
PubMedSearch : Krejci_1991_EMBO.J_10_1285
PubMedID: 1840520

Title : The Structure and Significance of Multiple Cholinesterase Forms -
Author(s) : Massoulie J , Bon S , Krejci E , Coussen F , Duval N , Chatel JM , Anselmet A , Legay C , Vallette FM , Grassi J
Ref : In: Cholinesterases: Structure, Function, Mechanism, Genetics, and Cell Biology , (Massoulie J, Barnard EA, Chatonnet A, Bacou F, Doctor BP, Quinn DM) American Chemical Society, Washington, DC :2 , 1991
PubMedID:

Title : Complex alternative splicing of acetylcholinesterase transcripts in Torpedo electric organ\; primary structure of the precursor of the glycolipid-anchored dimeric form - Sikorav_1988_EMBO.J_7_2983
Author(s) : Sikorav JL , Duval N , Anselmet A , Bon S , Krejci E , Legay C , Osterlund M , Reimund B , Massoulie J
Ref : EMBO Journal , 7 :2983 , 1988
Abstract : In this paper, we show the existence of alternative splicing in the 3' region of the coding sequence of Torpedo acetylcholinesterase (AChE). We describe two cDNA structures which both diverge from the previously described coding sequence of the catalytic subunit of asymmetric (A) forms (Schumacher et al., 1986; Sikorav et al., 1987). They both contain a coding sequence followed by a non-coding sequence and a poly(A) stretch. Both of these structures were shown to exist in poly(A)+ RNAs, by S1 mapping experiments. The divergent region encoded by the first sequence corresponds to the precursor of the globular dimeric form (G2a), since it contains the expected C-terminal amino acids, Ala-Cys. These amino acids are followed by a 29 amino acid extension which contains a hydrophobic segment and must be replaced by a glycolipid in the mature protein. Analyses of intact G2a AChE showed that the common domain of the protein contains intersubunit disulphide bonds. The divergent region of the second type of cDNA consists of an adjacent genomic sequence, which is removed as an intron in A and Ga mRNAs, but may encode a distinct, less abundant catalytic subunit. The structures of the cDNA clones indicate that they are derived from minor mRNAs, shorter than the three major transcripts which have been described previously (14.5, 10.5 and 5.5 kb). Oligonucleotide probes specific for the asymmetric and globular terminal regions hybridize with the three major transcripts, indicating that their size is determined by 3'-untranslated regions which are not related to the differential splicing leading to A and Ga forms.
ESTHER : Sikorav_1988_EMBO.J_7_2983
PubMedSearch : Sikorav_1988_EMBO.J_7_2983
PubMedID: 3181125