Nematode

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Nematode Acetylcholinesterases are Encoded by Multiple Genes and Perform Non-Overlapping Functions

Journal Title, Volume, Page: 
Chemico-Biological Interactions Volumes 157-158, 15 December 2005, Pages 263-268
Year of Publication: 
2005
Authors: 
Ayman S. Hussein
Faculty of Science, An-Najah National University, PO Box 7, Nablus, Palestine
Current Affiliation: 
Faculty of Medicine & Health Sciences, Department of Biomedical Sciences, An-Najah National University, Nablus, Palestine
Murray E. Selkirk
Division of Cell and Molecular Biology, Imperial College London, London SW7 2AY, UK
Ovadia Lazari
Department of Veterinary Clinical Science, University of Liverpool, South Wirral, CH64 7TE, UK
Jacqueline B. Matthews
Division of Parasitology, Moredun Research Institute, Midlothian EH26 OPZ, UK
Preferred Abstract (Original): 

Nematodes are unusual in that diverse molecular forms of acetylcholinesterase are the product of distinct genes. This is best characterised in the free living organism Caenorhabditis elegans, in which 3 genes are known to give rise to distinct enzymes, with a fourth likely to be non-functional. ACE-1 is an amphiphilic tetramer associated with a hydrophobic non-catalytic subunit, analogous to vertebrate T enzymes, whereas ACE-2 and ACE-3 are glycosylphosphatidylinositol-linked amphiphilic dimers. The different ace genes show distinct anatomical patterns of expression in muscles, sensory neurons and motor neurons, with only a few examples of coordinated expression. Clear homologues of ace-1 and ace-2 have now been isolated from a variety of parasitic nematodes, and the predicted proteins have very similar C-terminal amino acid sequences, implying an analogous means of anchorage to membranes. In addition to these membrane-bound enzymes, many parasitic nematodes which colonise mucosal surfaces secrete acetylcholinesterases to the external (host) environment. These hydrophilic enzymes are separately encoded in the genome, so that some parasites may thus have a total complement of six ace genes. The secretory enzymes have been characterised from the intestinal nematode Nippostrongylus brasiliensis and the lungworm Dictyocaulus viviparus. These show a number of common features, including a truncated C-terminus and an insertion at the molecular surface, when compared to other nematode acetylcholinesterases. Although the function of these enzymes has not been determined, they most likely alter host physiological responses to promote survival of the parasite.

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Trichinella Spiralis Secretes A Homologue Of Prosaposin

Journal Title, Volume, Page: 
Molecular and Biochemical Parasitology Volume 135, Issue 1, May 2004, Pages 49-56
Year of Publication: 
2004
Authors: 
Ayman S. Hussein
Department of Biological Sciences, Biochemistry Building, Imperial College London, London SW7 2AZ, UK
Current Affiliation: 
Faculty of Medicine & Health Sciences, Department of Biomedical Sciences, An-Najah National University, Nablus, Palestine
Murray E. Selkirk
Department of Biological Sciences, Biochemistry Building, Imperial College London, London SW7 2AZ, UK
Anne E. Chambers
National Institute for Medical Research, Mill Hill, London NW7 1AA, UK
David Goulding
Centre for Molecular Microbiology and Infection, Imperial College London, London SW7 2AZ, UK
Marie-Pierre Gares
Centre for Molecular Microbiology and Infection, Imperial College London, London SW7 2AZ, UK
Celia Vásquez-Lopez
Department of Biological Sciences, Biochemistry Building, Imperial College London, London SW7 2AZ, UK
Teresa Gárate
Instituto de Salud Carlos III, Centro Nacional de Microbiologia, Majadahonda, Madrid, Spain
R.Michael E. Parkhouse
Instituto Gulbenkian de Ciência, PGDB, Apartado 14, 2781-901 Oeiras, Portugal
Kleoniki Gounaris
Department of Biological Sciences, Biochemistry Building, Imperial College London, London SW7 2AZ, UK
Preferred Abstract (Original): 

Infective larvae and adult stage Trichinella spiralis secrete a protein homologous to prosaposin, the precursor of sphingolipid activator proteins (saposins) A–D originally defined in vertebrates. The protein contains four saposin domains, with the six cysteine residues which form the three intramolecular disulphide bonds in close register in each case. It differs substantially from vertebrate prosaposins in the N-terminal prodomain, the region separating saposins A and B, and completely lacks the C-terminal domain which has been demonstrated to be essential for lysosomal targetting in these organisms. The protein is secreted in unprocessed form with an estimated mass of 56 kDa, and contains a single N-linked glycan which is bound by the monoclonal antibody NIM-M1, characteristic of the TSL-1 antigens which are capped by tyvelose (3,6-dideoxy-d-arabinohexose). Immuno-electron microscopy localised the protein to membrane-bound vesicles and more complex multi-lamellar organelles in diverse tissues including the hypodermis, intestine and stichosomes, although it was absent from the dense-core secretory granules typical of the latter. Possible functions of a secreted prosaposin are discussed.

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A Distinct Family of Acetylcholinesterases is Secreted by Nippostrongylus Brasiliensis

Journal Title, Volume, Page: 
Molecular and Biochemical Parasitology, Volume:123, Issue:2, 28 August 2002, Pages:125-134
Year of Publication: 
2002
Authors: 
Ayman S. Hussein
Department of Biological Sciences, Biochemistry Building, Imperial College of Science, Technology and Medicine, South Kensington Campus, London SW7 2AY, UK
Current Affiliation: 
Faculty of Medicine & Health Sciences, Department of Biomedical Sciences, An-Najah National University, Nablus, Palestine
Michal Harel
Murray E. Selkirk
Department of Biological Sciences, Biochemistry Building, Imperial College of Science, Technology and Medicine, South Kensington Campus, London SW7 2AY, UK
Preferred Abstract (Original): 

A third variant of acetylcholinesterase (AChE A) secreted by the parasitic nematode Nippostrongylus brasiliensis has been isolated which shows 63–64% identity to AChE B and AChE C, with a truncated carboxyl terminus and a short internal insertion relative to AChEs from other species. Three of the fourteen aromatic residues which line the active site gorge in Torpedo AChE are substituted by non-aromatic residues (Y70T, W279D and F288M). All three enzymes have 8 cysteine residues in conserved positions, including 6 which have been implicated in disulphide bonds in other AChEs. Phylogenetic analysis suggests that these enzymes form a distinct group which evolved after speciation and are most closely related to ACE-2 of Caenorhabditis elegans. Recombinant AChE A secreted by Pichia pastoris was monomeric and hydrophilic, with a substrate preference for acetylthiocholine and negligible activity against butyrylthiocholine. A model structure of AChE A built from the coordinates of the Torpedo californica AChE suggests that W345 (F331 in Torpedo) limits the docking of butyrylcholine. This model is consistent with mutational analysis of the nematode enzymes. Expression of AChE A is regulated at the transcriptional level independently of the other 2 secreted variants, with maximal expression by fourth stage larvae and young adult worms. These enzymes thus appear to represent an unusual family of AChEs with conserved structural features which operate outside the normal boundaries of known functions in regulation of endogenous neurotransmitter activity.

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Determinants of Substrate Specificity of a Second Non-Neuronal Secreted Acetylcholinesterase from the Parasitic Nematode Nippostrongylus Brasiliensis

Journal Title, Volume, Page: 
European Journal of Biochemistry, Volume 267, Issue 8, pages 2276–2282, April 2000
Year of Publication: 
2000
Authors: 
Ayman S. Hussein
Department of Biochemistry, Imperial College of Science, Technology and Medicine, London, UK
Current Affiliation: 
Faculty of Medicine & Health Sciences, Department of Biomedical Sciences, An-Najah National University, Nablus, Palestine
Angela M. Smith
Department of Biochemistry, Imperial College of Science, Technology and Medicine, London, UK
Matilde R. Chacón
Department of Biochemistry, Imperial College of Science, Technology and Medicine, London, UK
Murray E. Selkirk
Department of Biochemistry, Imperial College of Science, Technology and Medicine, London, UK
Preferred Abstract (Original): 

We recently reported on a non-neuronal secreted acetylcholinesterase (AChE B) from the nematode parasite Nippostrongylus brasiliensis. Here we describe the primary structure and enzymatic properties of a second secreted variant, termed AChE C after the designation of native AChE isoforms from this parasite. As for the former enzyme, AChE C is truncated at the carboxyl terminus in comparison with the Torpedo AChE, and three of the 14 aromatic residues that line the active site gorge are substituted by nonaromatic residues, corresponding to Tyr70 (Ser), Trp279 (Asn) and Phe288 (Met). A recombinant form of AChE C was highly expressed by Pichia pastoris. The enzyme was monomeric and hydrophilic, and displayed a marked preference for acetylthiocholine as substrate. A double mutation (W302F/W345F, corresponding to positions 290 and 331 in Torpedo) rendered the enzyme 10-fold less sensitive to excess substrate inhibition and two times less susceptible to the bis quaternary inhibitor BW284C51, but did not radically affect substrate specificity or sensitivity to the 'peripheral site' inhibitor propidium iodide. In contrast, a triple mutant (M300G/W302F/W345F) efficiently hydrolysed propionylthiocholine and butyrylthiocholine in addition to acetylthiocholine, while remaining insensitive to the butyrylcholinesterase-specific inhibitor iso-OMPA and displaying a similar profile of excess substrate inhibition as the double mutant. These data highlight a conserved pattern of active site architecture for nematode secreted AChEs characterized to date, and provide an explanation for the substrate specificity that might otherwise appear inconsistent with the primary structure in comparison to other invertebrate AChEs.

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