title: Expression and purification of an active cysteine protease of Haemonchus contortus using Caenorhabditis elegans.
abstract: Many proteolytic enzymes of parasitic nematodes have been identified as possible targets of control. Testing these as vaccine or drug targets is often difficult due to the problems of expressing proteases in a correctly folded, active form in standard expression systems. In an effort to overcome these difficulties we have tested Caenorhabditis elegans as an expression system for a Haemonchus contortus cathepsin L cysteine protease, Hc-CPL-1. Recombinant Hc-CPL-1 with a polyhistidine tag added to the C-terminal was expressed in an active and glycosylated form in C. elegans. Optimal expression was obtained expressing Hc-cpl-1 under control of the promoter of the homologous C. elegans cpl-1 gene. The recombinant protein was purified from liquid cultures by nickel chelation chromatography in sufficient amounts for vaccination studies to be carried out. This study provides proof of principle that active, post-translationally modified parasitic nematode proteases can be expressed in C. elegans and this approach can be extended for expression of known protective antigens.
authors: Division of Veterinary Infection and Immunity, University of Glasgow Veterinary School, Bearsden Road, Glasgow G61 1QH, UK.
authors: Linda
authors: L
authors: Murray
authors: Peter
authors: P
authors: Geldhof
authors: Douglas
authors: D
authors: Clark
authors: David P
authors: DP
authors: Knox
authors: Collette
authors: C
authors: Britton
mesh_terms: D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics*; D003546:Cysteine Endopeptidases / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics; D005260:Female; D015870:Gene Expression; D006031:Glycosylation; D006188:Haemonchiasis / Q000276:immunology / Q000662:veterinary; D006189:Haemonchus / Q000201:enzymology* / Q000235:genetics; D015801:Helminth Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008297:Male; D009154:Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics; D012756:Sheep; D012757:Sheep Diseases / Q000276:immunology / Q000469:parasitology
chemical_list: D015801:Helminth Proteins; D003546:Cysteine Endopeptidases