title: Prokaryotic expression of ovis aries conglutinin encoding neck and carbohydrate recognition domain and its functional characterization.
abstract: Conglutinin, a soluble pattern recognition receptor of innate immune system in bovines is known for its potential defensive activity against microorganisms either by direct agglutination in the presence of calcium or by acting as opsonin. In the present study, sheep (Ovis aries) conglutinin encoding neck and carbohydrate recognition domain (rSCGN) was expressed in the E coli BL21 expression host. The recombinant conglutinin revealed molecular weight of 27 kDa in SDS PAGE and also in western blotting using antibuffalo conglutinin polyclonal serum. The protein was characterized further for its functional activity in various assays. In ELISA based sugar and LPS binding assay, the rSCGN revealed its high binding activity toward N-acetyl glucosamine and E. coli LPS in the presence and the absence of calcium ions, respectively. Hemagglutination of chicken red blood cells caused by Newcastle disease virus was not inhibited in the presence of rSCGN as it lacked complete collagenous region present in the native protein. In virus neutralization test, the recombinant protein was found to reduce multiplication of bovine herpes virus-1 propagated in MDBK cells. This prokaryotically expressed 27 kDa recombinant sheep conglutinin can serve as antigen in future studies to develop sandwich ELISA for assessing the level of native conglutinin in sheep serum.
authors.affiliation: a Center for Wildlife Conservation, Management & Disease Surveillance , Indian Veterinary Research Institute , Izatnagar , India.
authors.forename: S Chandra
authors.initials: SC
authors.lastname: Mohan
authors.forename: Mohini
authors.initials: M
authors.lastname: Saini
authors.forename: D
authors.initials: D
authors.lastname: Ramesh
authors.forename: M
authors.initials: M
authors.lastname: Shynu
authors.forename: Sasmita
authors.initials: S
authors.lastname: Barik
authors.forename: Asit
authors.initials: A
authors.lastname: Das
authors.forename: Anil Kumar
authors.initials: AK
authors.lastname: Sharma
authors.forename: Vinod Kumar
authors.initials: VK
authors.lastname: Chaturvedi
authors.forename: Praveen Kumar
authors.initials: PK
authors.lastname: Gupta
mesh_terms: D000117:Acetylglucosamine / Q000378:metabolism; D000818:Animals; D002645:Chickens; D037222:Collectins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D004912:Erythrocytes / Q000821:virology; D004926:Escherichia coli; D008070:Lipopolysaccharides / Q000378:metabolism; D009522:Newcastle disease virus; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D012712:Serum Globulins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D034561:Sheep, Domestic
chemical_list: D037222:Collectins; D008070:Lipopolysaccharides; D011994:Recombinant Proteins; D012712:Serum Globulins; C024014:conglutinin; D000117:Acetylglucosamine
keywords: Innate immunity; Ovis aries; Prokaryotic expression; Recombinant conglutinin; Sugar binding