title: Expression, characterisation and antigenicity of a truncated Hendra virus attachment protein expressed in the protozoan host Leishmania tarentolae.
abstract: Hendra virus (HeV) is an emerging zoonotic paramyxovirus within the genus Henipavirus that has caused severe morbidity and mortality in humans and horses in Australia since 1994. HeV infection of host cells is mediated by the membrane bound attachment (G) and fusion (F) glycoproteins, that are essential for receptor binding and fusion of viral and cellular membranes. The eukaryotic unicellular parasite Leishmania tarentolae has recently been established as a powerful tool to express recombinant proteins with mammalian-like glycosylation patterns, but only few viral proteins have been expressed in this system so far. Here, we describe the purification of a truncated, Strep-tag labelled and soluble version of the HeV attachment protein (sHeV G) expressed in stably transfected L. tarentolae cells. After Strep-tag purification the identity of sHeV G was confirmed by immunoblotting and mass spectrometry. The functional binding of sHeV G to the HeV cell entry receptor ephrin-B2 was confirmed in several binding assays. Generated polyclonal rabbit antiserum against sHeV G reacted with both HeV and Nipah virus (NiV) G proteins in immunofluorescence assay and efficiently neutralised NiV infection, thus further supporting the preserved antigenicity of the purified protein.
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Novel and Emerging Infectious Diseases, Greifswald-Insel Riems, Germany.
authors.forename: Kerstin
authors.initials: K
authors.lastname: Fischer
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Novel and Emerging Infectious Diseases, Greifswald-Insel Riems, Germany.
authors.forename: Vinicius Pinho
authors.initials: VP
authors.lastname: dos Reis
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Molecular Virology and Cell Biology, Greifswald-Insel Riems, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Finke
authors.affiliation: Institute of Virology, Philipps University of Marburg, Marburg, Germany.
authors.forename: Lucie
authors.initials: L
authors.lastname: Sauerhering
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Novel and Emerging Infectious Diseases, Greifswald-Insel Riems, Germany.
authors.forename: Eileen
authors.initials: E
authors.lastname: Stroh
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Molecular Virology and Cell Biology, Greifswald-Insel Riems, Germany.
authors.forename: Axel
authors.initials: A
authors.lastname: Karger
authors.affiliation: Institute of Virology, Philipps University of Marburg, Marburg, Germany.
authors.forename: Andrea
authors.initials: A
authors.lastname: Maisner
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Novel and Emerging Infectious Diseases, Greifswald-Insel Riems, Germany.
authors.forename: Martin H
authors.initials: MH
authors.lastname: Groschup
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Novel and Emerging Infectious Diseases, Greifswald-Insel Riems, Germany.
authors.forename: Sandra
authors.initials: S
authors.lastname: Diederich
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Novel and Emerging Infectious Diseases, Greifswald-Insel Riems, Germany. Electronic address: anne.buschmann@fli.bund.de.
authors.forename: Anne
authors.initials: A
authors.lastname: Balkema-Buschmann
mesh_terms: D000818:Animals; D001315:Australia; D036388:Ephrin-B2 / Q000378:metabolism; D045404:Hendra Virus / Q000737:chemistry* / Q000235:genetics / Q000276:immunology / Q000502:physiology; D006736:Horses; D006801:Humans; D007891:Leishmania / Q000235:genetics* / Q000378:metabolism; D009842:Oligopeptides / Q000378:metabolism; D015202:Protein Engineering; D011817:Rabbits; D011991:Receptors, Virus / Q000378:metabolism; D011994:Recombinant Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D053585:Virus Attachment; D053586:Virus Internalization
chemical_list: C099517:Ala-Trp-Arg-His-Pro-Gln-Phe-Gly-Gly; D036388:Ephrin-B2; D009842:Oligopeptides; D011991:Receptors, Virus; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; C116204:attachment protein G
keywords: Attachment protein; Binding assay; Hendra virus; Leishmania tarentolae; Protein expression