title: Expression and antigenicity of recombinant human respiratory syncytial virus glycoproteins having different affinity tags.
abstract: Human respiratory syncytial virus (HRSV) is a main cause of lower respiratory tract infections in infants and the elderly. Glycoprotein (G) is major antigen on the viral surface, and plays a key role for virus entry. Therefore, purification of the glycoprotein of HRSV is critical for the development of HRSV vaccine and serological diagnosis. In this study, we report the design and characterization of glycoprotein engineered rationally to enhance the protein solubility and to facilitate efficient purification. We permuted HRSV glycoproteins with two tags: (i) an immunoglobulin (Ig) M signal peptide and a protein A B domain tag to render HRSV glycoprotein secret into the culture media and (ii) a foldon and 6 × histidine tag with or without transmembrane domain. Three recombinant baculoviruses were constructed: (i) transmembrane-truncated HRSV glycoprotein (amino acid positions 66-298) inserted with the N-terminal IgM signal peptide and protein A B domain (MG-GΔTM), (ii) truncated HRSV glycoprotein (amino acid positions 66-298) fused with a C-terminal foldon and 6 × histidine tag (GΔTM-FH), and (iii) full-length HRSV glycoprotein (amino acid positions 1-298) fused with a C-terminal foldon and 6 × histidine tag (G-FH). Highly soluble recombinant MG-GΔTM protein was clearly purified using one-step affinity chromatography with IgG-sepharose resin, whereas the recombinant G-FH protein and truncated GΔTM-FH were purified partially using nickel-resin. Although, the antigenicity of GΔTM-FH was stronger than highly mannose-rich MG-GΔTM protein, MG-GΔTM induced neutralizing antibodies efficiently in the mice to protect from infectious HRSV.
authors.affiliation: Division of Respiratory Viruses, Center for Infectious Diseases, Korea National Institute of Health, Cheongju-si, Chungbuk 28159, Republic of Korea.
authors.forename: Han Saem
authors.initials: HS
authors.lastname: Lee
authors.affiliation: Division of Respiratory Viruses, Center for Infectious Diseases, Korea National Institute of Health, Cheongju-si, Chungbuk 28159, Republic of Korea.
authors.forename: A-Reum
authors.initials: AR
authors.lastname: Kim
authors.affiliation: Division of Influenza Viruses, Center for Infectious Diseases, Korea National Institute of Health, Cheongju-si, Chungbuk 28159, Republic of Korea.
authors.forename: Kisoon
authors.initials: K
authors.lastname: Kim
authors.affiliation: Division of Respiratory Viruses, Center for Infectious Diseases, Korea National Institute of Health, Cheongju-si, Chungbuk 28159, Republic of Korea.
authors.forename: Wan-Ji
authors.initials: WJ
authors.lastname: Lee
authors.affiliation: Division of Respiratory Viruses, Center for Infectious Diseases, Korea National Institute of Health, Cheongju-si, Chungbuk 28159, Republic of Korea.
authors.forename: Sung Soon
authors.initials: SS
authors.lastname: Kim
authors.affiliation: Division of Respiratory Viruses, Center for Infectious Diseases, Korea National Institute of Health, Cheongju-si, Chungbuk 28159, Republic of Korea. Electronic address: youjin3693@gmail.com.
authors.forename: You-Jin
authors.initials: YJ
authors.lastname: Kim
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000276:immunology*; D000914:Antibodies, Viral / Q000276:immunology*; D005260:Female; D015870:Gene Expression; D006023:Glycoproteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D006801:Humans; D007114:Immunization; D051379:Mice; D008807:Mice, Inbred BALB C; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D012136:Respiratory Syncytial Viruses / Q000235:genetics* / Q000276:immunology; D061987:Sf9 Cells; D018411:Spodoptera; D014764:Viral Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification
chemical_list: D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D006023:Glycoproteins; D011994:Recombinant Proteins; D014764:Viral Proteins
keywords: Antigenicity; Baculovirus; Glycoprotein; Human respiratory syncytial virus; IgG-sepharose; Neutralizing antibody