Human recombinant Puumala virus antibodies: cross-reaction with other hantaviruses and use in diagnostics.

Salonen, Eeva‐Marjatta; Parren, Paul W.H.I.; Graus, Y.; Lundkvist, Åke; Fisicaro, Paola; Vapalahti, Olli; Kallio‐Kokko, Hannimari; Vaheri, Antti; Burton, Dennis R.

doi:10.1099/0022-1317-79-4-659

Cited by 19 publications

(12 citation statements)

References 36 publications

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“…In addition, binding of neutralizing and non-neutralizing MAbs to HNTV G C was mapped to a region that include most of domain III but no specific epitope was determined [43]. Several neutralizing MAb against PUUV have been selected [44–46], two of which were shown to recognize G C (human MAb 1C9 and bank vole MAb 4G2). A peptide scan assay was used to identify the linear epitopes for these MAb [39, 47, 48].…”

Section: Resultsmentioning

confidence: 99%

Crystal Structure of Glycoprotein C from a Hantavirus in the Post-fusion Conformation

et al. 2016

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Hantaviruses are important emerging human pathogens and are the causative agents of serious diseases in humans with high mortality rates. Like other members in the Bunyaviridae family their M segment encodes two glycoproteins, GN and GC, which are responsible for the early events of infection. Hantaviruses deliver their tripartite genome into the cytoplasm by fusion of the viral and endosomal membranes in response to the reduced pH of the endosome. Unlike phleboviruses (e.g. Rift valley fever virus), that have an icosahedral glycoprotein envelope, hantaviruses display a pleomorphic virion morphology as GN and GC assemble into spikes with apparent four-fold symmetry organized in a grid-like pattern on the viral membrane. Here we present the crystal structure of glycoprotein C (GC) from Puumala virus (PUUV), a representative member of the Hantavirus genus. The crystal structure shows GC as the membrane fusion effector of PUUV and it presents a class II membrane fusion protein fold. Furthermore, GC was crystallized in its post-fusion trimeric conformation that until now had been observed only in Flavi- and Togaviridae family members. The PUUV GC structure together with our functional data provides intriguing evolutionary and mechanistic insights into class II membrane fusion proteins and reveals new targets for membrane fusion inhibitors against these important pathogens.

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Section: Resultsmentioning

confidence: 99%

Crystal Structure of Glycoprotein C from a Hantavirus in the Post-fusion Conformation

et al. 2016

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“…The RNA source of the third library originated from splenic lymphocytes from a splenectomized patient (donor C) possessing a serum antibody titer of 1:19,000 against MV measured in a standard diagnostic enzyme-linked immunosorbent assay (ELISA) performed at the Swedish Institute for Infectious Disease Control (SIIDC), Stockholm, Sweden. The same library has previously been used for isolation of Fab molecules specific for Puumala virus (10,29). The IgG1 heavy-chain Fd regions and whole and light chains from the three libraries (A, B, and C) were expressed in the phage display vector pComb3H essentially as described earlier (8,26,29).…”

Section: Methodsmentioning

confidence: 99%

“…The same library has previously been used for isolation of Fab molecules specific for Puumala virus (10,29). The IgG1 heavy-chain Fd regions and whole and light chains from the three libraries (A, B, and C) were expressed in the phage display vector pComb3H essentially as described earlier (8,26,29).…”

Section: Methodsmentioning

confidence: 99%

Neutralizing Human Fab Fragments against Measles Virus Recovered by Phage Display

Nicacio

Williamson

Parren

et al. 2002

J Virol

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Five human recombinant Fab fragments (Fabs) specific for measles virus (MV) proteins were isolated from three antibody phage display libraries generated from RNAs derived from bone marrow or splenic lymphocytes from three MV-immune individuals. All Fabs reacted in an enzyme-linked immunosorbent assay with MV antigens. In radioimmunoprecipitation assays two of the Fabs, MV12 and MT14, precipitated an Ϸ80-kDa protein band corresponding to the hemagglutinin (H) protein from MV-infected Vero cell cultures, while two other Fabs, MT64 and GL29, precipitated an Ϸ60-kDa protein corresponding the nucleocapsid (N) protein. In competition studies with MV fusion, H-and N protein-specific monoclonal antibodies (MAbs), the H-specific Fabs predominantly blocked the binding of H-specific MAbs, while the N-specific Fabs blocked MAbs to N. In addition, N-specific Fabs bound to denatured MV N protein in Western blotting. The specificity of the fifth Fab, MV4, could not be determined. By plaque reduction assays, three of the five Fabs, MV4, MV12, and MT14, exhibited neutralizing activity (80% cutoff) against MV (LEC-KI strain) at concentrations ranging between Ϸ2 and 7 g ml ؊1 . Neutralization capacity against MV strains Edmonston and Schwarz was also detected, albeit at somewhat higher Fab concentrations. In conclusion, three neutralizing Fabs were isolated, two of them reactive against the H glycoprotein of MV and another reactive against an undefined epitope. This is the first study in which MV-neutralizing human recombinant Fab antibodies have been isolated from phage display libraries.

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“…In subsequent studies with the same Mabs, sequence analysis of the genomes of Mab escape mutants revealed that noncontiguous amino acids form neutralizing epitopes (i.e., conformational epitopes) on the envelope proteins [Wang et al, 1993;Kikuchi et al, 1998]. Conformational, neutralizing epitopes were also identified on PUUV using phage display to express random peptides and by reacting the expression products with two G2-specific Mabs to PUUV [Salonen et al, 1998, Heiskanen et al, 1999, Nicacio et al, 2000. Thus, evidence to date suggests that neutralizing epitopes of the G1 and G2 proteins of HFRS-causing hantaviruses are frequently conformational.…”

Section: Introductionmentioning

confidence: 99%

Generation of an HFRS patient‐derived neutralizing recombinant antibody to Hantaan virus G1 protein and definition of the neutralizing domain

Liang¹,

Mähler

Koch

et al. 2002

Journal of Medical Virology

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Hantaan virus (HTNV) in the Hantavirus genus, family Bunyaviridae, is the major cause of severe hemorrhagic fever with renal syndrome (HFRS). We prepared a combinatorial phage display library of human Fabs to HTNV from RNA extracted from the blood lymphocytes of a convalescent HFRS patient. We selected two G1 glycoproteinspecific clones and one nucleocapsid protein (N)-specific clone from the Fab library for further studies. The human Fab antibodies were converted to IgG form in baculovirus/insect cells system by using cassette vectors that we developed earlier. Characterization of the recombinant antibodies revealed that the two G1-specific IgGs, could bind to and neutralize HTNV but not Seoul virus (SEOV). The N-specific IgG did not neutralize either HTNV or SEOV. Sequence analysis revealed that the two G1-specific clones differed by only one predicted amino acid in their complementarity determining regions, CDR3. Epitope mapping studies were carried out with one of the two G1-specific clones and synthetic peptides representing portions of HTNV G1. Results indicated that the recombinant antibody recognizes the core amino acid sequence LTKTLVIGQ, which is found near the C-terminus of HTNV G1. These results are the first to define a neutralizing epitope on the G1 protein of HTNV using an antibody derived from an HFRS patient.

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Human recombinant Puumala virus antibodies: cross-reaction with other hantaviruses and use in diagnostics.

Cited by 19 publications

References 36 publications

Crystal Structure of Glycoprotein C from a Hantavirus in the Post-fusion Conformation

Crystal Structure of Glycoprotein C from a Hantavirus in the Post-fusion Conformation

Neutralizing Human Fab Fragments against Measles Virus Recovered by Phage Display

Generation of an HFRS patient‐derived neutralizing recombinant antibody to Hantaan virus G1 protein and definition of the neutralizing domain

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