scholarly journals Probing the Flavivirus Membrane Fusion Mechanism by Using Monoclonal Antibodies

2007 ◽  
Vol 81 (20) ◽  
pp. 11526-11531 ◽  
Author(s):  
Karin Stiasny ◽  
Samantha Brandler ◽  
Christian Kössl ◽  
Franz X. Heinz
Keyword(s):  
Monoclonal Antibodies ◽  
Fusion Protein ◽  
Binding Sites ◽  
Fusion Inhibition ◽  
Tick Borne Encephalitis ◽  
Target Membrane ◽  
Initial Interaction ◽  
Tick Borne Encephalitis Virus ◽  
Late Stages

ABSTRACT In this study, we investigated in a flavivirus model (tick-borne encephalitis virus) the mechanisms of fusion inhibition by monoclonal antibodies directed to the different domains of the fusion protein (E) and to different sites within each of the domains by using in vitro fusion assays. Our data indicate that, depending on the location of their binding sites, the monoclonal antibodies impaired early or late stages of the fusion process, by blocking the initial interaction with the target membrane or by interfering with the proper formation of the postfusion structure of E, respectively. These data provide new insights into the mechanisms of flavivirus fusion inhibition by antibodies and their possible contribution to virus neutralization.

scholarly journals Effect of Membrane Curvature-Modifying Lipids on Membrane Fusion by Tick-Borne Encephalitis Virus

2004 ◽  
Vol 78 (16) ◽  
pp. 8536-8542 ◽  
Author(s):  
Karin Stiasny ◽  
Franz X. Heinz
Keyword(s):  
Fusion Protein ◽  
Membrane Fusion ◽  
Low Ph ◽  
Encephalitis Virus ◽  
Soluble Form ◽  
Inverted Cone ◽  
Outer Leaflet ◽  
Tick Borne Encephalitis ◽  
Target Membrane ◽  
Tick Borne Encephalitis Virus

ABSTRACT Enveloped viruses enter cells by fusion of their own membrane with a cellular membrane. Incorporation of inverted-cone-shaped lipids such as lysophosphatidylcholine (LPC) into the outer leaflet of target membranes has been shown previously to impair fusion mediated by class I viral fusion proteins, e.g., the influenza virus hemagglutinin. It has been suggested that these results provide evidence for the stalk-pore model of fusion, which involves a hemifusion intermediate (stalk) with highly bent outer membrane leaflets. Here, we investigated the effect of inverted-cone-shaped LPCs and the cone-shaped oleic acid (OA) on the membrane fusion activity of a virus with a class II fusion protein, the flavivirus tick-borne encephalitis virus (TBEV). This study included an analysis of lipid mixing, as well as of the steps preceding or accompanying fusion, i.e., binding to the target membrane and lipid-induced conformational changes in the fusion protein E. We show that the presence of LPC in the outer leaflet of target liposomes strongly inhibited TBEV-mediated fusion, whereas OA caused a very slight enhancement, consistent with a fusion mechanism involving a lipid stalk. However, LPC also impaired the low-pH-induced binding of a soluble form of the E protein to liposomes and its conversion into a trimeric postfusion structure that requires membrane binding at low pH. Because inhibition is already observed before the lipid-mixing step, it cannot be determined whether impairment of stalk formation is a contributing factor in the inhibition of fusion by LPC. These data emphasize, however, the importance of the composition of the target membrane in its interactions with the fusion peptide that are crucial for the initiation of fusion.

scholarly journals Membrane Interactions of the Tick-Borne Encephalitis Virus Fusion Protein E at Low pH

2002 ◽  
Vol 76 (8) ◽  
pp. 3784-3790 ◽  
Author(s):  
Karin Stiasny ◽  
Steven L. Allison ◽  
Juliane Schalich ◽  
Franz X. Heinz
Keyword(s):  
Fusion Protein ◽  
Membrane Binding ◽  
Low Ph ◽  
Encephalitis Virus ◽  
Soluble Form ◽  
Viral Fusion ◽  
Sequence Element ◽  
Viral Fusion Protein ◽  
Tick Borne Encephalitis ◽  
Tick Borne Encephalitis Virus

ABSTRACT Membrane fusion of the flavivirus tick-borne encephalitis virus is triggered by the mildly acidic pH of the endosome and is mediated by envelope protein E, a class II viral fusion protein. The low-pH trigger induces an oligomeric rearrangement in which the subunits of the native E homodimers dissociate and the monomeric subunits then reassociate into homotrimers. Here we provide evidence that membrane binding is mediated by the intermediate monomeric form of E, generated by low-pH-induced dissociation of the dimer. Liposome coflotation experiments revealed that association with target membranes occurred only when liposomes were present at the time of acidification, whereas pretreating virions at low pH in the absence of membranes resulted in the loss of their ability to stably attach to liposomes. With the cleavable cross-linker ethylene glycolbis(succinimidylsuccinate), it was shown that a truncated soluble form of the E protein (sE) could bind to membranes only when the dimers were free to dissociate at low pH, and binding could be blocked by a monoclonal antibody that recognizes the fusion peptide, which is at the distal tip of the E monomer but is buried in the native dimer. Surprisingly, analysis of the membrane-associated sE proteins revealed that they had formed trimers. This was unexpected because this protein lacks a sequence element in the C-terminal stem-anchor region, which was shown to be essential for trimerization in the absence of a target membrane. It can therefore be concluded that the formation of a trimeric form of sE is facilitated by membrane binding. Its stability is apparently maintained by contacts between the ectodomains only and is not dependent on sequence elements in the stem-anchor region as previously assumed.  

scholarly journals Tick populations from endemic and non-endemic areas in Germany show differential susceptibility to TBEV

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Katrin Liebig ◽  
Mathias Boelke ◽  
Domenic Grund ◽  
Sabine Schicht ◽  
Andrea Springer ◽  
...  
Keyword(s):  
Differential Susceptibility ◽  
Feeding Rates ◽  
Infection Rates ◽  
Lower Saxony ◽  
Tick Borne Encephalitis ◽  
The Central Nervous System ◽  
Tick Borne Encephalitis Virus ◽  
In Vitro Feeding ◽  
Tbev Infection

Abstract Tick-borne encephalitis virus (TBEV) is endemic in twenty-seven European countries, transmitted via the bite of an infected tick. TBEV is the causative agent of one of the most important viral diseases of the central nervous system (CNS). In Germany, 890 human cases were registered between the years 2018–2019. The castor bean tick, Ixodes ricinus, is the TBEV vector with the highest importance in Central Europe, including Germany. Despite the nationwide distribution of this tick species, risk areas of TBEV are largely located in Southern Germany. To increase our understanding of TBEV-tick interactions, we collected ticks from different areas within Germany (Haselmühl/Bavaria, Hanover/Lower Saxony) and infected them via an in vitro feeding system. A TBEV isolate was obtained from an endemic focus in Haselmühl. In two experimental series conducted in 2018 and 2019, ticks sampled in Haselmühl (TBEV focus) showed higher artificial feeding rates, as well as higher TBEV infections rates than ticks from the non-endemic area (Hanover). Other than the tick origin, year and month of the infection experiment as well as co-infection with Borrelia spp., had a significant impact on TBEV Haselmühl infection rates. Taken together, these findings suggest that a specific adaptation of the tick populations to their respective TBEV virus isolates or vice versa, leads to higher TBEV infection rates in those ticks. Furthermore, co-infection with other tick-borne pathogens such as Borrelia spp. can lower TBEV infection rates in specific populations.

scholarly journals Characterization of Monoclonal Antibodies against Hokkaido Strain Tick-Borne Encephalitis Virus

2000 ◽  
Vol 44 (6) ◽  
pp. 533-536 ◽  
Author(s):  
Kimiyo Komoro ◽  
Daisuke Hayasaka ◽  
Tetsuya Mizutani ◽  
Hiroaki Kariwa ◽  
Ikuo Takashima
Keyword(s):  
Monoclonal Antibodies ◽  
Encephalitis Virus ◽  
Tick Borne Encephalitis ◽  
Tick Borne Encephalitis Virus

scholarly journals Spectrum of antiviral activity of 4-aminopyrimidine N-oxides against a broad panel of tick-borne encephalitis virus strains

2020 ◽  
Vol 28 ◽  
pp. 204020662094346
Author(s):  
Evgenia V Dueva ◽  
Ksenia K Tuchynskaya ◽  
Liubov I Kozlovskaya ◽  
Dmitry I Osolodkin ◽  
Kseniya N Sedenkova ◽  
...  
Keyword(s):  
Encephalitis Virus ◽  
Northern Eurasia ◽  
Virus Particles ◽  
E Protein ◽  
Tick Borne Encephalitis ◽  
Tbev Strain ◽  
Amino Phenol ◽  
Tick Borne Encephalitis Virus ◽  
Immature Virus

Tick-borne encephalitis is an important human arbovirus neuroinfection spread across the Northern Eurasia. Inhibitors of tick-borne encephalitis virus (TBEV) strain Absettarov, presumably targeting E protein n-octyl-β-d-glucoside (β-OG) pocket, were reported earlier. In this work, these inhibitors were tested in vitro against seven strains representing three main TBEV subtypes. The most potent compound, 2-[(2-methyl-1-oxido-5,6,7,8-tetrahydroquinazolin-4-yl)amino]-phenol, showed EC50 values lower than 22 µM against all the tested strains. Nevertheless, EC50 values for virus samples of certain strains demonstrated a substantial variation, which appeared to be consistent with the presence of E protein not only in infectious virions, but also in non-infectious and immature virus particles, protein aggregates, and membrane complexes.

scholarly journals Site-specific inhibition of myosin-mediated motility in vitro by monoclonal antibodies.

1985 ◽  
Vol 100 (4) ◽  
pp. 1024-1030 ◽  
Author(s):  
P F Flicker ◽  
G Peltz ◽  
M P Sheetz ◽  
P Parham ◽  
J A Spudich
Keyword(s):  
Electron Microscopy ◽  
Monoclonal Antibodies ◽  
Binding Sites ◽  
Functional Assay ◽  
Specific Inhibition ◽  
Myosin Molecule ◽  
Site Specific ◽  
Dictyostelium Myosin ◽  
Cell Biol

Monoclonal antibodies directed against seven different sites on Dictyostelium myosin (Peltz, G., J. A. Spudich, and P. Parham, 1985, J. Cell Biol., 100: 1016-1023) were tested for their ability to inhibit movement of myosin in vitro, using the Nitella-based myosin-mediated bead movement assay (Sheetz, M. P., R. Chasan, and J. A. Spudich, 1984, J. Cell Biol., 99: 1867-1871). To complement this functional assay, we located the binding sites of these antibodies by electron microscopy, using the rotary shadowing technique. One antibody bound to the 18,000-dalton light chain and inhibited movement completely. All of the remaining antibodies bound to various positions along the rod portion of the myosin molecule, which is approximately 1,800 A long. Antibodies that bound to the rod about 470, 680, and 1400 A from the head-tail junction did not alter myosin movement. One antibody appeared to bind very close to the head-tail junction and to inhibit movement 50%. Surprisingly, three antibodies that bound about 1,200 A from the head-tail junction inhibited movement completely. This inhibition did not depend on using intact IgG, since Fab' fragments had the same effect.

scholarly journals Antiviral Properties of Water Extracts of Mycelium Inonotus rheades (Pers.) P. Karst. (1882) against the Virus of Tick-Borne Encephalitis Virus in vitro

2021 ◽  
Vol 6 (1) ◽  
pp. 55-59
Author(s):  
M. A. Khasnatinov ◽  
T. G. Gornostai ◽  
I. S. Solovarov ◽  
M. S. Polyakova ◽  
G. A. Danchinova ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Antiviral Agents ◽  
Encephalitis Virus ◽  
Water Soluble ◽  
Xylotrophic Basidiomycetes ◽  
Tick Borne Encephalitis ◽  
Wood Polysaccharides ◽  
Tick Borne Encephalitis Virus ◽  
The Relationship

Background. Tick-borne encephalitis virus is dangerous and widespread pathogen that is transmitted to humans through the bites of hard ticks. Wild fungi, such as xylotrophic basidiomycetes, are widely used in traditional medicine to treat the infectious diseases and are promising natural sources of new antiviral agents. It was previously shown that aqueous extracts from the mycelium of the Inonotus rheades (Pers.) P. Karst. (1882) fungus exhibit significant antiviral activity against tick-borne encephalitis virus, however, the mechanisms of this activity remain unclear.Aim. To analyze the relationship between the virucidal properties of I. rheades extract and the substrate on which the cultivation was carried out.Materials and methods. The mycelium was grown either in a standard liquid medium with wort or on wooden disks from birch. Extracts of water-soluble polysaccharides were prepared from both mycelium samples. The concentration of infectious tick-borne encephalitis virus was determined using the method of titration of plaque-forming components (PFU). Approximately 30 000 PFU of tick-borne encephalitis virus was mixed with an equal volume of corresponding I. rheades extract at concentration of 8 mg/mL and incubated for 30 min at 37 °C. Afterwards, the residual infectivity of tick-borne encephalitis virus was determined using the identical virus sample incubated with sterile water as a reference.Results. It was found that treatment of tick-borne encephalitis virus with extracts from I. rheades mycelium resulted in inhibition of the infectivity of the virus in the cell culture. However, the same strain of I. rheades, grown on medium with wort, did not exhibit antiviral properties.Conclusions. Virucidal substances are likely to be not the main metabolites of the mycelium of I. rheades, but are rather metabolized wood polysaccharides. Further research is needed to more accurately identify the active ingredients and assess their antiviral activity.

scholarly journals Tick-Borne Encephalitis Virus Vaccine-Induced Human Antibodies Mediate Negligible Enhancement of Zika Virus Infection In Vitro and in a Mouse Model

mSphere ◽  
2018 ◽  
Vol 3 (1) ◽  
Author(s):  
James Duehr ◽  
Silviana Lee ◽  
Gursewak Singh ◽  
Gregory A. Foster ◽  
David Krysztof ◽  
...  
Keyword(s):  
Mouse Model ◽  
Dengue Virus ◽  
Human Plasma ◽  
Mouse Models ◽  
Zika Virus ◽  
Encephalitis Virus ◽  
Tick Borne Encephalitis ◽  
Tick Borne Encephalitis Virus

ABSTRACT Recent reports in the scientific literature have suggested that anti-dengue virus (DENV) and anti-West Nile virus (WNV) immunity exacerbates Zika virus (ZIKV) pathogenesis in vitro and in vivo in mouse models. Large populations of immune individuals exist for a related flavivirus (tick-borne encephalitis virus [TBEV]), due to large-scale vaccination campaigns and endemic circulation throughout most of northern Europe and the southern Russian Federation. As a result, the question of whether anti-TBEV immunity can affect Zika virus pathogenesis is a pertinent one. For this study, we obtained 50 serum samples from individuals vaccinated with the TBEV vaccine FSME-IMMUN (Central European/Neudörfl strain) and evaluated their enhancement capacity in vitro using K562 human myeloid cells expressing CD32 and in vivo using a mouse model of ZIKV pathogenesis. Among the 50 TBEV vaccinee samples evaluated, 29 had detectable reactivity against ZIKV envelope (E) protein by enzyme-linked immunosorbent assay (ELISA), and 36 showed enhancement of ZIKV infection in vitro. A pool of the most highly reacting and enhanced samples resulted in no significant change in the morbidity/mortality of ZIKV disease in immunocompromised Stat2−/− mice. Our results suggest that humoral immunity against TBEV is unlikely to enhance Zika virus pathogenesis in humans. No clinical reports indicating that TBEV vaccinees experiencing enhanced ZIKV disease have been published so far, and though the epidemiological data are sparse, our findings suggest that there is little reason for concern. This study also displays a clear relationship between the phylogenetic distance between two flaviviruses and their capacity for pathogenic enhancement. IMPORTANCE The relationship between serial infections of two different serotypes of dengue virus and more severe disease courses is well-documented in the literature, driven by so-called antibody-dependent enhancement (ADE). Recently, studies have shown the possibility of ADE in cells exposed to anti-DENV human plasma and then infected with ZIKV and also in mouse models of ZIKV pathogenesis after passive transfer of anti-DENV human plasma. In this study, we evaluated the extent to which this phenomenon occurs using sera from individuals immunized against tick-borne encephalitis virus (TBEV). This is highly relevant, since large proportions of the European population are vaccinated against TBEV or otherwise seropositive.

scholarly journals Corepression of the P1 Addiction Operon by Phd and Doc

1998 ◽  
Vol 180 (23) ◽  
pp. 6342-6351 ◽  
Author(s):  
Roy Magnuson ◽  
Michael B. Yarmolinsky
Keyword(s):  
Fusion Protein ◽  
Binding Sites ◽  
Wild Type ◽  
Dna Complexes ◽  
Present Evidence ◽  
Cooperative Interactions ◽  
Regulatory Properties

ABSTRACT The P1 plasmid addiction operon encodes Doc, a toxin that kills plasmid-free segregants, and Phd, an unstable antidote that neutralizes the toxin. Additionally, these products repress transcription of the operon. The antidote binds to two adjacent sites in the promoter. Here we present evidence concerning the regulatory role of the toxin, which we studied with the aid of a mutation,docH66Y. The DocH66Y protein retained the regulatory properties of the wild-type protein, but not its toxicity. In vivo, DocH66Y enhanced repression by Phd but failed to affect repression in the absence of Phd, suggesting that DocH66Y contacts Phd. In vitro, a MalE-DocH66Y fusion protein was found to bind Phd. Binding of toxin to antidote may be the physical basis for the neutralization of toxin. DocH66Y failed to bind DNA in vitro yet enhanced the affinity, cooperativity, and specificity with which Phd bound the operator. Although DocH66Y enhanced the binding of Phd to two adjacent Phd-binding sites, DocH66Y had relatively little effect on the binding of Phd to a single Phd-binding site, indicating that DocH66Y mediates cooperative interactions between adjacent Phd-binding sites. Several electrophoretically distinct protein-DNA complexes were observed with different amounts of DocH66Y relative to Phd. Maximal repression and specificity of DNA binding were observed with subsaturating amounts of DocH66Y relative to Phd. Analogous antidote-toxin pairs appear to have similar autoregulatory circuits. Autoregulation, by dampening fluctuations in the levels of toxin and antidote, may prevent the inappropriate activation of the toxin.

Sign in / Sign up

Export Citation Format

Share Document