- Open Access
Peptide inhibitors of dengue virus and West Nile virus infectivity
© Hrobowski et al; licensee BioMed Central Ltd. 2005
- Received: 20 May 2005
- Accepted: 01 June 2005
- Published: 01 June 2005
Viral fusion proteins mediate cell entry by undergoing a series of conformational changes that result in virion-target cell membrane fusion. Class I viral fusion proteins, such as those encoded by influenza virus and human immunodeficiency virus (HIV), contain two prominent alpha helices. Peptides that mimic portions of these alpha helices inhibit structural rearrangements of the fusion proteins and prevent viral infection. The envelope glycoprotein (E) of flaviviruses, such as West Nile virus (WNV) and dengue virus (DENV), are class II viral fusion proteins comprised predominantly of beta sheets. We used a physio-chemical algorithm, the Wimley-White interfacial hydrophobicity scale (WWIHS)  in combination with known structural data to identify potential peptide inhibitors of WNV and DENV infectivity that target the viral E protein. Viral inhibition assays confirm that several of these peptides specifically interfere with target virus entry with 50% inhibitory concentration (IC50) in the 10 μM range. Inhibitory peptides similar in sequence to domains with a significant WWIHS scores, including domain II (IIb), and the stem domain, were detected. DN59, a peptide corresponding to the stem domain of DENV, inhibited infection by DENV (>99% inhibition of plaque formation at a concentrations of <25 μM) and cross-inhibition of WNV fusion/infectivity (>99% inhibition at <25 μM) was also demonstrated with DN59. However, a potent WNV inhibitory peptide, WN83, which corresponds to WNV E domain IIb, did not inhibit infectivity by DENV. Additional results suggest that these inhibitory peptides are noncytotoxic and act in a sequence specific manner. The inhibitory peptides identified here can serve as lead compounds for the development of peptide drugs for flavivirus infection.
- West Nile Virus
- Japanese Encephalitis Virus
- Inhibitory Peptide
- Dengue Hemorrhagic Fever
- Fusion Peptide
Enveloped viruses utilize membrane-bound fusion proteins to mediate attachment and entry into specific target host cells. During the virion assembly process, newly synthesized envelope proteins are targeted to the endoplasmic reticulum and golgi apparatus where initial folding and post-transcriptional processing occurs, including multimerization, glycosylation, and proteolysis. This initial folding and processing is required to achieve a conformation where the proteins are held in a metastable state prior to virion release. Post virion release, the multimeric envelope proteins are poised to undergo structural rearrangement leading to fusion of the virion and the new target cell lipid bilayer membranes. Depending on the virus system, the rearrangement trigger can take the form of specific receptor binding, multiple receptor binding, decreased pH following receptor mediated endocytosis, or a combination of triggers.
The prototypic viral envelope fusion protein, the hemagglutinin of influenza virus, contains short alpha helical domains in the trimeric virion configuration. In response to receptor binding and decreased pH, the short helices rearrange with adjoining sequences to produce a longer helix, thus exposing an N-terminal fusion peptide that is believed to interact directly with the target cell membrane. This is followed by a hinge-like bending of the entire complex to adjoin and fuse the two lipid membranes [2, 3]. The structural rearrangements that result in extrusion of the fusion peptide and subsequent collapse involve alterations in packing between regions both within individual fusion proteins as well as between monomeric subunits in the trimeric structures. Several disparate viruses, including arenaviruses, coronaviruses, filoviruses, orthomyxoviruses, paramyxoviruses and retroviruses, encode similar proteins that together are classified as class I fusion proteins. These class I viral fusion proteins vary in length and sequence, but are similar in overall structure [4, 5].
Qureshi et al. (1990) demonstrated that a peptide from one of the two extended helical domains of the HIV-1 transmembrane protein can block virion infectivity. Subsequently, the FDA approved anti-HIV-1 drug Fuzeon™ (aka DP178, T-20, enfuvirtide) and other N- and C-helix inhibitory peptides were developed [6, 7]. These results have greatly motivated the search for other HIV-1 inhibitory peptides [8, 9]. Additional peptide mimics of the fusion proteins of other retroviruses, and of orthomyxoviruses, paramyxoviruses, filoviruses, coronaviruses, and herpesviruses have also been identified and shown to inhibit viral entry [10–18]
The envelope fusion proteins of several virus types, including the flaviviruses and alphaviruses, have a structure distinct from class I viral fusion proteins. The envelope glycoprotein (E) of the flavivirus tick-borne encephalitis virus (TBEV) consists of three domains: a structurally central amino terminal domain (domain I), a dimerization domain (domain II) and a carboxyl terminal Ig-like domain (domain III), all containing predominantly beta sheet folds . The primary sequence of E1, the fusion protein of Semliki Forest virus, an alphavirus, revealed a remarkable fit to the scaffold of TBEV E  suggesting the existence of a second class of viral fusion proteins. The dengue virus (DENV) E protein has also been shown to have a class II structure . Recent studies of flavivirus virions and proteins by cryoelectron microscopy and crystal structure analysis have lead to a greatly increased understanding of the function of these class II viral envelope proteins, including the structural rearrangements they undergo during maturation, triggering and fusion [21–28].
The flaviviruses, which include DENV, West Nile virus (WNV), yellow fever virus, Japanese encephalitis virus (JEV), and TBEV, among others, are transmitted between vertebrate hosts by insect vectors. The most serious manifestations of DENV infection are dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). There are four serotypes of DENV (1–4), which together cause an estimated 50 million human infections per year , and each can cause DF, DHF or DSS. Because of the phenomenon of antibody-dependent enhancement (ADE), or other immune phenomena, protection against one DENV serotype increases the risk of DHF or DSS when the individual is exposed to another serotype [30–32]. Cross-reactive, but non-neutralizing antibodies can mediate entry of DENV into macrophages, dendritic cells and other viral target cells via Fc receptors, increasing virus titers and thus pathology. Multivalent DENV vaccines have shown some promise in humans [32–39] and in nonhuman primate studies [40, 41], but face several obstacles. Antiviral drugs, which target each of the four serotypes of DENV without enhancing pathogenesis of any serotype, are urgently needed. The recent introduction of WNV in the United States further highlights the public health challenges posed by flaviviruses. No effective vaccine or antiviral drug therapy is currently available against either DENV or WNV.
Although there are many differences between the structures of class I and class II viral fusion proteins, we hypothesized that they function through a similar membrane fusion mechanism involving rearrangements of domains, and that peptides mimicking portions of class II viral fusion proteins would inhibit virion fusion and entry steps thereby serving as lead compounds for the development of antivirals. We used a physio-chemical algorithm, the Wimley-White interfacial hydrophobicity scale  in combination with known structural data to predict regions of the DENV and WNV E proteins that may play roles in protein-protein rearrangements or bilayer membrane interactions during the entry and fusion process. Several of these peptides specifically inhibit DENV or WNV infection.
Identification of Flavivirus inhibitory peptides
Initial peptides synthesized and tested for inhibitory activity.
% Inhibition +/- SD
17 +/- 10
7 +/- 4
25 +/- 8
93 +/- 2
4 +/- 13
56 +/- 5
70 +/- 2
Determination of 50% inhibitory concentrations
Specificity of peptide inhibitory activity
Non-synergistic activity of combined peptides
When added in combination, peptides that block entry at different steps or that target different domains may produce greater inhibition of DENV-2 infectivity than either peptide alone. Synergistic or antagonistic effects are also possible, if a peptide that alters protein-protein interactions allows greater or lesser access to E domains targeted by another peptide. Since the WN53, WN83 and DN59 peptides all inhibited WNV entry, the possibility of antagonistic or synergistic function was examined by testing WN53 and DN59 alone or in combination at three concentrations (5, 10 and 20 μM). At all three concentrations, the peptide combination was more effective than WN53 alone, but less effective than DN59 alone. This indicates that the activity of the WN53 peptide has an antagonistic effect on the function of the DN59 peptide (Fig. 6).
Synthetic peptides corresponding to sequences in DENV and WNV E proteins were identified that inhibited infectivity of these viral pathogens of major public health importance. The inhibitory effects of these peptides were dose dependent with IC50s in the range of 10 μM. Several of the most potent of these peptides showed no inhibitory activity against SINV, an alphavirus that possesses a class II viral fusion protein with a similar overall structure as flavivirus E. Scrambled peptides with the same amino acid composition as the inhibitory peptides, but with a different primary sequence, failed to inhibit DENV and WNV infection. None of the DENV or WNV inhibitory peptides induced gross cytopathic effects, killing of cultured cells or showed evidence of in vitro cellular toxicity. These results indicate that these inhibitory peptides function through a sequence-specific mechanism and are not merely cytotoxic.
Membrane fusion by both class I and II viral fusion proteins is initiated by interaction of the fusion peptide with the target cell membrane. In class I viral fusion proteins, a subsequent rearrangement of a trimer of the proteins, each with two α helices, to form a six-helix bundle brings the viral and cell membranes into closer proximity. Inhibitors of viruses with class I fusion proteins, such as Fuzeon™ that mimic a portion of one or the other of the two α helices, interfere with a step proximal to six-helix bundle formation possibly by forming an inactive aggregate with the opposite helix. Recent studies indicate that after insertion of the fusion peptide, class II viral fusion proteins likewise undergo rearrangements. In this case, intraprotein interactions may occur between the stem domain and domains I, II and/or III [21–28, 46]. According to this model, the viral and cellular membranes are brought closer by interactions of the stem with other portions of E, resulting in bilayer fusion. DN59, WN53 and WN83 peptides may interfere with the intramolecular interactions between the stem and other portions of class II viral fusion proteins, a possibility suggested previously [23, 24, 46].
Two of the inhibitory peptides (WN53 and WN83) are designed from overlapping regions of the E protein domain I/II junction and are specifically inhibitory against WNV. Recently, other investigators have hypothesized that small molecule inhibitors to this domain I/II junction region might be developed. Modis et al (2003; 2004) predicted that interactions near this region (the k-l loop) that are involved in the rotational changes between these domains might be blocked by small molecule inhibitors. However, our similar peptides designed from the analogous region of the DENV E protein (D57 and D81) failed to inhibit DENV infectivity.
The possibility that WN53, WN83 and DN59 interact with some target cell surface component to exert their inhibitory effects cannot be ruled out. However, the majority of flavivirus neutralizing antibodies that appear to be involved in receptor blocking bind to domain III, and soluble domain III itself can block flavivirus entry, apparently through competition for cellular receptors [47–51]. In contrast, the domains that correspond to WN53, WN83 and DN59 peptides, IIb and the pre-anchor stem, appear to be involved in structural rearrangements during fusion, rather than direct interactions with cellular receptors. Interestingly, previous observations indicate that some monoclonal antibodies block virion entry at a post attachment step, indicating that they may interfere with conformational changes necessary for fusion . That possibility that some antibodies can gain access to regions important for conformational changes and block these changes suggests that these inhibitory peptide regions might be candidates for novel vaccine designs that either utilize the inhibitory peptide regions directly as antigens, or target the regions that interact with the inhibitory peptides. Further studies are needed to define the exact mechanism of inhibition of these DENV and WNV peptides, and the specific nature or location of their interactions with viral targets.
Alignment of preanchor domain sequences from representative flaviviruses.
Pre-anchor sequence b
IC50s in the μM range have been considered promising for class I viral fusion protein inhibitor development [54, 55]. Thus, the peptides identified here can serve as lead compounds that may be developed as peptide drugs against the four serotypes of DENV, WNV and potentially other flaviviruses. We anticipate that such peptide inhibitors may be as successful as the HIV-1 inhibitory peptide Fuzeon™. Unlike persistant HIV infections, immune responses against DENV and other flaviviruses are capable of clearing the viruses in individuals that survive the initial infection. By reducing the viral load during the initial stages of infection, it may be possible to extend the window of time during which an immune response could arise, and thus enable more individuals to control, eliminate and survive infections by these agents. Evidence for the ability to therapeutically intervene in flavivirus-induced diseases has been demonstrated with the recent observation that administration of neutralizing antibodies against WNV can be curative, even after symptom initiation . Development of resistant mutants will be a concern, but should be a less problematic than in the case of long-term treatment of persistent retroviral infections.
It is worth noting that the HIV inhibitor Fuzeon™, was initially identified using a predictive strategy without the availability of structural data [6, 7, 57]. The fact that we developed these peptides using a predictive algorithm validates our approach as well as the accuracy of the flavivirus E protein structural data. A similar approach may be useful for the large number of other viruses with class II envelope fusion proteins with or without known structures.
Design and synthesis of peptides
Sequences of DENV and WNV E with positive Wimley-White interfacial hydrophobicity scale scores were determined using the program Membrane Protein eXplorer . After consideration of the known secondary structures for several subdomains of E, selected peptides were synthesized by solid-phase conventional N-α-9-flurenylmethyloxycarbonyl chemistry (Genemed Synthesis, San Francisco, CA). Peptides were purified by reverse-phase high performance liquid chromatography and confirmed by amino acid analysis and electrospray mass spectrometry. Peptide stock solutions were prepared in 20% (v/v) dimethyl sulfoxide (DMSO): 80% (v/v) H20, and concentrations determined by absorbance of aromatic side chains at 280 nm. Scrambled peptides sequences were obtained by drawing from a hat.
Viruses and Cells
DENV strain New Guinea-2 and WNV strain Egypt 101 were obtained from R. Tesh at the World Health Organization Arbovirus Reference Laboratory at the University of Texas at Galveston. DENV and WNV were propagated in the African green monkey kidney epithelial cell line, LLCKM-2, a gift of K. Olsen at Colorado State University. Sindbis virus (SINV) containing the enhanced green fluorescent protein (EGFP) protein expression cassette was obtained from K. Ryman at Louisiana State University at Shreveport and was propagated in baby hamster kidney cells. All cells were grown in Dulbecco's modified eagle medium (DMEM) with 10% (v/v) fetal bovine serum (FBS), 100 U/ml penicillin G and 100 mg/ml streptomycin, at 37°C with 5% (v/v) CO2.
Viral plaque reduction assays
LLCKM-2 target cells were seeded at a density of 3 × 105 cells in each well of a 6-well plate 48 h prior to infection. Approximately 200-focus forming units (FFU) of DENV, WNV, or SINV/EGFP were incubated with or without peptides in serum-free DMEM for 1 h at rt. Virus/peptide or virus/control mixtures were allowed to infect confluent LLCKM-2 monolayers for 1 h at 37°C, after which time the medium was removed and the cells were washed once with phosphate buffered saline (PBS) and overlaid with fresh DMEM/10% (v/v) FBS containing 0.85% (w/v) SeaPlaque Agarose (Cambrex Bio Science, Rockland, ME). Cells were then incubated at 37°C with 5% CO2 for 1 day (Sindbis virus), 3 days (WNV) or 6 days (DNV). Sindbis virus infections were quantified by directly counting green fluorescing foci. Cultures infected with DENV were fixed with 10% formalin overnight at 4°C and permeablized with 70% (v/v) ethanol prior to immunostaining and visualization using a human polyclonal anti-flavivirus antibody (a gift of V. Vorndam, CDC, San Juan) followed by horseradish peroxidase (HRP) conjugated goat anti-human immunoglobulin (Pierce, Rockford, IL) and AEC chromogen substrate (Dako, Carpinteria, CA). WNV plaques were similarly visualized using a mouse anti-WNV antibody (Chemicon, Temecula, CA) and an HRP conjugated goat anti-mouse antibody (Dako, Carpinteria, CA).
Peptide cytotoxicity was measured using the TACS™ MTT cell proliferation assay (R&D systems, Minneapolis, MN) according to the manufacturer's instructions.
The authors would like to thank S. Isern, B. Sainz, W. Wimley and W. Gallaher for helpful discussions and technical assistance.
- White SH, Snider C, Jaysinghe S, Kim J: Membrane Protein Explorer version 2.2a. http://blancobiomoluciedu/mpex/. 2003Google Scholar
- Carr CM, Kim PS: A spring-loaded mechanism for the conformational change of influenza hemagglutinin. Cell. 1993, 73: 823-832. 10.1016/0092-8674(93)90260-W.View ArticlePubMedGoogle Scholar
- Wilson IA, Skehel JJ, Wiley DC: Structure of the haemagglutinin membrane glycoprotein of influenza virus at 3 A resolution. Nature. 1981, 289: 366-373. 10.1038/289366a0.View ArticlePubMedGoogle Scholar
- Gallaher WR: Similar structural models of the transmembrane glycoproteins of Ebola and avian sarcoma viruses. Cell. 1996, 85: 1-2. 10.1016/S0092-8674(00)81248-9.View ArticleGoogle Scholar
- Gallaher WR, Ball JM, Garry RF, Griffin MC, Montelaro RC: A general model for the transmembrane proteins of HIV and other retroviruses. AIDS Res Hum Retroviruses. 1989, 5: 431-440.View ArticlePubMedGoogle Scholar
- Wild CT, Shugars DC, Greenwell TK, McDanal CB, Matthews TJ: Peptides corresponding to a predictive alpha-helical domain of human immunodeficiency virus type 1 gp41 are potent inhibitors of virus infection. Proc Natl Acad Sci U S A. 1994, 91: 9770-9774.PubMed CentralView ArticlePubMedGoogle Scholar
- Wild C, Greenwell T, Matthews T: A synthetic peptide from HIV-1 gp41 is a potent inhibitor of virus-mediated cell-cell fusion. AIDS Research & Human Retroviruses. 1993, 9: 1051-1053.View ArticleGoogle Scholar
- Pozniak A: HIV fusion inhibitors. J HIV Ther. 2001, 6: 91-94.PubMedGoogle Scholar
- Sodroski JG: HIV-1 entry inhibitors in the side pocket. Cell. 1999, 99: 243-246. 10.1016/S0092-8674(00)81655-4.View ArticlePubMedGoogle Scholar
- Lambert DM, Barney S, Lambert AL, Guthrie K, Medinas R, Davis DE, Bucy T, Erickson J, Merutka G, Petteway SRJ: Peptides from conserved regions of paramyxovirus fusion (F) proteins are potent inhibitors of viral fusion. Proc Natl Acad Sci U S A. 1996, 93: 2186-2191. 10.1073/pnas.93.5.2186.PubMed CentralView ArticlePubMedGoogle Scholar
- Young JK, Li D, Abramowitz MC, Morrison TG: Interaction of peptides with sequences from the Newcastle disease virus fusion protein heptad repeat regions. J Virol. 1999, 73: 5945-5956.PubMed CentralPubMedGoogle Scholar
- Watanabe S, Takada A, Watanabe T, Ito H, Kida H, Kawaoka Y: Functional importance of the coiled-coil of the Ebola virus glycoprotein. J Virol. 2000, 74: 10194-10201. 10.1128/JVI.74.21.10194-10201.2000.PubMed CentralView ArticlePubMedGoogle Scholar
- Bosch BJ, van der Zee R, de Haan CA, Rottier PJ: The coronavirus spike protein is a class I virus fusion protein: structural and functional characterization of the fusion core complex. J Virol. 2003, 77: 8801-8811. 10.1128/JVI.77.16.8801-8811.2003.PubMed CentralView ArticlePubMedGoogle Scholar
- Bultmann H, Brandt CR: Peptides containing membrane-transiting motifs inhibit virus entry. J Biol Chem. 2002, 277: 36018-36023. 10.1074/jbc.M204849200.View ArticlePubMedGoogle Scholar
- Markosyan RM, Bates P, Cohen FS, Melikyan GB: A study of low pH-induced refolding of Env of avian sarcoma and leukosis virus into a six-helix bundle. Biophys J. 2004, 87: 3291-3298. 10.1529/biophysj.104.047696.PubMed CentralView ArticlePubMedGoogle Scholar
- Rapaport D, Ovadia M, Shai Y: A synthetic peptide corresponding to a conserved heptad repeat domain is a potent inhibitor of Sendai virus-cell fusion: an emerging similarity with functional domains of other viruses. Embo J. 1995, 14: 5524-5531.PubMed CentralPubMedGoogle Scholar
- Okazaki K, Kida H: A synthetic peptide from a heptad repeat region of herpesvirus glycoprotein B inhibits virus replication. J Gen Virol. 2004, 85: 2131-2137. 10.1099/vir.0.80051-0.View ArticlePubMedGoogle Scholar
- Yao Q, Compans RW: Peptides corresponding to the heptad repeat sequence of human parainfluenza virus fusion protein are potent inhibitors of virus infection. Virology. 1996, 223: 103-112. 10.1006/viro.1996.0459.View ArticlePubMedGoogle Scholar
- Rey FA, Heinz FX, Mandl C, Kunz C, Harrison SC: The envelope glycoprotein from tick-borne encephalitis virus at 2 A resolution. Nature. 1995, 375: 291-298. 10.1038/375291a0.View ArticlePubMedGoogle Scholar
- Lescar J, Roussel A, Wien MW, Navaza J, Fuller SD, Wengler G, Rey FA: The Fusion glycoprotein shell of Semliki Forest virus: an icosahedral assembly primed for fusogenic activation at endosomal pH. Cell. 2001, 105: 137-148. 10.1016/S0092-8674(01)00303-8.View ArticlePubMedGoogle Scholar
- Kuhn RJ, Zhang W, Rossmann MG, Pletnev SV, Corver J, Lenches E, Jones CT, Mukhopadhyay S, Chipman PR, Strauss EG, Baker TS, Strauss JH: Structure of dengue virus: implications for flavivirus organization, maturation, and fusion. Cell. 2002, 108: 717-725. 10.1016/S0092-8674(02)00660-8.PubMed CentralView ArticlePubMedGoogle Scholar
- Zhang Y, Corver J, Chipman PR, Zhang W, Pletnev SV, Sedlak D, Baker TS, Strauss JH, Kuhn RJ, Rossmann MG: Structures of immature flavivirus particles. Embo J. 2003, 22: 2604-2613. 10.1093/emboj/cdg270.PubMed CentralView ArticlePubMedGoogle Scholar
- Bressanelli S, Stiasny K, Allison SL, Stura EA, Duquerroy S, Lescar J, Heinz FX, Rey FA: Structure of a flavivirus envelope glycoprotein in its low-pH-induced membrane fusion conformation. Embo J. 2004, 23: 728-738. 10.1038/sj.emboj.7600064.PubMed CentralView ArticlePubMedGoogle Scholar
- Modis Y, Ogata S, Clements D, Harrison SC: Structure of the dengue virus envelope protein after membrane fusio. Nature. 2004, 427: 313-319. 10.1038/nature02165.View ArticlePubMedGoogle Scholar
- Modis Y, Ogata S, Clements D, Harrison SC: A ligand-binding pocket in the dengue virus envelope glycoprotein. Proc Natl Acad Sci U S A. 2003, 100: 6986-6991. 10.1073/pnas.0832193100.PubMed CentralView ArticlePubMedGoogle Scholar
- Mukhopadhyay S, Kim BS, Chipman PR, Rossmann MG, Kuhn RJ: Structure of West Nile virus. Science. 2003, 302: 248-10.1126/science.1089316.View ArticlePubMedGoogle Scholar
- Mukhopadhyay S, Kuhn RJ, Rossmann MG: A structural perspective of the flavivirus life cycle. Nat Rev Microbiol. 2005, 3: 13-22. 10.1038/nrmicro1067.View ArticlePubMedGoogle Scholar
- Rey FA: Dengue virus envelope glycoprotein structure: new insight into its interactions during viral entry. Proc Natl Acad Sci U S A. 2003, 100: 6899-6901. 10.1073/pnas.1332695100.PubMed CentralView ArticlePubMedGoogle Scholar
- WHO: Dengue and dengue haemorrhagic fever. 2002, http://www.who.int/mediacentre/factsheets/fs117/en/Google Scholar
- Hotta H, Sanchez LF, Takada H, Homma M, Kotani S: Enhancement of dengue virus infection in cultured mouse macrophages by lipophilic derivatives of muramyl peptides. Microbiol Immunol. 1985, 29: 533-541.View ArticlePubMedGoogle Scholar
- Halstead SB: Pathogenesis of dengue: challenges to molecular biology. Science. 1988, 239: 476-481.View ArticlePubMedGoogle Scholar
- Guy B, Chanthavanich P, Gimenez S, Sirivichayakul C, Sabchareon A, Begue S, Yoksan S, Luxemburger C, Lang J: Evaluation by flow cytometry of antibody-dependent enhancement (ADE) of dengue infection by sera from Thai children immunized with a live-attenuated tetravalent dengue vaccine. Vaccine. 2004, 22: 3563-3574. 10.1016/j.vaccine.2004.03.042.View ArticlePubMedGoogle Scholar
- Sabchareon A, Lang J, Chanthavanich P, Yoksan S, Forrat R, Attanath P, Sirivichayakul C, Pengsaa K, Pojjaroen-Anant C, Chokejindachai W, Jagsudee A, Saluzzo JF, Bhamarapravati N: Safety and immunogenicity of tetravalent live-attenuated dengue vaccines in Thai adult volunteers: role of serotype concentration, ratio, and multiple doses. Am J Trop Med Hyg. 2002, 66: 264-272.PubMedGoogle Scholar
- Edelman R, Wasserman SS, Bodison SA, Putnak RJ, Eckels KH, Tang D, Kanesa-Thasan N, Vaughn DW, Innis BL, Sun W: Phase I trial of 16 formulations of a tetravalent live-attenuated dengue vaccine. Am J Trop Med Hyg. 2003, 69: 48-60.PubMedGoogle Scholar
- Kanesa-Thasan N, Sun W, Ludwig GV, Rossi C, Putnak JR, Mangiafico JA, Innis BL, Edelman R: Atypical antibody responses in dengue vaccine recipients. Am J Trop Med Hyg. 2003, 69: 32-38.PubMedGoogle Scholar
- Kanesa-thasan N, Sun W, Kim-Ahn G, Van Albert S, Putnak JR, King A, Raengsakulsrach B, Christ-Schmidt H, Gilson K, Zahradnik JM, Vaughn DW, Innis BL, Saluzzo JF, Hoke CHJ: Safety and immunogenicity of attenuated dengue virus vaccines (Aventis Pasteur) in human volunteers. Vaccine. 2001, 19: 3179-3188. 10.1016/S0264-410X(01)00020-2.View ArticlePubMedGoogle Scholar
- Bhamarapravati N, Sutee Y: Live attenuated tetravalent dengue vaccine. Vaccine. 2000, 18 Suppl 2: 44-47. 10.1016/S0264-410X(00)00040-2.View ArticlePubMedGoogle Scholar
- Barrett AD: Current status of flavivirus vaccines. Ann N Y Acad Sci. 2001, 951: 262-271.View ArticlePubMedGoogle Scholar
- Sun W, Edelman R, Kanesa-Thasan N, Eckels KH, Putnak JR, King AD, Houng HS, Tang D, Scherer JM, Hoke CHJ, Innis BL: Vaccination of human volunteers with monovalent and tetravalent live-attenuated dengue vaccine candidates. Am J Trop Med Hyg. 2003, 69: 24-31.PubMedGoogle Scholar
- Guirakhoo F, Pugachev K, Zhang Z, Myers G, Levenbook I, Draper K, Lang J, Ocran S, Mitchell F, Parsons M, Brown N, Brandler S, Fournier C, Barrere B, Rizvi F, Travassos A, Nichols R, Trent D, Monath T: Safety and efficacy of chimeric yellow Fever-dengue virus tetravalent vaccine formulations in nonhuman primates. J Virol. 2004, 78: 4761-4775. 10.1128/JVI.78.9.4761-4775.2004.PubMed CentralView ArticlePubMedGoogle Scholar
- Guirakhoo F, Pugachev K, Arroyo J, Miller C, Zhang ZX, Weltzin R, Georgakopoulos K, Catalan J, Ocran S, Draper K, Monath TP: Viremia and immunogenicity in nonhuman primates of a tetravalent yellow fever-dengue chimeric vaccine: genetic reconstructions, dose adjustment, and antibody responses against wild-type dengue virus isolates. Virology. 2002, 298: 146-159. 10.1006/viro.2002.1462.View ArticlePubMedGoogle Scholar
- Wimley WC, White SH: Experimentally determined hydrophobicity scale for proteins at membrane interfaces. Nat Struct Biol. 1996, 3: 842-848. 10.1038/nsb1096-842.View ArticlePubMedGoogle Scholar
- Giannecchini S, Bonci F, Pistello M, Matteucci D, Sichi O, Rovero P, Bendinelli M: The membrane-proximal tryptophan-rich region in the transmembrane glycoprotein ectodomain of feline immunodeficiency virus is important for cell entry. Virology. 2004, 320: 156-166. 10.1016/j.virol.2003.12.001.View ArticlePubMedGoogle Scholar
- Garry RF, Dash S: Proteomics computational analyses suggest that hepatitis C virus E1 and pestivirus E2 envelope glycoproteins are truncated class II fusion proteins. Virology. 2003, 307: 255-265. 10.1016/S0042-6822(02)00065-X.View ArticlePubMedGoogle Scholar
- Garry CE, Garry RF: Proteomics computational analyses suggest that the carboxyl terminal glycoproteins of Bunyaviruses are class II viral fusion protein (beta-penetrenes). Theor Biol Med Model. 2004, 1: 10-10.1186/1742-4682-1-10.PubMed CentralView ArticlePubMedGoogle Scholar
- Gibbons DL, Vaney MC, Roussel A, Vigouroux A, Reilly B, Lepault J, Kielian M, Rey FA: Conformational change and protein-protein interactions of the fusion protein of Semliki Forest virus. Nature. 2004, 427: 320-325. 10.1038/nature02239.View ArticlePubMedGoogle Scholar
- Beasley DW, Barrett AD: Identification of neutralizing epitopes within structural domain III of the West Nile virus envelope protein. J Virol. 2002, 76: 13097-13100. 10.1128/JVI.76.24.13097-13100.2002.PubMed CentralView ArticlePubMedGoogle Scholar
- Volk DE, Beasley DW, Kallick DA, Holbrook MR, Barrett AD, Gorenstein DG: Solution structure and antibody binding studies of the envelope protein domain III from the New York strain of West Nile virus. J Biol Chem. 2004, 279: 38755-38761. 10.1074/jbc.M402385200.View ArticlePubMedGoogle Scholar
- Chen Y, Maguire T, Marks RM: Demonstration of binding of dengue virus envelope protein to target cells. J Virol. 1996, 70: 8765-8772.PubMed CentralPubMedGoogle Scholar
- Bhardwaj S, Holbrook M, Shope RE, Barrett AD, Watowich SJ: Biophysical characterization and vector-specific antagonist activity of domain III of the tick-borne flavivirus envelope protein. J Virol. 2001, 75: 4002-4007. 10.1128/JVI.75.8.4002-4007.2001.PubMed CentralView ArticlePubMedGoogle Scholar
- Wu KP, Wu CW, Tsao YP, Kuo TW, Lou YC, Lin CW, Wu SC, Cheng JW: Structural basis of a Flavivirus recognized by its neutralizing antibody: Solution structure of the domain III of the Japanese Encephalitis virus envelope protein. J Biol Chem. 2003Google Scholar
- Hung SL, Lee PL, Chen HW, Chen LK, Kao CL, King CC: Analysis of the steps involved in Dengue virus entry into host cells. Virology. 1999, 257: 156-167. 10.1006/viro.1999.9633.View ArticlePubMedGoogle Scholar
- Allison SL, Stiasny K, Stadler K, Mandl CW, Heinz FX: Mapping of functional elements in the stem-anchor region of tick-borne encephalitis virus envelope protein E. J Virol. 1999, 73: 5605-5612.PubMed CentralPubMedGoogle Scholar
- Liu S, Xiao G, Chen Y, He Y, Niu J, Escalante CR, Xiong H, Farmar J, Debnath AK, Tien P, Jiang S: Interaction between heptad repeat 1 and 2 regions in spike protein of SARS-associated coronavirus: implications for virus fusogenic mechanism and identification of fusion inhibitors. Lancet. 2004, 363: 938-947. 10.1016/S0140-6736(04)15788-7.View ArticlePubMedGoogle Scholar
- Sia SK, Carr PA, Cochran AG, Malashkevich VN, Kim PS: Short constrained peptides that inhibit HIV-1 entry. Proc Natl Acad Sci U S A. 2002, 99: 14664-14669. 10.1073/pnas.232566599.PubMed CentralView ArticlePubMedGoogle Scholar
- Oliphant T, Engle M, Nybakken GE, Doane C, Johnson S, Huang L, Gorlatov S, Mehlhop E, Marri A, Chung KM, Ebel GD, Kramer LD, Fremont DH, Diamond MS: Development of a humanized monoclonal antibody with therapeutic potential against West Nile virus. Nat Med. 2005, 11: 522-30. 10.1038/nm1240.PubMed CentralView ArticlePubMedGoogle Scholar
- Qureshi NM, Coy DH, Garry RF, LA H: Characterization of a putative cellular receptor for HIV-1 transmembrane glycoprotein using synthetic peptides. AIDS. 1990, 4: 553-558.View ArticlePubMedGoogle Scholar
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