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Jianhua Sui - One of the best experts on this subject based on the ideXlab platform.
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viral entry of hepatitis b and d viruses and bile salts transportation share common molecular determinants on sodium taurocholate cotransporting polypeptide
Journal of Virology, 2014Co-Authors: Huan Yan, Bo Peng, Zhiyi Jing, Bijie Ren, Yang Liu, Jianhua SuiAbstract:ABSTRACT The liver bile acids transporter sodium taurocholate cotransporting polypeptide (NTCP) is responsible for the majority of sodium-dependent bile salts uptake by hepatocytes. NTCP also functions as a cellular receptor for viral entry of hepatitis B virus (HBV) and hepatitis D virus (HDV) through a specific interaction between NTCP and the pre-S1 Domain of HBV large envelope protein. However, it remains unknown if these two functions of NTCP are independent or if they interfere with each other. Here we show that binding of the pre-S1 Domain to human NTCP blocks taurocholate uptake by the receptor; conversely, some bile acid substrates of NTCP inhibit HBV and HDV entry. Mutations of NTCP residues critical for bile salts binding severely impair viral infection by HDV and HBV; to a lesser extent, the residues important for sodium binding also inhibit viral infection. The mutation S267F, corresponding to a single nucleotide polymorphism (SNP) found in about 9% of the East Asian population, renders NTCP without either taurocholate transporting activity or the ability to support HBV or HDV infection in cell culture. These results demonstrate that molecular determinants critical for HBV and HDV entry overlap with that for bile salts uptake by NTCP, indicating that viral infection may interfere with the normal function of NTCP, and bile acids and their derivatives hold the potential for further development into antiviral drugs. IMPORTANCE Human hepatitis B virus (HBV) and its satellite virus, hepatitis D virus (HDV), are important human pathogens. Available therapeutics against HBV are limited, and there is no drug that is clinically available for HDV infection. A liver bile acids transporter (sodium taurocholate cotransporting polypeptide [NTCP]) critical for maintaining homeostasis of bile acids serves as a functional receptor for HBV and HDV. We report here that the NTCP-binding lipopeptide that originates from the first 47 amino acids of the pre-S1 Domain of the HBV L protein blocks taurocholate transport. Some bile salts dose dependently inhibit HBV and HDV infection mediated by NTCP; molecular determinants of NTCP critical for HBV and HDV entry overlap with that for bile acids transport. This work advances our understanding of NTCP-mediated HBV and HDV infection in relation to NTCP9s physiological function. Our results also suggest that bile acids or their derivatives hold potential for development into novel drugs against HBV and HDV infection.
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potent neutralization of severe acute respiratory syndrome sars coronavirus by a human mab to S1 protein that blocks receptor association
Proceedings of the National Academy of Sciences of the United States of America, 2004Co-Authors: Jianhua Sui, Swee Kee Wong, Wenhui Li, Michael J. Moore, Akikazu Murakami, Azaibi Tamin, Leslie J Matthews, Aimee St Clair Tallarico, Mobolaji Olurinde, Hyeryun ChoeAbstract:Effective prophylaxis and antiviral therapies are urgently needed in the event of reemergence of the highly contagious and often fatal severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) infection. We have identified eight recombinant human single-chain variable region fragments (scFvs) against the S1 Domain of spike (S) protein of the SARS-CoV from two nonimmune human antibody libraries. One scFv 80R efficiently neutralized SARS-CoV and inhibited syncytia formation between cells expressing the S protein and those expressing the SARS-CoV receptor angiotensin-converting enzyme 2 (ACE2). Mapping of the 80R epitope showed it is located within the N-terminal 261–672 amino acids of S protein and is not glycosylation-dependent. 80R scFv competed with soluble ACE2 for association with the S1 Domain and bound S1 with high affinity (equilibrium dissociation constant, Kd = 32.3 nM). A human IgG1 form of 80R bound S1 with a 20-fold higher affinity of 1.59 nM comparable to that of ACE2 (Kd = 1.70 nM), and neutralized virus 20-fold more efficiently than the 80R scFv. These data suggest that the 80R human monoclonal antibody may be a useful viral entry inhibitor for the emergency prophylaxis and treatment of SARS, and that the ACE2-binding site of S1 could be an attractive target for subunit vaccine and drug development.
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angiotensin converting enzyme 2 is a functional receptor for the sars coronavirus
Nature, 2003Co-Authors: Wenhui Li, Natalya Vasilieva, Swee Kee Wong, Michael A Berne, Mohan Somasundaran, Katherine Luzuriaga, John L. Sullivan, Jianhua Sui, Michael J. Moore, Thomas C GreenoughAbstract:Spike (S) proteins of coronaviruses, including the coronavirus that causes severe acute respiratory syndrome (SARS), associate with cellular receptors to mediate infection of their target cellS1,2. Here we identify a metallopeptidase, angiotensin-converting enzyme 2 (ACE2)3,4, isolated from SARS coronavirus (SARS-CoV)-permissive Vero E6 cells, that efficiently binds the S1 Domain of the SARS-CoV S protein. We found that a soluble form of ACE2, but not of the related enzyme ACE1, blocked association of the S1 Domain with Vero E6 cells. 293T cells transfected with ACE2, but not those transfected with human immunodeficiency virus-1 receptors, formed multinucleated syncytia with cells expressing S protein. Furthermore, SARS-CoV replicated efficiently on ACE2-transfected but not mock-transfected 293T cells. Finally, anti-ACE2 but not anti-ACE1 antibody blocked viral replication on Vero E6 cells. Together our data indicate that ACE2 is a functional receptor for SARS-CoV.
Philippe Gripon - One of the best experts on this subject based on the ideXlab platform.
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efficient inhibition of hepatitis b virus infection by acylated peptides derived from the large viral surface protein
Journal of Virology, 2005Co-Authors: Philippe Gripon, Isabelle Cannie, Stephan UrbanAbstract:The lack of an appropriate in vitro infection system for the major human pathogen hepatitis B virus (HBV) has prevented a molecular understanding of the early infection events of HBV. We used the novel HBV-infectible cell line HepaRG and primary human hepatocytes to investigate the interference of infection by HBV envelope protein-derived peptides. We found that a peptide consisting of the authentically myristoylated N-terminal 47 amino acids of the pre-S1 Domain of the large viral envelope protein (L protein) specifically prevented HBV infection, with a 50% inhibitory concentration (IC50) of 8 nM. The replacement of myristic acid with other hydrophobic moieties resulted in changes in the inhibitory activity, most notably by a decrease in the IC50 to picomolar concentrations for longer unbranched fatty acids. The obstruction of HepaRG cell susceptibility to HBV infection after short preincubation times with the peptides suggested that the peptides efficiently target and inactivate a receptor at the hepatocyte surface. Our data both shed light on the molecular mechanism of HBV entry into hepatocytes and provide a basis for the development of potent hepadnaviral entry inhibitors as a novel therapeutic concept for the treatment of hepatitis Beta.
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efficient inhibition of hepatitis b virus infection by acylated peptides derived from the large viral surface protein
Journal of Virology, 2005Co-Authors: Philippe Gripon, Isabelle Cannie, Stephan UrbanAbstract:The lack of an appropriate in vitro infection system for the major human pathogen hepatitis B virus (HBV) has prevented a molecular understanding of the early infection events of HBV. We used the novel HBV-infectible cell line HepaRG and primary human hepatocytes to investigate the interference of infection by HBV envelope protein-derived peptides. We found that a peptide consisting of the authentically myristoylated N-terminal 47 amino acids of the pre-S1 Domain of the large viral envelope protein (L protein) specifically prevented HBV infection, with a 50% inhibitory concentration (IC50) of 8 nM. The replacement of myristic acid with other hydrophobic moieties resulted in changes in the inhibitory activity, most notably by a decrease in the IC50 to picomolar concentrations for longer unbranched fatty acids. The obstruction of HepaRG cell susceptibility to HBV infection after short preincubation times with the peptides suggested that the peptides efficiently target and inactivate a receptor at the hepatocyte surface. Our data both shed light on the molecular mechanism of HBV entry into hepatocytes and provide a basis for the development of potent hepadnaviral entry inhibitors as a novel therapeutic concept for the treatment of hepatitis Β.
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an 80 kilodalton protein that binds to the pre S1 Domain of hepatitis b virus
Journal of Virology, 2000Co-Authors: Chun Jeih Ryu, Dae Yeon Cho, Philippe Gripon, Hee Sun Kim, Christiane Guguenguillouzo, Hyo Jeong HongAbstract:It has been suggested that hepatitis B virus (HBV) binds to a receptor on the plasma membrane of human hepatocytes via the pre-S1 Domain of the large envelope protein as an initial step in HBV infection. However, the nature of the receptor remains controversial. In an attempt to identify a cell surface receptor for HBV, purified recombinant fusion protein of the pre-S1 Domain of HBV with glutathione S-transferase (GST), expressed in Escherichia coli, was used as a ligand. The surface of human hepatocytes or HepG2 cells was biotinylated, and the cell lysate (precleared lysate) which did not bind to GST and glutathione-Sepharose beads was used as a source of receptor molecules. The precleared lysate of the biotinylated cells was incubated with the GST-pre-S1 fusion protein, and the bound proteins were visualized by Western blotting and enhanced chemiluminescence. An approximately 80-kDa protein (p80) was shown to bind specifically to the pre-S1 Domain of the fusion protein. The receptor binding assay using serially or internally deleted segments of pre-S1 showed that amino acid residues 12 to 20 and 82 to 90 are essential for the binding of pre-S1 to p80. p80 also bound specifically to the pre-S1 of native HBV particles. Analysis of the tissue and species specificity of p80 expression in several available human primary cultures and cell lines of different tissue origin showed that p80 expression is not restricted to human hepatocytes. Taken together the results suggest that p80 may be a component of the viral entry machinery.
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infection process of the hepatitis b virus depends on the presence of a defined sequence in the pre S1 Domain
Journal of Virology, 1999Co-Authors: Le J Seyec, Christiane Guguenguillouzo, Philippe Chouteau, Isabelle Cannie, Philippe GriponAbstract:During the life cycle of hepatitis B virus (HBV), the large envelope protein (L) plays a pivotal role. Indeed, this polypeptide is essential for viral assembly and probably for the infection process. By performing mutagenesis experiments, we have previously excluded a putative involvement of the pre-S2 Domain of the L protein in viral infectivity. In the present study, we have evaluated the role of the pre-S1 region in HBV infection. For this purpose, 21 mutants of the L protein were created. The entire pre-S1 Domain was covered by contiguous deletions of 5 amino acids. First, after transfection into HepG2 cells, the efficient expression of both glycosylated and unglycosylated L mutant proteins was verified. The secretion rate of envelope proteins was modified positively or negatively by deletions, indicating that the pre-S1 Domain contains several regulating sequences able to influence the surface protein secretion. The ability of mutant proteins to support the production of virions was then studied. Only the four C-terminal deletions, covering the 17 amino acids suspected to interact with the cytoplasmic nucleocapsids, inhibited virion release. Finally, the presence of the modified pre-S1 Domain at the external side of all secreted virions was confirmed, and their infectivity was assayed on normal human hepatocytes in primary culture. Only a short sequence including amino acids 78 to 87 tolerates internal deletions without affecting viral infectivity. These results confirm the involvement of the L protein in the infection step and demonstrate that the sequence between amino acids 3 and 77 is involved in this process.
Shi Bo Jiang - One of the best experts on this subject based on the ideXlab platform.
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antigenic and immunogenic characterization of recombinant baculovirus expressed severe acute respiratory syndrome coronavirus spike protein implication for vaccine design
Journal of Virology, 2006Co-Authors: Susanne Heck, Sara Lustigman, Shi Bo JiangAbstract:The spike (S) glycoprotein of severe acute respiratory syndrome coronavirus (SARS-CoV) mediates the receptor interaction and immune recognition and is considered a major target for vaccine design. However, its antigenic and immunogenic properties remain to be elucidated. In this study, we immunized mice with full-length S protein (FL-S) or its extracellular Domain (EC-S) expressed by recombinant baculoviruses in insect cells. We found that the immunized mice developed high titers of anti-S antibodies with potent neutralizing activities against SARS pseudoviruses constructed with the S proteins of Tor2, GD03T13, and SZ3, the representative strains of 2002 to 2003 and 2003 to 2004 human SARS-CoV and palm civet SARS-CoV, respectively. These data suggest that the recombinant baculovirus-expressed S protein vaccines possess excellent immunogenicity, thereby inducing highly potent neutralizing responses against human and animal SARS-CoV variants. The antigenic structure of the S protein was characterized by a panel of 38 monoclonal antibodies (MAbs) isolated from the immunized mice. The epitopes of most anti-S MAbs (32 of 38) were localized within the S1 Domain, and those of the remaining 6 MAbs were mapped to the S2 Domain. Among the anti-S1 MAbs, 17 MAbs targeted the N-terminal region (amino acids [aa] 12 to 327), 9 MAbs recognized the receptor-binding Domain (RBD; aa 318 to 510), and 6 MAbs reacted with the C-terminal region of S1 Domain that contains the major immunodominant site (aa 528 to 635). Strikingly, all of the RBD-specific MAbs had potent neutralizing activity, 6 of which efficiently blocked the receptor binding, confirming that the RBD contains the main neutralizing epitopes and that blockage of the receptor association is the major mechanism of SARS-CoV neutralization. Five MAbs specific for the S1 N-terminal region exhibited moderate neutralizing activity, but none of the MAbs reacting with the S2 Domain and the major immunodominant site in S1 showed neutralizing activity. All of the neutralizing MAbs recognize conformational epitopes. These data provide important information for understanding the antigenicity and immunogenicity of S protein and for designing SARS vaccines. This panel of anti-S MAbs can be used as tools for studying the structure and function of the SARS-CoV S protein.
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receptor binding Domain of sars cov spike protein induces highly potent neutralizing antibodies implication for developing subunit vaccine
Biochemical and Biophysical Research Communications, 2004Co-Authors: Yusen Zhou, Michael Farzan, Shuwen Liu, Zhihua Kou, Shi Bo JiangAbstract:Abstract The spike (S) protein of severe acute respiratory syndrome (SARS) coronavirus (CoV), a type I transmembrane envelope glycoprotein, consists of S1 and S2 Domains responsible for virus binding and fusion, respectively. The S1 contains a receptor-binding Domain (RBD) that can specifically bind to angiotensin-converting enzyme 2 (ACE2), the receptor on target cells. Here we show that a recombinant fusion protein (designated RBD-Fc) containing 193-amino acid RBD (residues 318–510) and a human IgG1 Fc fragment can induce highly potent antibody responses in the immunized rabbits. The antibodies recognized RBD on S1 Domain and completely inhibited SARS-CoV infection at a serum dilution of 1:10,240. Rabbit antisera effectively blocked binding of S1, which contains RBD, to ACE2. This suggests that RBD can induce highly potent neutralizing antibody responses and has potential to be developed as an effective and safe subunit vaccine for prevention of SARS.
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immunological reaction between the peptides from S1 Domain of sars coronavirus s protein and the serum from sars patients
Academic journal of the first medical college of PLA, 2004Co-Authors: Jia Jie Zhang, Jin Lin Hou, Zhan Hui Wang, Shi Bo JiangAbstract:OBJECTIVE To examine the immunological reactions between the peptides of SARS coronavirus (SARS-Cov) S-protein and the serum of SARS patients for identifying the SARS-Cov epitope. METHODS The peptides from S1 Domain of SARS-Cov S-protein were synthesized by peptide synthesizer, and the immunological reaction of the peptides with the serum of SARS patients were examined by means of ELISA. RESULTS The optical density value of the immunological reaction of synthesized 8 peptides with SARS patient serum was 1.5-2.6 times higher than that with normal serum. CONCLUSION The 8 peptides from S1 Domain of S protein appear to have low immunogenicity to the serum of SARS patients, indicating that these peptides may not be the epitope of the SARS-Cov.
Gaiping Zhang - One of the best experts on this subject based on the ideXlab platform.
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isolation and identification of a recombinant porcine epidemic diarrhea virus with a novel insertion in S1 Domain
Frontiers in Microbiology, 2021Co-Authors: Yunchao Liu, Qiang Wei, Yumei Chen, Wenqiang Jiao, Hua Feng, Jucai Wang, Yuhang Zhang, Gaiping ZhangAbstract:Porcine epidemic diarrhea virus (PEDV) is the major pathogen that causes diarrhea and high mortality in newborn piglets with devastating impact to the pig industry. Recombination and mutation are the main driving forces of viral evolution and genetic diversity of PEDV. In 2016, an outbreak of diarrhea in piglets occurred in an intensive pig farm in Central China. A novel PEDV isolate (called HNAY) was successfully isolated from clinical samples. Sequence analysis and alignment showed that HNAY possessed 21-nucleotide (nt) insertion in its S1 gene, which has never been reported in other PEDV isolates. Moreover, the sequence of the insertion was identical with the sequence fragment in PEDV N gene. Notably, the HNAY strain exhibited two unique mutations (T500A and L521Y) in the neutralizing epitopes of the S1 protein that were different from those of other PEDV variant strains and CV777-based vaccine strains. Additionally, PEDV HNAY might be derived from a natural recombination between two Chinese variant PEDV strains. Animal experiments demonstrated that HNAY displayed higher pathogenicity compared with two other clinical isolates. This study lays the foundation for better understanding of the genetic evolution and molecular pathogenesis of PEDV.
Stephan Urban - One of the best experts on this subject based on the ideXlab platform.
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fine mapping of pre s sequence requirements for hepatitis b virus large envelope protein mediated receptor interaction
Journal of Virology, 2010Co-Authors: Andreas Schulze, Alexa Schieck, Walter Mier, Stephan UrbanAbstract:Previous studies showed that the N-terminal 75 amino acids of the pre-S1 Domain of the hepatitis B virus (HBV) L protein are essential for HBV and hepatitis delta virus (HDV) infectivity. Consistently, synthetic lipopeptides encompassing this sequence or only parts of it efficiently block HBV and HDV infection, presumably through specific interference with a cellular receptor. Crucial for both virus infectivity and the inhibitory activity of the peptides are N-terminal myristoylation and a highly conserved motif within the N-terminal 48 amino acids. To refine the sequence requirements, we synthesized a series of HBV pre-S1 peptides containing deletions, point mutations, d-amino acid exchanges, or genotype-specific sequence permutations. Using the HepaRG cell line and a genotype D-derived virus, we determined the specific inhibitory activities of the peptides and found that (i) lipopeptides with an artificial consensus sequence inhibit HBV genotype D infection more potently than the corresponding genotype D peptides; (ii) point mutations, d-amino acid exchanges, or deletions introduced into the highly conserved part of the pre-S1 Domain result in an almost complete loss of activity; and (iii) the flanking sequences comprising amino acids 2 to 8, 16 to 20, and, to a less pronounced extent, 34 to 48 gradually increase the inhibitory activity, while amino acids 21 to 33 behave indifferently. Taken together, our data suggest that HBV pre-S1-mediated receptor interference and, thus, HBV receptor recognition form a highly specific process. It requires an N-terminal acyl moiety and a highly conserved sequence that is present in primate but not rodent or avian hepadnaviruses, indicating different entry pathways for the different family members.
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efficient inhibition of hepatitis b virus infection by acylated peptides derived from the large viral surface protein
Journal of Virology, 2005Co-Authors: Philippe Gripon, Isabelle Cannie, Stephan UrbanAbstract:The lack of an appropriate in vitro infection system for the major human pathogen hepatitis B virus (HBV) has prevented a molecular understanding of the early infection events of HBV. We used the novel HBV-infectible cell line HepaRG and primary human hepatocytes to investigate the interference of infection by HBV envelope protein-derived peptides. We found that a peptide consisting of the authentically myristoylated N-terminal 47 amino acids of the pre-S1 Domain of the large viral envelope protein (L protein) specifically prevented HBV infection, with a 50% inhibitory concentration (IC50) of 8 nM. The replacement of myristic acid with other hydrophobic moieties resulted in changes in the inhibitory activity, most notably by a decrease in the IC50 to picomolar concentrations for longer unbranched fatty acids. The obstruction of HepaRG cell susceptibility to HBV infection after short preincubation times with the peptides suggested that the peptides efficiently target and inactivate a receptor at the hepatocyte surface. Our data both shed light on the molecular mechanism of HBV entry into hepatocytes and provide a basis for the development of potent hepadnaviral entry inhibitors as a novel therapeutic concept for the treatment of hepatitis Beta.
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efficient inhibition of hepatitis b virus infection by acylated peptides derived from the large viral surface protein
Journal of Virology, 2005Co-Authors: Philippe Gripon, Isabelle Cannie, Stephan UrbanAbstract:The lack of an appropriate in vitro infection system for the major human pathogen hepatitis B virus (HBV) has prevented a molecular understanding of the early infection events of HBV. We used the novel HBV-infectible cell line HepaRG and primary human hepatocytes to investigate the interference of infection by HBV envelope protein-derived peptides. We found that a peptide consisting of the authentically myristoylated N-terminal 47 amino acids of the pre-S1 Domain of the large viral envelope protein (L protein) specifically prevented HBV infection, with a 50% inhibitory concentration (IC50) of 8 nM. The replacement of myristic acid with other hydrophobic moieties resulted in changes in the inhibitory activity, most notably by a decrease in the IC50 to picomolar concentrations for longer unbranched fatty acids. The obstruction of HepaRG cell susceptibility to HBV infection after short preincubation times with the peptides suggested that the peptides efficiently target and inactivate a receptor at the hepatocyte surface. Our data both shed light on the molecular mechanism of HBV entry into hepatocytes and provide a basis for the development of potent hepadnaviral entry inhibitors as a novel therapeutic concept for the treatment of hepatitis Β.
