scholarly journals Vaccinia Virus 4c (A26L) Protein on Intracellular Mature Virus Binds to the Extracellular Cellular Matrix Laminin

2006 ◽  
Vol 81 (5) ◽  
pp. 2149-2157 ◽  
Author(s):  
Wen-Ling Chiu ◽  
Chi-Long Lin ◽  
Min-Hsiang Yang ◽  
Der-Lii M. Tzou ◽  
Wen Chang
Keyword(s):  
Vaccinia Virus ◽  
Envelope Protein ◽  
Binding Activity ◽  
Viral Envelope ◽  
Wild Type ◽  
Viral Envelope Protein ◽  
Reading Frame ◽  
Mature Virus ◽  
Laminin Binding

ABSTRACT Vaccinia virus intracellular mature virus (IMV) binds to glycosaminoglycans (GAGs) on cells via three virion proteins, H3L, A27L, and D8L. In this study, we demonstrated that binding of IMV to BSC40 cells was competitively inhibited by soluble laminin but not by fibronectin or collagen V, suggesting that this cell surface extracellular matrix (ECM) protein may play a role in vaccinia virus entry. Moreover, IMV infection of GAG− sog9 cells was also inhibited by laminin, demonstrating that virion binding to laminin does not involve a prior interaction with GAGs. Furthermore, comparative envelope protein analyses of wild-type vaccinia virus strain Western Reserve, which binds to laminin, and of a mutant virus, IA27L, which does not, showed that the A26L open reading frame (ORF), encoding an envelope protein, was mutated in IA27L, resulting in A26L being absent from the IMV. Expression of the wild-type A26L ORF in IA27L resulted in laminin binding activity. Moreover, recombinant A26L protein bound to laminin in vitro with a high affinity, providing direct evidence that A26L is the laminin binding protein on IMV. In summary, these results reveal a novel role for the vaccinia viral envelope protein A26L in binding to the ECM protein laminin, an association that is proposed to facilitate IMV entry.

scholarly journals The YXXL Sequences of a Transmembrane Protein of Bovine Leukemia Virus Are Required for Viral Entry and Incorporation of Viral Envelope Protein into Virions

1999 ◽  
Vol 73 (2) ◽  
pp. 1293-1301 ◽  
Author(s):  
Kazunori Inabe ◽  
Masako Nishizawa ◽  
Shigeru Tajima ◽  
Kazuyoshi Ikuta ◽  
Yoko Aida
Keyword(s):  
Viral Entry ◽  
Envelope Protein ◽  
Bovine Leukemia Virus ◽  
Leukemia Virus ◽  
Transient Transfection ◽  
Viral Envelope ◽  
Tyrosine Residue ◽  
Wild Type ◽  
Viral Envelope Protein

ABSTRACT The cytoplasmic domain of an envelope transmembrane glycoprotein (gp30) of bovine leukemia virus (BLV) has two overlapping copies of the (YXXL)2 motif. The N-terminal motif has been implicated in in vitro signal transduction pathways from the external to the intracellular compartment and is also involved in infection and maintenance of high viral loads in sheep that have been experimentally infected with BLV. To determine the role of YXXL sequences in the replication of BLV in vitro, we changed the tyrosine or leucine residues of the N-terminal motif in an infectious molecular clone of BLV, pBLV-IF, to alanine to produce mutated proviruses designated Y487A, L490A, Y498A, L501A, and Y487/498A. Transient transfection of African green monkey kidney COS-1 cells with proviral DNAs that encoded wild-type and mutant sequences revealed that all of the mutated proviral DNAs synthesized mature envelope proteins and released virus particles into the growth medium. However, serial passages of fetal lamb kidney (FLK) cells, which are sensitive to infection with BLV, after transient transfection revealed that mutation of a second tyrosine residue in the N-terminal motif completely prevented the propagation of the virus. Similarly, Y498A and Y487/498A mutant BLV that was produced by the stably transfected COS-1 cells exhibited significantly reduced levels of cell-free virion-mediated transmission. Analysis of the protein compositions of mutant viruses demonstrated that lower levels of envelope protein were incorporated by two of the mutant virions than by wild-type and other mutant virions. Furthermore, a mutation of a second tyrosine residue decreased the specific binding of BLV particles to FLK cells and the capacity for viral penetration. Our data indicate that the YXXL sequences play critical roles in both viral entry and the incorporation of viral envelope protein into the virion during the life cycle of BLV.

scholarly journals The Conserved Poxvirus L3 Virion Protein Is Required for Transcription of Vaccinia Virus Early Genes

2005 ◽  
Vol 79 (23) ◽  
pp. 14719-14729 ◽  
Author(s):  
Wolfgang Resch ◽  
Bernard Moss
Keyword(s):  
Vaccinia Virus ◽  
Promoter Sequence ◽  
Wild Type ◽  
Virion Protein ◽  
Reading Frame ◽  
Virus Particles ◽  
Mature Virus

ABSTRACT We provide the initial characterization of the product of the vaccinia virus L3L open reading frame (VACWR090), which is conserved in all sequenced members of the poxvirus family. The predicted polypeptide contains no motifs or other features that provided a clue to the role of the L3 protein, and no functional information was available regarding a homolog discovered in Plasmodium falciparum. The L3 protein was expressed following viral DNA replication, a finding consistent with a putative late promoter sequence, and was packaged as a non-membrane protein in mature virus particles. A recombinant virus, in which the L3L gene was regulated by the Escherichia coli lac operator/repressor system, had a conditional lethal phenotype. The virus replicated in the presence of inducer, but in its absence, the yields of infectious virus were reduced by 99%. When cells were infected without inducer, however, no defect in gene expression or morphogenesis was noted. Virus particles lacking L3, which assembled in the absence of inducer, were indistinguishable from wild-type virions with regard to morphology, major structural proteins, and DNA content but were noninfectious. L3-deficient virions were able to bind and penetrate cells but produced extremely small amounts of viral early mRNA. A defect in transcription was demonstrated by in vitro studies with permeabilized virions, but soluble extracts of L3-deficient virions showed normal levels of template-dependent transcriptional activity, indicating that only transcription of the packaged genome is impaired.

scholarly journals Characterization of the VP39 envelope protein from Singapore grouper iridovirus

2015 ◽  
Vol 61 (12) ◽  
pp. 924-937 ◽  
Author(s):  
Honglian Zhang ◽  
Sheng Zhou ◽  
Liqun Xia ◽  
Xiaohong Huang ◽  
Youhua Huang ◽  
...  
Keyword(s):  
Envelope Protein ◽  
Core Gene ◽  
Immunofluorescence Assay ◽  
Current Data ◽  
Viral Envelope ◽  
Economic Losses ◽  
Immunogold Electron Microscopy ◽  
Viral Envelope Protein ◽  
Drug Inhibition

Singapore grouper iridovirus (SGIV) is a major pathogen that causes heavy economic losses to the grouper aquaculture industry in China and Southeast Asian countries. In the present study, a viral envelope protein, VP39, encoded by SGIV ORF39L, was identified and characterized. SGIV ORF39L was found in all sequenced iridoviruses and is now considered to be a core gene of the family Iridoviridae. ORF39L was classified as a late gene during in vitro infection using reverse transcription–polymerase chain reaction, western blotting, and a drug inhibition analysis. An indirect immunofluorescence assay revealed that the VP39 protein was confined to the cytoplasm, especially at viral assembly sites. Western blot and matrix-assisted laser desorption/ionization-time of flight tandem mass spectrometry analyses suggested that VP39 is an envelope protein. Immunogold electron microscopy further confirmed that VP39 is a viral envelope protein. Furthermore, a mouse anti-VP39 polyclonal antibody exhibited SGIV-neutralizing activity in vitro, suggesting that VP39 is involved in SGIV infection. Taken together, the current data suggest that VP39 represents a conserved envelope protein of iridoviruses that contributes to viral infection.

scholarly journals Mutations in the Vaccinia Virus A33R and B5R Envelope Proteins That Enhance Release of Extracellular Virions and Eliminate Formation of Actin-Containing Microvilli without Preventing Tyrosine Phosphorylation of the A36R Protein

2003 ◽  
Vol 77 (22) ◽  
pp. 12266-12275 ◽  
Author(s):  
Ehud Katz ◽  
Brian M. Ward ◽  
Andrea S. Weisberg ◽  
Bernard Moss
Keyword(s):  
Vaccinia Virus ◽  
Envelope Protein ◽  
Actin Polymerization ◽  
Single Amino Acid ◽  
Wild Type Virus ◽  
Virus Release ◽  
Type Virus ◽  
Wild Type ◽  
Extracellular Virus

ABSTRACT The spread of vaccinia virus in cell cultures is mediated by virions that adhere to the tips of specialized actin-containing microvilli and also by virions that are released into the medium. The use of a small plaque-forming A36R gene deletion mutant to select spontaneous second-site mutants exhibiting enhanced virus release was described previously. Two types of mutations were found: C-terminal truncations of the A33R envelope protein and a single amino acid substitution of the B5R envelope protein. In the present study, we transferred each type of mutation into a wild-type virus background in order to study their effects in vitro and in vivo. The two new mutants conserved the enhanced virus release properties of the original isolates; the A33R mutant produced considerably more extracellular virus than the B5R mutant. The extracellular virus particles contained the truncated A33R protein in one case and the mutated B5R protein in the other. Remarkably, both mutants failed to form actin tails and specialized microvilli, despite the presence of an intact A36R gene. The synthesis of the A36R protein as well as its physical association with the mutated or wild-type A33R protein was demonstrated. Moreover, the A36R protein was tyrosine phosphorylated, a step mediated by a membrane-associated Src kinase that regulates the nucleation of actin polymerization. The presence of large numbers of adherent virions on the cell surface argued against rapid dissociation as having a key role in preventing actin tail formation. Thus, the A33R and B5R proteins may be more directly involved in the formation or stabilization of actin tails than had been previously thought. When mice were inoculated intranasally, the A33R mutant was highly attenuated and the B5R mutant was mildly attenuated compared to wild-type virus. Enhanced virus release, therefore, did not compensate for the loss of actin tails and specialized microvilli.

scholarly journals Efficient Inhibition of Hepatitis B Virus Infection by Acylated Peptides Derived from the Large Viral Surface Protein

2005 ◽  
Vol 79 (3) ◽  
pp. 1613-1622 ◽  
Author(s):  
Philippe Gripon ◽  
Isabelle Cannie ◽  
Stephan Urban
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Envelope Protein ◽  
Viral Envelope ◽  
Hbv Infection ◽  
Viral Envelope Protein ◽  
Therapeutic Concept ◽  
Heparg Cell ◽  
B Virus

ABSTRACT 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.

scholarly journals Antiviral Peptides Targeting the West Nile Virus Envelope Protein

2006 ◽  
Vol 81 (4) ◽  
pp. 2047-2055 ◽  
Author(s):  
Fengwei Bai ◽  
Terrence Town ◽  
Deepti Pradhan ◽  
Jonathan Cox ◽  
Ashish ◽  
...  
Keyword(s):  
West Nile Virus ◽  
Envelope Protein ◽  
Phage Display Library ◽  
Viral Envelope ◽  
Host Cells ◽  
Viral Envelope Protein ◽  
Virus Envelope ◽  
West Nile ◽  
Inhibition Concentration

ABSTRACT West Nile virus (WNV) can cause fatal murine and human encephalitis. The viral envelope protein interacts with host cells. A murine brain cDNA phage display library was therefore probed with WNV envelope protein, resulting in the identification of several adherent peptides. Of these, peptide 1 prevented WNV infection in vitro with a 50% inhibition concentration of 67 μM and also inhibited infection of a related flavivirus, dengue virus. Peptide 9, a derivative of peptide 1, was a particularly potent inhibitor of WNV in vitro, with a 50% inhibition concentration of 2.6 μM. Moreover, mice challenged with WNV that had been incubated with peptide 9 had reduced viremia and fatality compared with control animals. Peptide 9 penetrated the murine blood-brain barrier and was found in the brain parenchyma, implying that it may have antiviral activity in the central nervous system. These short peptides serve as the basis for developing new therapeutics for West Nile encephalitis and, potentially, other flaviviruses.

scholarly journals Inhibition of p53 DNA Binding Function by the MDM2 Protein Acidic Domain

2011 ◽  
Vol 286 (18) ◽  
pp. 16018-16029 ◽  
Author(s):  
Brittany Cross ◽  
Lihong Chen ◽  
Qian Cheng ◽  
Baozong Li ◽  
Zhi-Min Yuan ◽  
...  
Keyword(s):  
Dna Binding ◽  
Binding Activity ◽  
Wild Type ◽  
Methyl Transferase ◽  
Reading Frame ◽  
Acidic Domain ◽  
Dna Binding Activity ◽  
Binding Function

MDM2 regulates p53 predominantly by promoting p53 ubiquitination. However, ubiquitination-independent mechanisms of MDM2 have also been implicated. Here we show that MDM2 inhibits p53 DNA binding activity in vitro and in vivo. MDM2 binding promotes p53 to adopt a mutant-like conformation, losing reactivity to antibody Pab1620, while exposing the Pab240 epitope. The acidic domain of MDM2 is required to induce p53 conformational change and inhibit p53 DNA binding. Alternate reading frame binding to the MDM2 acidic domain restores p53 wild type conformation and rescues DNA binding activity. Furthermore, histone methyl transferase SUV39H1 binding to the MDM2 acidic domain also restores p53 wild type conformation and allows p53-MDM2-SUV39H1 complex to bind DNA. These results provide further evidence for an ubiquitination-independent mechanism of p53 regulation by MDM2 and reveal how MDM2-interacting repressors gain access to p53 target promoters and repress transcription. Furthermore, we show that the MDM2 inhibitor Nutlin cooperates with the proteasome inhibitor Bortezomib by stimulating p53 DNA binding and transcriptional activity, providing a rationale for combination therapy using proteasome and MDM2 inhibitors.

scholarly journals Sinapic Acid Suppresses SARS CoV-2 Replication by Targeting Its Envelope Protein

Antibiotics ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 420
Author(s):  
Raha Orfali ◽  
Mostafa E. Rateb ◽  
Hossam M. Hassan ◽  
Mona Alonazi ◽  
Mokhtar R. Gomaa ◽  
...  
Keyword(s):  
Antiviral Activity ◽  
Envelope Protein ◽  
Sinapic Acid ◽  
Structural Features ◽  
Viral Envelope ◽  
Structural Motif ◽  
Protective Measure ◽  
Viral Envelope Protein ◽  
E Protein

SARS CoV-2 is still considered a global health issue, and its threat keeps growing with the emergence of newly evolved strains. Despite the success in developing some vaccines as a protective measure, finding cost-effective treatments is urgent. Accordingly, we screened a number of phenolic natural compounds for their in vitro anti-SARS CoV-2 activity. We found sinapic acid (SA) selectively inhibited the viral replication in vitro with an half-maximal inhibitory concentration (IC50) value of 2.69 µg/mL with significantly low cytotoxicity (CC50 = 189.3 µg/mL). Subsequently, we virtually screened all currently available molecular targets using a multistep in silico protocol to find out the most probable molecular target that mediates this compound’s antiviral activity. As a result, the viral envelope protein (E-protein) was suggested as the most possible hit for SA. Further in-depth molecular dynamic simulation-based investigation revealed the essential structural features of SA antiviral activity and its binding mode with E-protein. The structural and experimental results presented in this study strongly recommend SA as a promising structural motif for anti-SARS CoV-2 agent development.

scholarly journals Vaccinia Virus Envelope H3L Protein Binds to Cell Surface Heparan Sulfate and Is Important for Intracellular Mature Virion Morphogenesis and Virus Infection In Vitro and In Vivo

2000 ◽  
Vol 74 (7) ◽  
pp. 3353-3365 ◽  
Author(s):  
Chi-Long Lin ◽  
Che-Sheng Chung ◽  
Hans G. Heine ◽  
Wen Chang
Keyword(s):  
Cell Surface ◽  
Vaccinia Virus ◽  
Heparan Sulfate ◽  
Mutant Virus ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Virion Morphogenesis

ABSTRACT An immunodominant antigen, p35, is expressed on the envelope of intracellular mature virions (IMV) of vaccinia virus. p35 is encoded by the viral late gene H3L, but its role in the virus life cycle is not known. This report demonstrates that soluble H3L protein binds to heparan sulfate on the cell surface and competes with the binding of vaccinia virus, indicating a role for H3L protein in IMV adsorption to mammalian cells. A mutant virus defective in expression of H3L (H3L−) was constructed; the mutant virus has a small plaque phenotype and 10-fold lower IMV and extracellular enveloped virion titers than the wild-type virus. Virion morphogenesis is severely blocked and intermediate viral structures such as viral factories and crescents accumulate in cells infected with the H3L− mutant virus. IMV from the H3L− mutant virus are somewhat altered and less infectious than wild-type virions. However, cells infected by the mutant virus form multinucleated syncytia after low pH treatment, suggesting that H3L protein is not required for cell fusion. Mice inoculated intranasally with wild-type virus show high mortality and severe weight loss, whereas mice infected with H3L− mutant virus survive and recover faster, indicating that inactivation of the H3L gene attenuates virus virulence in vivo. In summary, these data indicate that H3L protein mediates vaccinia virus adsorption to cell surface heparan sulfate and is important for vaccinia virus infection in vitro and in vivo. In addition, H3L protein plays a role in virion assembly.

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