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Antiviral Research Oct 2012New polyomaviruses are continually being identified, and it is likely that links between this virus family and disease will continue to emerge. Unfortunately, a specific...
New polyomaviruses are continually being identified, and it is likely that links between this virus family and disease will continue to emerge. Unfortunately, a specific treatment for polyomavirus-associated disease is lacking. Because polyomaviruses express large Tumor Antigen, TAg, we hypothesized that small molecule inhibitors of the essential ATPase activity of TAg would inhibit viral replication. Using a new screening platform, we identified inhibitors of TAg's ATPase activity. Lead compounds were moved into a secondary assay, and ultimately two FDA approved compounds, bithionol and hexachlorophene, were identified as the most potent TAg inhibitors known to date. Both compounds inhibited Simian Virus 40 replication as assessed by plaque assay and quantitative PCR. Moreover, these compounds inhibited BK virus, which causes BKV Associated Nephropathy. In neither case was host cell viability compromised at these concentrations. Our data indicate that directed screening for TAg inhibitors is a viable method to identify polyomavirus inhibitors, and that bithionol and hexachlorophene represent lead compounds that may be further modified and/or ultimately used to combat diseases associated with polyomavirus infection.
Topics: Adenosine Triphosphatases; Antigens, Viral, Tumor; Antiviral Agents; BK Virus; Drug Evaluation, Preclinical; Enzyme Inhibitors; Humans; Real-Time Polymerase Chain Reaction; Simian virus 40; Viral Plaque Assay; Virus Replication
PubMed: 22898086
DOI: 10.1016/j.antiviral.2012.07.012 -
Journal of Virology Jun 1972Velocity sedimentation analysis of simian virus 40 degraded in alkaline buffers, pH 10.5, yields two components: soluble protein containing the largest polypeptides, VP1...
Velocity sedimentation analysis of simian virus 40 degraded in alkaline buffers, pH 10.5, yields two components: soluble protein containing the largest polypeptides, VP1 and VP2, of the virion, and a deoxynucleoprotein complex (DNP-I) containing the viral deoxyribonucleic acid (DNA) and the small polypeptides, VP4, 5, and 6, and all or part of VP3. Dissociation of DNP-I by equilibrium centrifugation in CsCl yields a complex (DNP-II) of the viral DNA and residual, tightly bound polypeptide; VP4, 5, and 6, but not VP3, are recovered after separation from DNP-II. Treatment of the virus with beta-mercaptoethanol and iodination experiments suggest that VP1 and VP2 might exist as compact structures cross-linked with disulfide bonds, perhaps forming the capsid. VP4, 5, and 6 form a relatively stable complex with the viral DNA and are supposed to be the internal proteins. The location of VP3 is not well defined; at least a portion of it is tightly bound to the viral DNA.
Topics: Acetates; Amino Acids; Animals; Autoradiography; Binding Sites; Carbon Isotopes; Cell Line; Centrifugation, Density Gradient; DNA, Viral; Electrophoresis, Disc; Haplorhini; Iodine; Kidney; Mercaptoethanol; Microscopy, Electron; Nucleoproteins; Peptides; Simian virus 40; Staining and Labeling; Thymidine; Tritium; Uranium; Viral Proteins
PubMed: 4113888
DOI: 10.1128/JVI.9.6.923-929.1972 -
ACS Nano Oct 2017Crystallization is a fundamental and ubiquitous process that is well understood in the case of atoms or small molecules, but its outcome is still hard to predict in the...
Crystallization is a fundamental and ubiquitous process that is well understood in the case of atoms or small molecules, but its outcome is still hard to predict in the case of nanoparticles or macromolecular complexes. Controlling the organization of virus nanoparticles into a variety of 3D supramolecular architectures is often done by multivalent ions and is of great interest for biomedical applications such as drug or gene delivery and biosensing, as well as for bionanomaterials and catalysis. In this paper, we show that slow dialysis, over several hours, of wild-type Simian Virus 40 (wt SV40) nanoparticle solution against salt solutions containing MgCl, with or without added NaCl, results in wt SV40 nanoparticles arranged in a body cubic center crystal structure with Im3m space group, as a thermodynamic product, in coexistence with soluble wt SV40 nanoparticles. The nanoparticle crystals formed above a critical MgCl concentrations. Reentrant melting and resolubilization of the virus nanoparticles took place when the MgCl concentrations passed a second threshold. Using synchrotron solution X-ray scattering we determined the structures and the mass fraction of the soluble and crystal phases as a function of MgCl and NaCl concentrations. A thermodynamic model, which balances the chemical potentials of the Mg ions in each of the possible states, explains our observations. The model reveals the mechanism of both the crystallization and the reentrant melting and resolubilization and shows that counterion entropy is the main driving force for both processes.
Topics: Crystallization; Nanoparticles; Simian virus 40; Solubility; Thermodynamics
PubMed: 28956913
DOI: 10.1021/acsnano.7b03131 -
Journal of Virology Sep 1982Simian virus 40 chromosomes were separated into various species by a two-step purification consisting of low-ionic-strength glycerol gradient sedimentation followed by...
Simian virus 40 chromosomes were separated into various species by a two-step purification consisting of low-ionic-strength glycerol gradient sedimentation followed by low-ionic-strength agarose gel electrophoresis. For each species of simian virus 40 chromosome purified, the comigrating DNA and proteins were identified by agarose or polyacrylamide gel electrophoresis, respectively. Two species of chromosomes were identified which contained form I and form II DNA and large amounts of viral protein; they migrated more slowly than most of the free simian virus 40 chromosomes, which contained very little viral protein. The nuclease susceptibility of these chromosomes suggests to us that they are intermediates in encapsidation, and we describe an encapsidation model.
Topics: Capsid; DNA, Viral; Deoxyribonucleases; Deoxyribonucleoproteins; Electrophoresis, Agar Gel; Morphogenesis; Simian virus 40; Viral Proteins
PubMed: 6292464
DOI: 10.1128/JVI.43.3.830-839.1982 -
Proceedings of the National Academy of... Mar 1979The primary nucleotide sequence of three regions of BK virus (BKV) variant (MM) DNA has been determined. The region between map positions 0.715 and 0.900 includes the... (Comparative Study)
Comparative Study
The primary nucleotide sequence of three regions of BK virus (BKV) variant (MM) DNA has been determined. The region between map positions 0.715 and 0.900 includes the initiation points and partial coding sequences of the putative VP2 and VP3 proteins of BKV(MM), the amino acid sequences of which show over 80% homology with those of VP2 and VP3 of simian virus 40. The sequence of a potential leader protein X, 66 amino acids long for BKV(MM) and 62 long for simian virus 40, is also deduced. The regions between 0.595 and 0.398 and 0.310 and 0.175 include the coding sequence for the entire small t antigen and most of the large T antigen of BKV(MM). The DNA sequence within these regions comprises over 50% of the complete BKV(MM) genome and shows a 70% sequence homology with the corresponding regions of simian virus 40 DNA. This high degree of homology is at variance with the reported homology values of 11--20% estimated by hybridization measurements of heteroduplex analyses. Possible explanations for the discrepancy are discussed.
Topics: BK Virus; Base Sequence; DNA, Viral; Genetic Code; Nucleic Acid Hybridization; Polyomavirus; Simian virus 40; Species Specificity
PubMed: 220609
DOI: 10.1073/pnas.76.3.1179 -
Proceedings of the National Academy of... Jul 1977The infection of confluent monkey cells with simian virus 40 induced the synthesis of both cellular DNA and histones. However, during the course of infection, cellular...
The infection of confluent monkey cells with simian virus 40 induced the synthesis of both cellular DNA and histones. However, during the course of infection, cellular histone synthesis was uncoupled from cellular DNA replication and became coupled to viral DNA replication. The synthesis of all five host histone species was induced after virus infection and they appeared to be more highly phosphorylated than their couterparts in uninfected cells. At late times after infection, the cells contained twice as much histones as did uninfected cells. All the histone species except H1 were incorporated into virions. Compared to cellular histones, virion histones were enriched in the arginine-rich species H3 and H4. Although both old and newly synthesized cellular histones were incorporated into virions, there were about 5 times more newly synthesized than old histone polypeptides in virions.
Topics: Cell Line; Contact Inhibition; DNA Replication; DNA, Viral; Histones; Kinetics; Phosphates; Simian virus 40; Virus Replication
PubMed: 197521
DOI: 10.1073/pnas.74.7.2805 -
Journal of Virology May 1976Viable mutants of simian virus 40 (SV40), with deletions ranging in size from 15 to 200 base pairs, have been obtained by infecting CV-1P cells with circularly permuted... (Comparative Study)
Comparative Study
Viable mutants of simian virus 40 (SV40), with deletions ranging in size from 15 to 200 base pairs, have been obtained by infecting CV-1P cells with circularly permuted linear SV40 DNA. The linear DNA was produced by cleavage of closed circular DNA with DNase I in the presence of Mn2+, followed, in some cases, by mild digestion with lambda 5'-exonuclease. The SV40 map location and the size of each deletion were determined by using the S1 nuclease mapping procedure (Shenk et al., 1975) and the change in size of fragments produced by Hind II + III endonuclease cleavage. Deletions in at least three regions of the SV40 chromosome have slight or no effect on the rate or yield of viral multiplication and on vira-induced cellular transformation. These regions are located at the following coordinates on the SV40 physical map: 0.17 to 0.18; 0.54 to 0.59; and 0.68 to 0.74.
Topics: Cell Line; Cell Transformation, Neoplastic; Chromosome Mapping; DNA Restriction Enzymes; DNA, Viral; Deoxyribonucleases; Endonucleases; Genetic Linkage; Haemophilus; Mutation; Simian virus 40; Virus Replication
PubMed: 178902
DOI: 10.1128/JVI.18.2.664-671.1976 -
Proceedings of the National Academy of... Feb 1975The transforming ability in 10% serum medium of the temperature-sensitive mutants of simian virus 40 in the complementation group III (ts640 type mutants) was greatly...
The transforming ability in 10% serum medium of the temperature-sensitive mutants of simian virus 40 in the complementation group III (ts640 type mutants) was greatly reduced when the infected rat 3Y1 cells were incubated at the restrictive temperature of 40 degrees or incubated first at 40 degrees for 3 days and then shifted to the permissive temperature of 33 degrees. Transformation did occur efficiently after incubation at 33 degrees or after an initial incubation at 33 degrees for 5 days followed by a shift to 40 degrees. When growth properties of 3Y1 cells transformed at 33 degrees by the group III mutants were examined at 40 degrees, several aspects of the transformed state were rendered temperature-sensitive. These aspects were the ability of cells to grow in low serum (1.5%) medium and to make colonies, in 10% serum medium, on monolayers of untransformed 3Y1 cells and in soft agar. It is concluded that a simian virus 40 gene (cistron III) controls the initiation, as well as at least some aspects of the maintenance, of transformation and that the initiation reaction is a more heat-labile event than the maintenance reaction(s) under the experimental conditions.
Topics: Animals; Cell Transformation, Neoplastic; Clone Cells; Embryo, Mammalian; Fibroblasts; Genes; Mutation; Rats; Simian virus 40; Temperature
PubMed: 164665
DOI: 10.1073/pnas.72.2.673 -
Applied Microbiology May 1972The immunoperoxidase technique was used in an electron microscopy study to localize the virions of herpes zoster virus and simian virus 40 in cell cultures. Intranuclear...
The immunoperoxidase technique was used in an electron microscopy study to localize the virions of herpes zoster virus and simian virus 40 in cell cultures. Intranuclear and intracytoplasmic virions of herpes zoster virus were easily and specifically identified due to intense staining by the finely granular, black reaction product. With simian virus 40, intranuclear virions were not stained, whereas intracytoplasmic particles appeared densely black. There was essentially no background staining. Advantages of this technique over the ferritin-labeled antibody method include simpler preparative procedures for reagents, greater penetrability of the antibody conjugate, and internal amplification which substantially improves the ability to localize sites of antigen-antibody reaction. We believe that the immunoperoxidase method can be successfully applied to a wide variety of problems involving viral antigens.
Topics: Animals; Antigen-Antibody Reactions; Antigens, Viral; Cell Line; Cell Nucleus; Cells, Cultured; Cytoplasm; Fibroblasts; Haplorhini; Herpes Zoster; Herpesviridae; Humans; Inclusion Bodies, Viral; Microscopy, Electron; Peroxidases; Plants, Edible; Simian virus 40; Staining and Labeling
PubMed: 4113252
DOI: 10.1128/am.23.5.1001-1009.1972 -
Structure (London, England : 1993) Feb 1996Simian virus 40 (SV40) and murine polyomavirus (polyoma) are non-enveloped DNA tumor viruses. Their structurally similar capsids, about 500 degrees in diameter, are...
BACKGROUND
Simian virus 40 (SV40) and murine polyomavirus (polyoma) are non-enveloped DNA tumor viruses. Their structurally similar capsids, about 500 degrees in diameter, are formed by 72 pentamers of the major coat protein VP1.
RESULTS
We describe in this paper the structure determination of SV40 and polyoma at 3.8 degree resolution, focusing particularly on methodological issues, and on a comparison of the overall molecular organization in the two related virus particles. Initial phases for SV40 were obtained by single isomorphous replacement at 6.5 degree. Phases were refined and the resolution extended to 3.8 degree by a combination of strict 5-fold and partial 30-fold electron-density averaging. The structure of polyoma was subsequently determined by systematically translating and rotating the individual VP1 pentamers, in order to find the maximum correlation between calculated and observed structure factors. The resolution was then extended to 3.8 degree, also by phase refinement through electron-density averaging.
CONCLUSION
The strategies for density averaging and for molecular replacement, used to determine the SV40 and polyoma structures, are likely to be generally useful. The individual building blocks, the VP1 pentamers, are essentially identical in both cases, as are the local details of their interactions with neighboring pentamers. Nevertheless, the arrangement of the pentamers with respect to each other is somewhat different in the two viruses. Whereas SV40 is almost spherical, with all pentamers at identical radii, the pentamers in polyoma that lie on icosahedral fivefold axes are displaced outward by about 5 degree.
Topics: Capsid; Capsid Proteins; Electrons; Polyomavirus; Protein Conformation; Simian virus 40
PubMed: 8805522
DOI: 10.1016/s0969-2126(96)00019-6