Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The sequence arrangement within the nontranscribed portion of the inverted terminal repetition of the vaccinia virus genome exists in quasi-stable and unstable forms that are not distinguishable on the basis of viral infectivity. The unstable forms, which composed about 20% of a serially passaged stock of virus, were recognized by terminal heterogeneity on restriction endonuclease analysis. Instead of a single terminal fragment from each end of the genome, an array of eight or more fragments differing in size by 1650-base-pair increments was detected. This feature was not eliminated by repeated plaque purification, indicating that the population of DNA molecules with various numbers of reiterations can rapidly evolve from the DNA of a single virus particle. However, at each successive round of plaque purification, about 20% of the unstable isolates revert back to the more stable form. Stable forms are characterized by the presence of a set of 13-17 tandem 70-base-pair repeats on each side of a 435-base-pair intervening sequence near both ends of the genome. In contrast, the unstable forms possess sets of tandem repeats and intervening sequences that alternate many times in series. The transition between the two genomic forms and the evolution of the unstable form appear to be mediated by recombinational events.
 ...
PMID:Instability and reiteration of DNA sequences within the vaccinia virus genome. 626 19

Deletions contained within the genomes of unstable and stable variants of vaccinia virus (strain WR) were analyzed. Restriction endonuclease mapping and hybridization to specific 32P-labeled DNA probes indicated that more than 6 X 10(6) daltons of DNA were deleted from the variants. In each case, the deletion occurred on the left side of the genome and started very close to the junction of the inverted terminal repetition and unique sequence. Both variants also contained a new SstI side on the right side of the genome. Hybridization selection and cell-free translation experiments indicated that these variants lost the ability to synthesize at least eight early mRNA's mapping within the deleted region. Although the deleted DNA was not essential for replication of the WR strain of vaccinia virus under laboratory conditions of infection, it presumably has a defined role under other circumstances. This conclusion was based on the conservation within the Elstree strain of vaccinia, the Utrecht strain of rabbitpox, and the Brighton strain of cowpox virus of sequences homologous to the deleted DNA. Moreover, mRNA's that hybridized to the deleted vaccinia virus DNA segment and encoded similar size polypeptides were made in cells infected with rabbitpox and cowpox viruses.
 ...
PMID:Deletion of a 9,000-base-pair segment of the vaccinia virus genome that encodes nonessential polypeptides. 627 95

The effect of Shope fibroma virus (SFV) infection on host DNA synthesis was investigated. The cytocidal strain, SFV-I, inhibited the incorporation of [3H]thymidine into nuclear DNA very shortly (2 h) after infection, whereas the noncytocidal strain, SFV-W, did so later (10 h postinfection) and to a lesser extent. Furthermore, a two- to threefold stimulation of host DNA synthesis was recorded in SFV-W-infected cells 3 to 4 h after infection. Since virion-associated nucleases have been implicated in the shutoff of host synthesis, these and other enzymatic activities were measured in purified virion preparations. The SFV strains and vaccinia virus contained equivalent amounts of DNA-dependent RNA polymerase, ATPase, and protein kinase activities. However, in SFV-W the pH 4.5 exonuclease activity was lower than in SFV-I and vaccinia virus, and the level of pH 7.8 endonuclease was almost undetectable. To test whether the lack of endonucleolytic activity had some effect on the removal of the cross-links in the parental DNA that occurs after viral penetration, the fate of the virion SFV DNA was followed. The majority (80%) of the SFV-I and SFV-W DNA molecules extracted after viral adsorption sedimented in alkaline sucrose gradients as cross-linked. After 3 h of infection, 75% of the SFV-I DNA molecules lacked cross-links, whereas 78% of the SFV-W DNA still remained cross-linked. The same results were obtained when the presence of cross-links was tested in restriction fragments. Taken together, these results indicate that virion-associated nucleases are involved in the early shutoff of host DNA synthesis and in the elimination of cross-links from the parental viral DNA.
 ...
PMID:Shope fibroma virus. II. Role of the virion-associated nucleases. 628 6

The 180,000 base pair (bp), covalently closed, linear duplex DNA genome of vaccinia virus contains a 10,000 bp inverted terminal repetition within which are one set of 13 and one set of 18 tandem 70 bp repeating units. A 967 bp segment containing the innermost 70 bp repeat and an adjacent region notable for a scarcity of restriction endonuclease sites has been sequenced. This was facilitated by the cloning of TaqI and partial TaqI fragments in pBR322. We found that the innermost 70 bp repeat overlaps one of two adjacent 125 bp repeats, following which are eight repeats of 54 bp, parts of 54 bp and 70 bp repeats, and four consecutive 6 to 7 bp repeats. The 70, 125, and 54 bp repeating units have extensive sequence homologies and redundancies that suggest evolution by unequal crossing over. Schemes whereby unequal crossovers of 54 bp repeats lead to a recombinant segment 86% homologous to the 125 bp repeat and unequal crossovers of 125 bp repeats lead to a recombinant segment 94% homologous to the 70 bp repeat were considered. This propensity for sequence divergence should provide a useful marker for comparing the relatedness of poxviruses.
 ...
PMID:Sequence homologies of diverse length tandem repetitions near ends of vaccinia virus genome suggest unequal crossing over. 629 46

DNA from several independent strains of Shope fibroma virus, a tumorogenic leporipoxvirus of rabbits, was isolated and analyzed by restriction endonuclease digestion and Southern blotting. The restriction profiles indicated a high degree of sequence conservation among the isolates but blotting under standard stringencies revealed no detectable cross homology with a member of the orthopoxvirus group, vaccinia. The genome of the fibroma virus was calculated to be in excess of 160 kilobases and shown to possess two features analogous to the orthopoxvirus group: (1) the terminal restriction fragments possess covalently closed hairpin structures; and (2) the terminal sequences are present as inverted repeats of greater than 10 kilobases. The terminal 3.6 kilobase BamHI restriction fragment was cloned in pBR322 after removal of the hairpin structure with mung bean single strand-specific endonuclease and addition of BamHI linkers. SFV sequences within this terminal region were shown, using 32P SFV cloned terminal probe, to have none of the sequence heterogeneity characteristic of vaccinia DNA termini. The remaining 20 internal SFV BamHI restriction fragments were propagated in bacterial plasmids either as intact fragments, or after secondary digestion with HindIII, and together constitute the complete cloned SFV sequence library.
 ...
PMID:Physical characterization and molecular cloning of the Shope fibroma virus DNA genome. 631 37

The production and selection of infectious vaccinia virus recombinants expressing foreign genes was facilitated by the construction of plasmid vectors. These vectors contain all or part of the vaccinia virus thymidine kinase (TK) gene interrupted by multiple unique restriction endonuclease sites placed adjacent to the TK promoter or another promoter translocated within the TK gene. The insertion of a continuous coding sequence for a foreign protein at one of the unique restriction endonuclease sites juxtaposes the transcriptional start site of a vaccinia promoter and the translational start site of a foreign gene. After transfection of vaccinia virus-infected cells with such plasmids, homologous recombination occurs between the vaccinia virus sequences flanking the chimeric gene and the same sequences within the virus genome. Recombinants formed in this manner have the chimeric gene inserted within the body of the vaccinia virus TK gene under control of a vaccinia virus promoter. Since recombinants have an interrupted TK gene, they are selected on the basis of their TK- phenotype and then checked for the presence and expression of the foreign gene. Infectious recombinant viruses expressing the procaryotic enzyme chloramphenicol acetyltransferase were constructed to optimize the system. The absence of chloramphenicol acetyltransferase activity in uninfected cells or in cells infected with wild-type vaccinia virus and the availability of a sensitive and quantitative enzyme assay allowed an estimation of the relative strengths of various promoter constructs. The expression of chloramphenicol acetyltransferase was detected within 1 h after infection of cells with recombinant virus, reflecting the early nature of the promoters used.
 ...
PMID:General method for production and selection of infectious vaccinia virus recombinants expressing foreign genes. 632 70

Two genomic variants of vaccinia virus isolated from serially propagated stocks were used to demonstrate marker rescue. The smaller (S variant) virus contains a 6.3 megadalton (MDal) deletion of unique DNA sequences present in the 123-MDal larger (L variant) virus. The deletion was mapped at 6.85 MDal from the left terminus of the genome, just outside of the inverted terminal repetition. Rescue of the unique deleted DNA sequences by infectious S variant virus was obtained in CV-1 cells by using the calcium orthophosphate precipitation technique of intact or restriction endonuclease-treated L-variant DNA. Restriction fragments that overlapped the deletion allowed marker rescue, but restriction of the L-variant DNA within the unique deleted sequences gave negative results. Restriction endonuclease analysis of the DNA obtained from twice-plaque-purified recombinant virus derived from the rescue of overlap donor fragments gave a restriction pattern identical to that of L-variant virus, indicating that the donor DNA was inserted into the rescuing virus by double recombination. No amplification of the unique sequences was observed from intact L-variant DNA in the absence of infectious S-variant virus, suggesting that deproteinized vaccinia DNA is noninfectious and that the donor DNA was neither integrated into the host DNA nor present as an episomal structure. By using 1 microgram of intact L-variant DNA per CV-1 monolayer in a 6-cm Petri dish, approximately 1--5% of the plaques contained the L-variant genotype, and the dose--response curve was essentially linear from 0.1 to 2 microgram of DNA.
 ...
PMID:Molecular genetics of vaccinia virus: demonstration of marker rescue. 695 Nov 97

Cultures of murine Friend erythroleukemia (FL) cells, which are chronically infected with leukemia virus, were inoculated with vaccinia virus. The yield of vaccinia virus was determined by assaying plaque-forming units in mouse L2 cells, and the yield of leukemia virus was determined by measuring reverse transcriptase (RNA-dependent DNA nucleotidyltransferase) activity released into the culture fluid. Although no facilitation of one virus by the other was detected, persistently infected cultures were established. Electron microscopic examination revealed the presence of vaccinia and leukemia viruses in the same cell. The permanent lines of cells persistently infected with vaccinia were designated FLvac. Their morphology, growth rate, cloning efficiency, and ability to respond to the induction of erythrodifferentiation by treatment with dimethyl sulfoxide were not appreciably altered as compared to the parental FL cells. However, the persistently infected cells showed a marked decrease in tumorigenicity when assayed in DBA/2 mice. The infectious virus produced by FLvac cells and by L2 cells were indistinguishable as judged by restriction endonuclease patterns of virion DNA, structural proteins, and the activities of two virion-associated DNases. The yield of infectious vaccinia virus from FLvac cells generally declined after about 60 serial passages. Although some cell lines no longer yield infectious virus, they are resistant to challenge with vaccinia at concentrations that are cytolytic for L2 cells. The mechanism responsible for the establishment of the persistent infection remains unclear because defective particles, interferon production, and temperature-sensitive mutants have not been detected.
 ...
PMID:Persistent infection of Friend erythroleukemia cells with vaccinia virus. 695 93

The large differences between the G+C content of the orf virus genome and those of other characterized poxviruses have precluded the use of DNA hybridization to establish a gene map of orf virus. Here we have sequenced the ends of cloned restriction endonuclease fragments of the nZ2 strain of orf virus (OV) and used the translated sequences to search protein data bases. Sequence from 15 points found high-scoring matches to data base entries, including 18 vaccinia virus (VAC) genes. We also present 2 kb of sequence from a region near the right terminus of the OV genome and show that it encodes homologs of VAC genes, F9L and F10L. The data presented here in conjunction with published and as yet unpublished data have allowed the construction of a gene map of OV on which 37 genes have been placed. Thirty-two of these genes have homologs in VAC. Alignment of the OV gene map with that of VAC revealed that each OV gene and its VAC counterpart occurred in the same order and orientation on their respective genomes. The intervals between many of the points of sequence were also found to be strikingly similar. The conserved spacing of genes between OV and VAC within the central 88.2 kb of the 139-kb OV genome is not maintained in the termini where insertion, deletion, and translocation have occurred. Parallels are drawn between the data presented here and related data from swinepox virus and capripox virus.
 ...
PMID:The establishment of a genetic map of orf virus reveals a pattern of genomic organization that is highly conserved among divergent poxviruses. 757 39

Influenza virus polymerase complexes that were expressed in the absence of genomic viral RNA and nucleoprotein were examined for endonuclease activity and transcriptase ability in vitro. Nuclear extracts of cells that express influenza virus polymerase through recombinant vaccinia virus infection did not display specific endonuclease activity in vitro. This polymerase presumably represents an early form of enzyme present in infected cells prior to ribonucleoprotein assembly. Upon addition of a virus-like model RNA template, containing the partially complementary sequence found at the ends of viral RNA, endonuclease activity is stimulated in a concentration-dependent and sequence-specific manner. Once stimulated, the polymerase is able to elongate from the added viral template. Thus, addition of viral template is required for polymerase activity, while the presence of nucleoprotein is not required for limited transcription. Also, full activation of this recombinant viral polymerase is dependent on the presence of both the 3' and 5' ends of the viral genome, as model RNA containing either end alone could not effectively trigger the endonuclease.
 ...
PMID:Recombinant influenza virus polymerase: requirement of both 5' and 3' viral ends for endonuclease activity. 810 13


<< Previous 1 2 3 4 5 6 Next >>