Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bursal lymphomas induced in chickens by avian leukosis viruses (ALVs) harbor proviral insertions that augment expression of an adjacent cellular oncogene, c-myc. To analyze such insertionally mutagenized c-myc genes in greater detail, we isolated molecular clones from two independent tumors. Precise proviral integration has occurred within the transcribed region of the c-myc gene in both mutant alleles. The proviruses bear different internal deletions that preclude the expression of the gag, pol, and env genes. The c-myc gene from bursal lymphoma LL4 contains a single copy of an ALV long terminal repeat (LTR), presumably the product of homologous recombination between LTRs at the ends of a normal provirus; the "solo" LTR is positioned in the correct orientation to act as a promoter for the c-myc gene. Bursal lymphoma LL3 contains an ALV provirus positioned upstream in the opposite transcriptional orientation to the coding exons of c-myc and deleted from a site within the leader region into the gag gene. In addition, the nucleotide sequence of the c-myc gene from tumor LL3 differs from the published sequence of the normal c-myc coding region at 3 positions of 180 determined. One of these changes, a silent nucleotide transition, is documented as a somatic mutation by restriction endonuclease mapping. It is flanked by two other candidate tumor-specific point mutations, one of which predicts an amino acid replacement, Pro----Thr at position 63. Thus, additional lesions that may affect the expression of viral genes and the quantity and nature of the putative c-myc gene product occur in provirally mutated c-myc alleles and may contribute to tumor progression.
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PMID:Proviral deletions and oncogene base-substitutions in insertionally mutagenized c-myc alleles may contribute to the progression of avian bursal tumors. 632 73

By the use of recombinant DNA technology and microinjection in cultured cells, the molecular genetic elements involved in the evolution of a retrovirus with the multipotential to infect, transform and replicate in host cell, have been critically examined in this investigation. Recently we have identified and purified the integrated and proviral DNA sequences specific for two rat endogenous helper leukemia viruses, WR- RaLV , originated from a chemically induced wild rat fibrosarcoma, and RHHV , isolated from a chemically induced rat hepatoma, HTC-H1 (1). By using a multidisciplinary approach combining restriction endonuclease analysis, reverse phase V-column chromatography, agarose gel electrophoresis, Southern blot transfer and filter nucleic acid hybridization, we were able to demonstrate that the rat helper leukemia viral DNA sequence was approximately 8.4-8.8 kb. The 8.8 kb RHHV DNA was molecularly cloned via the EK-1 certified vector pBR 322 plasmid into E. coli RRI cells. A successful recombinant clone, 8/32, that carried one entire RHHV 8.8 kb DNA sequence was mapped by restriction endonuclease analyses. Restricted DNA fragments of various sizes throughout the complete RHHV genome were isolated and purified for intranuclear microinjection into normal rat kidney cells. Release of type C infectious helper virus in these microinjected cells was investigated by superinfection on K-NRK, Kirsten sarcoma transformed non-producer cells. Recombination of the helper viral DNA sequence, en toto or of subgenomic sizes, carried in microinjected cells, with the sarcomagenic DNA sequence, carried in K-NRK cells, was also studied by genome-rescue and cell-transformation experiments. Our observations led to the conclusion that all critical genetic elements including the 5' LTR helper DNA sequence, gag, pol, and env genes, encoded for the biological activity of the type C helper virus resided within the 6.0 kb proximal to the 5' terminus of the endogenous rat type C helper virus DNA. They proved vitally essential for the recombination with the Src sequence during the evolution of an infectious, transforming and replication-competent retrovirus.
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PMID:Cloning of the rat endogenous helper leukemia virus DNA sequence and expression of the helper activity encoded by the cloned DNA sequence in normal rat kidney cells by microinjection. 632 7

Mag is a retrotransposon found as an insert in the Sericin 2 gene. It is present in a few copies--4 to 15--dispersed in the genome of different strains of Bombyx mori as well as in Bombyx mandarina. Flanked by a 5 bp target sequence with no sequence specificity, it is bordered by direct repeats of 77 nucleotides. Despite their unusual short size, these terminal repeats and their immediately adjacent sequences present all the signals necessary for transcription into genomic RNA and for reverse transcription. Mag contains two overlapping open reading frames which are organized as the gag and pol genes of retroviruses and encode putative nucleic acid binding peptide, protease, reverse transcriptase, RNase H and endonuclease in this order. Sequence comparison of these proteins places mag within the gypsy group of LTR retrotransposons next to the echinoderm element SURL.
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PMID:Structural features of mag, a gypsy-like retrotransposon of Bombyx mori, with unusual short terminal repeats. 781 9

A mutant of human immunodeficiency virus type 1 which carries a frameshift insertion in the integrase/endonuclease region of pol gene was constructed in vitro. Upon transfection into cells, although this mutant exhibited a normal phenotype with respect to expression of gag, pol, and env genes and to generation of progeny virions, no replication-competent virus in CD4-positive cells emerged. An assay for the single-step replication of a defective viral genome dependent on trans complementation by rev protein was established and used to monitor the early phase of viral infection process. Viral clones with a mutation in the vif, vpr, or vpu gene displayed no abnormality in the early phase. In contrast, the integrase mutant did not direct a marker gene expression after infection. Together with an observation that the mutant lacked the ability to integrate, these results indicated that the integration was required for efficient viral gene expression and productive infection of human immunodeficiency virus type 1.
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PMID:Integration is essential for efficient gene expression of human immunodeficiency virus type 1. 843 8

A repeated DNA sequence used for epidemiological studies of the human opportunistic pathogen Aspergillus fumigatus has been characterized. It is a retroelement of 6914 bp in length, bounded by long terminal repeats of 282 bp, with sequence and features characteristic of retroviruses and retrotransposons. A 5 bp duplication site was found at its borders. This element, designated Afut1, encodes amino acid sequences homologous to the reverse transcriptase, RNase H and endonuclease encoded by the pol genes of retroelements. Comparison of the peptidic sequences with other putative polypeptides of fungal LTR retrotransposons showed that Afut1 is a member of the gypsy group. This is the first report of a transposable element in A.fumigatus. Afut1 is a defective element: the putative coding domains contain multiple stop codons due exclusively to transitions from C:G to T:A.
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PMID:Afut1, a retrotransposon-like element from Aspergillus fumigatus. 862 74

One, two or four copies of the 'helix-hairpin-helix' (HhH) DNA-binding motif are predicted to occur in 14 homologous families of proteins. The predicted DNA-binding function of this motif is shown to be consistent with the crystallographic structure of rat polymerase beta, complexed with DNA template-primer [Pelletier, H., Sawaya, M.R., Kumar, A., Wilson, S.H. and Kraut, J. (1994) Science 264, 1891-1903] and with biochemical data. Five crystal structures of predicted HhH motifs are currently known: two from rat pol beta and one each in endonuclease III, AlkA and the 5' nuclease domain of Taq pol I. These motifs are more structurally similar to each other than to any other structure in current databases, including helix-turn-helix motifs. The clustering of the five HhH structures separately from other bi-helical structures in searches indicates that all members of the 14 families of proteins described herein possess similar HhH structures. By analogy with the rat pol beta structure, it is suggested that each of these HhH motifs bind DNA in a non-sequence-specific manner, via the formation of hydrogen bonds between protein backbone nitrogens and DNA phosphate groups. This type of interaction contrasts with the sequence-specific interactions of other motifs, including helix-turn-helix structures. Additional evidence is provided that alphaherpesvirus virion host shutoff proteins are members of the polymerase I 5'-nuclease and FEN1-like endonuclease gene family, and that a novel HhH-containing DNA-binding domain occurs in the kinesin-like molecule nod, and in other proteins such as cnjB, emb-5 and SPT6.
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PMID:The helix-hairpin-helix DNA-binding motif: a structural basis for non-sequence-specific recognition of DNA. 869 86

The simultaneous presence of multiple HIV-1 subtypes has become common in communities with the growth of the pandemic. As a consequence, the potentiality for an increased frequency of HIV-1 mixed infections caused by viruses of distinct subtypes could be expected. Thus, there is a need to estimate the prevalence and geographic distribution of infections caused by viruses of a singular subtype as well as coinfections caused by two or more HIV-1 strains of distinct subtypes. To address this need, we have developed a genetic method based on restriction fragment length polymorphism (RFLP) to screen for these two types of infections within infected populations. In this assay, restriction enzymes may be used to predict the phylogroup of HIV-1 infected samples. A 297 bp pol fragment spanning the entire viral protease gene and a 311 bp fragment of the p24 gag region are used for this analysis. The viral regions are amplified by nested PCR using DNA templates from uncultured peripheral blood mononuclear cells (PBMC) or virus culture. Classification of HIV-1 strains to well defined subtypes B, D, F, and A/C is done by sequential endonuclease restriction analysis of a PCR amplified-protease gene followed by analysis of the p24 gag region. The electrophoretic migration patterns visualized by ethidium bromide staining or by radiolabeled probes are then determined on a 10% polyacrylamide gel. In infections caused by viruses of a singular subtype, a single restriction pattern is detected, whereas in multiple infections caused by two or more viral strains of different subtypes, the combination of different digestion patterns are observed in infected individuals. Using this methodology we have screened for genetic variations in HIV-1 proviral DNA from thirty-three Brazilian samples. Our RFLP procedure classified thirty-two samples as single infections caused by viruses of subtypes B (31) and F (1), and one sample as dual infection caused by distinct viral strains. Subsequent sequence and phylogenetic analysis of the viral protease gene in lymphocytes of all these patients confirmed our RFLP findings in single infections, and demonstrated the existence of two distinct HIV-1 strains of subtypes F and D in a patient which lymphocytes showed the simultaneous presence of two different digestion patterns. As up to now, single infections caused by subtype D variants were not identified in Brazil, our data provide the first evidence of subtype D HIV-1 in this country. Because sequencing of HIV proviral DNA is not particularly practical for large-scale molecular epidemiological studies, the protease/gag-based RFLP screening method will be useful to predict the phylogroup of HIV-1, and to identify multiple infections caused by HIV-1 strains of distinct subtypes. We believe that this information is crucial for both evaluation of the HIV-1/AIDS pandemic and intervention strategies.
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PMID:Identification of single and dual infections with distinct subtypes of human immunodeficiency virus type 1 by using restriction fragment length polymorphism analysis. 893 82

Chicken repeat 1 (CR1) elements comprise a family of non-long terminal repeat (LTR) retrotransposons that have several noteworthy features. For example, whereas most other non-LTR elements have poly(A) tracts or other simple A-rich repeats at their 3' ends, the 3' ends of CR1 elements conform to the consensus [(CATTCTRT)(GATTCTRT)1-3]. CR1 elements also display an unusual bias for severe 5' truncations: only approx. 30 (out of a total of approx. 30 000) CR1 elements in the chicken genome include significant portions of the pol-like open reading frame (ORF) that we previously identified and partially sequenced [Burch et al. (1993) Proc. Natl. Acad. Sci. USA 90, 8199-8203]. In the present study we derived a consensus sequence for this entire ORF (ORF2) as well as an upstream ORF (ORF1) and part of a 5' untranslated region (UTR). The conceptual translation product of ORF2 is predicted to contain an endonuclease domain in addition to a reverse transcriptase domain. These results suggest that CR1 elements retrotranspose using a "nick and prime" mechanism similar (but not identical) to other families of non-LTR elements.
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PMID:Chicken repeat 1 (CR1) elements, which define an ancient family of vertebrate non-LTR retrotransposons, contain two closely spaced open reading frames. 933 79

DNA polymerase beta (beta-pol) cleaves the sugar-phosphate bond 3' to an intact apurinic/apyrimidinic (AP) site (i.e. AP lyase activity). The same bond is cleaved even if the AP site has been previously 5'-incised by AP endonuclease, resulting in a 5' 2-deoxyribose 5-phosphate (i.e. dRP lyase activity). We characterized these lyase reactions by steady-state kinetics with the amino-terminal 8-kDa domain of beta-pol and with the entire 39-kDa polymerase. Steady-state kinetic analyses show that the Michaelis constants for both the dRP and AP lyase activities of beta-pol are similar. However, kcat is approximately 200-fold lower for the AP lyase activity on an intact AP site than for an AP endonuclease-preincised site. The 8-kDa domain was also less efficient with an intact AP site than on a preincised site. The full-length enzyme and the 8-kDa domain efficiently remove the 5' dRP from a preincised AP site in the absence of Mg2+, and the pH profiles of beta-pol and 8-kDa domain dRP lyase catalytic efficiency exhibit a broad alkaline pH optimum. An inhibitory effect of pyridoxal 5'-phosphate on the dRP lyase activity is consistent with involvement of a primary amine (Lys72) as the Schiff base nucleophile during lyase chemistry.
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PMID:Human DNA polymerase beta deoxyribose phosphate lyase. Substrate specificity and catalytic mechanism. 961 42

A repeated DNA sequence in the genome of Neurospora crassa has been identified as a family of degenerate retroelements. Retroelements encode protein sequences with clear homology to the reverse transcriptase, RNase H and endonuclease products of the pol genes common to retroviruses and retrotransposons. These sequence comparisons place the N. crassa element within the gypsy group of retrotransposons, akin to other elements found in filamentous fungi. However, the Neurospora element is defective, as no flanking long terminal repeats (LTRs) could be distinguished and the pol gene homologues contain numerous stop codons as a result of multiple base substitutions. The base composition of the element displays significant under-representation of the dinucleotide CpA, the preferred target site of repeat-induced point mutation (RIP). The genomic sequences exhibit G:C to A:T transitions between copies which are diagnostic of RIP. The degenerate retroelement has accordingly been designated by the acronym dab-1 (dead and buried).
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PMID:DAB1: a degenerate retrotransposon-like element from Neurospora crassa. 964 50


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