Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
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Enzymatic activity, hydrolyzing DNA treated with beta-isopropyl-bis-beta-chloroethylamine (HN2-DNA), HN2-DNA exposed at 50 degrees for 1 h, and DNA treated with acid, to acid-soluble fragments was found in extracts from cells of M. lysodeikticus. The endonucleolytic component ofthe indicated activity manifests chromatographic properties on DEAE- and CM-cellulose, close to those for UV-endonuclease. Activity is manifested by UV-irradiated DNA, proflavin, and cyanide. Two electrophoretically homogeneous fractions of UV-endonuclease (after chromatography on DEAE- and CM-cellulose), with molecular weights about 13,000 and 15,000 daltons, exhibit endonucleolytic activity with respect to HN2-DNA, exposed at 50 degrees for 1 h, and with respect to "acid" DNA, treated for 6 min at 70 degrees in citrate buffer, pH 3.5. The activity with respect to the latter substrate is competitively suppressed by UV-irradiated DNA. The most probable substrate of UV-endonuclease, in addition to cyclobutane dimers, is the depurinized region of DNA.
Mol Biol 1975 Jan
PMID:The presence of an endonuclease acting on UV-irradiated and depurinized DNA in cells of Micrococcus lysodeikticus. 112 5

A 1.45 kb DNA sequence encoding the rat alpha 6 GABAA receptor subunit (nucleotides 33-1483) was cloned from a Sprague-Dawley rat brain cDNA library by PCR amplification. Dideoxy sequencing of two individual clones revealed that the nucleotide sequence differed at only one basepair (T480-->G) from that published previously. This difference altered the deduced amino acid sequence, producing a conservative amino acid substitution (His121-->Gln). A Gln residue is present at the same location in the bovine alpha 6 subunit. Restriction endonuclease analysis of the total PCR product demonstrated that this variant of the rat alpha 6 subunit was the only allele found in this particular rat brain library, the original allele was not present. These results were further verified by RNAse protection assays performed with RNA isolated from individual rat cerebella. alpha 6, beta 1, and gamma 2S subunits were transiently expressed in L929 cells for electrophysiological analysis. Whole-cell recordings obtained from the cells demonstrated that GABAA receptor channels with the expected GABA and benzodiazepine pharmacology were produced. Excised outside out single channel recordings from the same cells revealed that GABA elicited brief duration openings to a 33 pS main conductance level and to at least one smaller (approximately 21 pS) subconductance level. Thus this allelic variant of rat alpha 6 subunit could assemble with other subunits to form a functional GABAA receptor channel with similar properties to the original allelic form.
Brain Res Mol Brain Res 1992 Nov
PMID:Molecular and electrophysiological characterization of a allelic variant of the rat alpha 6 GABAA receptor subunit. 128 Dec 55

It has been shown that retrons, retro-elements in bacteria, produce a reverse transcriptase (RT) and multicopy single-stranded DNA (msDNA) whose 5' end is covalently linked to RNA (msdRNA) by a 2'-5' phosphodiester bond. Here, I show that a retron in clinical Escherichia coli strain 161 produces an msDNA unlinked to RNA. The msDNA produced by this retron is a 79-nucleotide-long single-stranded DNA with monophosphate on its 5' terminus. When the retron in strain 161 is cloned into E. coli K-12, the majority of msDNA produced in the clone is the same as the msDNA in the clinical strain. However, in the K-12 clone, about 10% of the msDNA produced is present as a DNA covalently linked to RNA. The DNA part of this RNA-DNA compound is an 83 nucleotides long with the same sequence as the unbranched msDNA, except for the presence of four additional nucleotides at the 5' side. From the analysis of the RNA-DNA compound and the results of in vitro synthesis, I show that the primary product of reverse transcription in this retron is an 83-nucleotide-long DNA covalently linked to RNA. This RNA-DNA compound is further processed to the final product, the 79-nucleotide-long msDNA with a terminal 5' monophosphate, by an endonucleolytic cleavage between the fourth and fifth positions of the DNA component of the RNA-DNA compound. The minimum region required for the production of such msDNA free of RNA contains only genes known to be required for the synthesis of branched msDNA-RNA compound in other retrons (msd, msr and ret). This suggests that either the RT has an endonuclease activity or that the msDNA-RNA compound is autocatalytically processed.
Mol Microbiol 1992 Dec
PMID:Structure and biosynthesis of unbranched multicopy single-stranded DNA by reverse transcriptase in a clinical Escherichia coli isolate. 128 91

Abasic (AP) sites in DNA are produced spontaneously and by many genotoxic agents. The repair of such damages is initiated by AP endonucleases, which are evidently ubiquitous. We employed the recently cloned cDNA, APE, that encodes the major human AP endonuclease, to isolate large genomic fragments that contain the intact APE gene. The sequence of 3 kb encompassing APE was determined (GenBank Accession No. M99703). The APE gene contains four small introns (ranging 130 to 566 bp) and five exons, the first of which is untranslated. The 0.5 kb of DNA sequence upstream of APE did revealed only a possible CCAAT box, but no other regulatory sites or a TATA box, consistent with the constitutive expression of AP endonuclease activity observed in other studies. The location of APE in the human genome was mapped to chromosome 14, bands q11.2-12, by fluorescence in situ hybridization of metaphase cells with DNA from the genomic clones and subclones. Although this locus has not been associated causally with genetic diseases of DNA repair, some translocations that affect 14q11.2-12 could compromise APE and lead to genetic instability.
Hum Mol Genet 1992 Dec
PMID:Human apurinic endonuclease gene (APE): structure and genomic mapping (chromosome 14q11.2-12). 128 93

Nucleosome formation on inverted repeats or on some alternations of purines and pyrimidines can be inhibited in vitro by DNA supercoiling through their supercoiling-induced structural transitions to cruciforms or Z-form DNA, respectively. We report here, as a result of study of single nucleosome reconstitutions on a DNA minicircle, that a physiological level of DNA supercoiling can also enhance nucleosome sequence preference. The 357 base-pair minicircle was composed of a promoter of phage SP6 RNA polymerase joined to a 256 base-pair fragment containing a sea urchin 5 S RNA gene. Nucleosome formation on the promoter was found to be enhanced on a topoisomer with in vivo superhelix density when compared to topoisomers of lower or higher superhelical densities, to the nicked circle, or to the linear DNA. In contrast, nucleosomes at other positions appeared to be insensitive to supercoiling. This observation relied on a novel procedure for the investigation of nucleosome positioning. The reconstituted circular chromatin was first linearized using a restriction endonuclease, and the linear chromatin so obtained was electrophoresed as nucleoprotein in a polyacrylamide gel. The gel showed well-fractionated bands whose mobilities were a V-like function of nucleosome positions, with the nucleosome near the middle migrating less. This behavior is similar to that previously observed for complexes of sequence-specific DNA-bending proteins with circularly permuted DNA fragments, and presumably reflects the change in the direction of the DNA axis between the entrance and the exit of the particle. Possible mechanisms for such supercoiling-induced modulation of nucleosome formation are discussed in the light of the supercoiling-dependent susceptibility to cleavage of the naked minicircle with S1 and Bal31 nucleases; and a comparison between DNase I cleavage patterns of the modulated nucleosome and of another, non-modulated, overlapping nucleosome.
J Mol Biol 1992 Apr 20
PMID:Chromatin reconstitution on small DNA rings. IV. DNA supercoiling and nucleosome sequence preference. 131 7

For studies on molecular mechanisms of mutagenesis, it would be advantageous to transfer mutant genes with specific alterations into mammalian cells and use the transformed cells in reversion analyses. In the present paper, we describe an efficient method for analyzing reversion events occurring in cells that possess multiple copies of a mutational target gene. This method involves amplification of the chromosomally integrated target genes with the polymerase chain reaction (PCR) and restriction endonuclease digestion of the amplified product. Single reversion events that either create or destroy restriction endonuclease recognition sequences that encompass the site of the original mutation can be identified in a background of 10-20 copies of the gene that retain the mutant sequence. Using this method, we have analyzed revertants induced by 5-bromodeoxyuridine (BrdU) in a Chinese hamster ovary cell line that possesses multiple copies of a mutant bacterial gpt gene containing a specific alteration. The results of this study not only demonstrate the effectiveness of this method for analyzing reversion of a single gene copy in transfectants possessing multiple copies of a mutant target gene, but also demonstrate that the sequence specificity for BrdU-induced mutations is the same in Chinese hamster cells as previously observed with mouse cells.
Somat Cell Mol Genet 1992 Mar
PMID:Analysis of sequence specificity of 5-bromodeoxyuridine-induced reversion in cells containing multiple copies of a mutant gpt gene. 131 57

The mitochondrial DNA molecules of two interfertile algal species, Chlamydomonas smithii and C. reinhardtii, are co-linear except for a 1075 bp intron (the alpha-insert) that is present in the cob gene of C. smithii. The alpha-insert, a group I intron (Cs cob.1) containing an open reading frame (ORF) which encodes a basic, hydrophilic protein of 237 amino acids, is unidirectionally transmitted to all diploid progeny during interspecific crosses. In this report, we show that the Cs cob.1-encoded protein is a site-specific endonuclease (I-Csm I) which could mediate the intron transfer via the gene conversion mechanism. The Cs cob.1 ORF was cloned into the vector pMALcr1 and over-expressed as a hybrid protein fused to maltose-binding protein (MBP). This fusion protein exhibited an in vivo endonuclease activity which specifically cleaved the intron homing site within the intronless cob gene.
Plant Mol Biol 1992 Mar
PMID:The group I intron of apocytochrome b gene from Chlamydomonas smithii encodes a site-specific endonuclease. 131 90

Nodulation by the Rhizobium strain IC3342 causes a leaf curl syndrome in certain tropical legumes such as pigeon pea (Cajanus cajan) (N.M. Upadhyaya, J.V.D.K. Kumar Rao, D.S. Letham, and P.J. Dart, Physiological and Molecular Plant Pathology 39:357-373, 1991). Transposon (Tn5) mutagenesis of this leaf curl-inducing (Curl+) Rhizobium strain yielded two Curl- Fix- and three Curl- Fix+ mutants. Plasmid visualization and subsequent Southern blot hybridization analyses with Tn5, nif and nod gene probes showed that the Tn5 element had inserted into the symbiotic (Sym) plasmid in three of the mutants. Restriction endonuclease analyses indicated that none of the Tn5 insertions were closely linked. Tn5-containing EcoRI fragments were cloned from each mutant and used as probes to isolate the corresponding wild-type DNA fragments from a cosmid (pLAFR3) genomic library. Fix+ and/or Curl+ phenotypes were restored in each mutant by the introduction of cosmids containing the corresponding wild-type DNA. A closely related but Curl- Rhizobium strain ANU240 was shown, by Southern hybridization, to contain conserved DNA sequences of all but one of the identified genetic regions of the Curl+ Rhizobium strain IC3342. Cosmids containing the genetic region unique to the strain IC3342, designated lcr1, conferred a Curl+ phenotype on the strain ANU240. DNA sequence analysis of the cloned lcr1 region revealed five open reading frames (ORFs). The ORF2 showed homology with the Escherichia coli regulatory gene ompR, and ORF4 showed homology with E. coli and Rhizobium meliloti regulatory genes fnr and fixK, respectively.
Mol Plant Microbe Interact
PMID:Isolation and characterization of Rhizobium (IC3342) genes that determine leaf curl induction in pigeon pea. 131 72

The efficiency of bacteriophages CP-54 and CP-55 plating on Bacillus thuringiensis var. kumantoensis H18 (Kum) is decreased about 10-fold as compared with the efficiency of plating on Bacillus thuringiensis var. galleriae H5 (Gal). Bacteriophages having propagated for one cycle in Kum cells might be further grown in this strain without growth restriction. Two site-specific restriction enzymes isolated from Bacillus thuringiensis var. kumantoensis were designated BtkI and BtkII. The endonuclease BtkI recognises the same nucleotide sequence CGCG in DNA as recognised by the restriction endonuclease FnuDII; BtkII recognises the same nucleotide sequence GATC as the endonuclease Sau3A.
Mol Gen Mikrobiol Virusol 1992
PMID:[Site-specific restrictases from Bacillus thuringiensis var. Kumantoensis]. 132 Jan 99

When Escherichia coli HB101 harbors pWR127, a plasmid comprising the viaB gene from Citrobacter freundii WR7004 and the ColE1-derived pACKC1, the strain produces the virulence (Vi) antigen. Vi antigen expression is abolished (Vi- phenotype), however, when an IS1 or IS1-like DNA element inserts into the viaB region. To determine the sites of IS1 insertion, pWR127 DNAs extracted from 95 independently isolated Vi- strains were analyzed by digestion with the restriction endonuclease PstI and agarose gel electrophoresis. Ten insertion sites were found distributed non-randomly in an area of about 1.3 kb. Nine Vi+ strains (two Citrobacter, two E. coli, and five Salmonella strains), four of which contain pWR127, were then tested for the presence of IS1 by DNA-DNA hybridization. Of the nine strains, five were stable Vi+ and did not contain IS1. The other four which generated Vi- strains, contained IS1. When pRR134, a plasmid that contains IS1 was transferred into a stable Vi+ Salmonella typhimurium strain carrying pWR127 (OU5140), Vi- strains were produced from which pWR127 derivatives carrying IS1 inserts could be isolated. It appears, therefore, that the presence of an IS1 or IS1-like element in a strain is required for conversion of the Vi+ expression state to the Vi- expression state.
Mol Gen Genet 1992 Aug
PMID:Role of IS1 in the conversion of virulence (Vi) antigen expression in Enterobacteriaceae. 132 99


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