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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

At least three promoters (1A, 1B, and 1C) control the expression of mRNA transcripts for the human glucocorticoid receptor (hGR) protein. An hGR 1A promoter/exon sequence (-218/+269) contains at least 12 deoxyribonuclease (DNase) I footprints that contain bound protein. Whereas four of these footprints (FP6, FP7, FP8, and FP11) contain bound hGR in protein-DNA complexes that are formed, only two (FP7 and FP11) appear to be important for the up-regulation of hGR 1A promoter/exon activity in T-lymphoblasts. Furthermore, the activity of these DNA elements depends upon the promoter context, leading to a redundant and complex regulation of expression of the hGR 1A promoter/exon. FP7 appears to be required for hormonal responsiveness in the absence of upstream sequences (+41/+191), whereas the hormonal responsiveness of FP11 requires a functional, adjacent FP12 DNA sequence. FP12 contains overlapping binding sites for the protooncogene transcription factors c-Myb and c-Ets. It seems likely that binding of either c-Myb or c-Ets to FP12 is necessary for the direct or indirect binding of the hGR to FP11 (a nonconsensus glucocorticoid response element), and the resultant steroid-responsiveness of the hGR 1A promoter/exon sequence. We propose that the identity of the accessory transcription factor bound to FP12 (c-Myb or c-Ets) may determine the nature of regulation (positive or negative) of hGR gene expression by hormone, and that this may be important for hormone-induced apoptosis in T cell acute lymphoblastic leukemia.
Mol Endocrinol 2004 Apr
PMID:Steroid-responsive sequences in the human glucocorticoid receptor gene 1A promoter. 1504 98

Members of the double-stranded RNA (dsRNA) specific RNase III family are known to use a conserved dsRNA-binding domain (dsRBD) to distinguish RNA A-form helices from DNA B-form ones, however, the basis of this selectivity and its effect on cleavage specificity remain unknown. Here, we directly examine the molecular requirements for dsRNA recognition and cleavage by the budding yeast RNase III (Rnt1p), and compare it to both bacterial RNase III and fission yeast RNase III (Pac1). We synthesized substrates with either chemically modified nucleotides near the cleavage sites, or with different DNA/RNA combinations, and investigated their binding and cleavage by Rnt1p. Substitution for the ribonucleotide vicinal to the scissile phosphodiester linkage with 2'-deoxy-2'-fluoro-beta-d-ribose (2' F-RNA), a deoxyribonucleotide, or a 2'-O-methylribonucleotide permitted cleavage by Rnt1p, while the introduction of a 2', 5'-phosphodiester linkage permitted binding, but not cleavage. This indicates that the position of the phosphodiester link with respect to the nuclease domain, and not the 2'-OH group, is critical for cleavage by Rnt1p. Surprisingly, Rnt1p bound to a DNA helix capped with an NGNN tetraribonucleotide loop indicating that the binding of at least one member of the RNase III family is not restricted to RNA. The results also suggest that the dsRBD may accommodate B-form DNA duplexes. Interestingly, Rnt1p, but not Pac1 nor bacterial RNase III, cleaved the DNA strand of a DNA/RNA hybrid, indicating that A-form RNA helix is not essential for cleavage by Rnt1p. In contrast, RNA/DNA hybrids bound to, but were not cleaved by Rnt1p, underscoring the critical role for the nucleotide located at 3' end of the tetraloop and suggesting an asymmetrical mode of substrate recognition. In cell extracts, the native enzyme effectively cleaved the DNA/RNA hybrid, indicating much broader Rnt1p substrate specificity than previously thought. The discovery of this novel RNA-dependent deoxyribonuclease activity has potential implications in devising new antiviral strategies that target actively transcribed DNA.
J Mol Biol 2004 Apr 23
PMID:Molecular requirements for duplex recognition and cleavage by eukaryotic RNase III: discovery of an RNA-dependent DNA cleavage activity of yeast Rnt1p. 1506 40

Two unusual enzymatic activities are required for the realization of a single molecule sequencing: a polymerase for copying a deoxyribonuclease (DNA) target into complementary flurophore-labeled DNA, and an exonuclease for the successive hydrolysis of the completely dye-labeled DNA. Recently, we found that the wild-type Klenow fragment of Escherichia coli DNA polymerase I is well-suited for the synthesis of DNA in a reaction set-up that contains exclusively specific rhodamine-labeled analogs of the natural pyrimidine nucleotides (dCTP and dTTP). This protocol describes the procedure used for the preparation of DNA that is labeled at all pyrimidine bases of one strand, as well as an example of enzymatic downstream processing of the DNA product.
Methods Mol Biol 2004
PMID:High-density labeling of DNA for single molecule sequencing. 1519 7

Type I restriction enzymes bind sequence-specifically to unmodified DNA and subsequently pull the adjacent DNA toward themselves. Cleavage then occurs remotely from the recognition site. The mechanism by which these members of the superfamily 2 (SF2) of helicases translocate DNA is largely unknown. We report the first single-molecule study of DNA translocation by the type I restriction enzyme EcoR124I. Mechanochemical parameters such as the translocation rate and processivity, and their dependence on force and ATP concentration, are presented. We show that the two motor subunits of EcoR124I work independently. By using torsionally constrained DNA molecules, we found that the enzyme tracks along the helical pitch of the DNA molecule. This assay may be directly applicable to investigating the tracking of other DNA-translocating motors along their DNA templates.
Nat Struct Mol Biol 2004 Sep
PMID:Real-time observation of DNA translocation by the type I restriction modification enzyme EcoR124I. 1533 77

Nuclease footprinting techniques were initially developed to investigate protein-deoxyribonucleic acid (DNA) interactions but these tools of molecular biology have also become instrumental for probing sequence-selective binding of small molecules to DNA. Here, the method is described and technical details are given for performing deoxyribonuclease (DNase) I footprinting with DNA-binding drugs. An example is presented where DNase I is used (as well as DNase II and micrococcal nuclease) to probe the patterns of sequence-selective recognition of DNA by the anticancer antibiotic actinomycin D. DNase I is a convenient endonuclease for detecting and locating the position of actinomycin-binding sites within GC-rich sequences.
Methods Mol Biol 2005
PMID:DNase I footprinting of small molecule binding sites on DNA. 1533 13

A new acid deoxyribonuclease (DNase) was purified from the cultured mycelia of Cordyceps sinensis, and designated CSDNase. CSDNase was purified by (NH(4))(2)SO(4) precipitation, Sephacryl S-100 HR gel filtration, weak anion-exchange HPLC, and gel filtration HPLC. The protein was single-chained, with an apparent molecular mass of ca. 34 kDa, as revealed by SDS-PAGE, and an isoelectric point of 7.05, as estimated by isoelectric focusing. CSDNase acted on both double-stranded (ds) and single- stranded (ss) DNA, but preferentially on dsDNA. The optimum pH of CSDNase was pH 5.5 and its optimum temperature 55. The activity of CSDNase was not dependent on divalent cations, but its enzymic activity was inhibited by high concentration of the cation: MgCl(2) above 150 mM, MnCl(2) above 200 mM, ZnCl(2) above 150 mM, CaCl(2) above 200 mM, NaCl above 300 mM, and KCl above 300 mM. CSDNase was found to hydrolyze DNA, and to generate 3-phosphate and 5-OH termini. These results indicate that the nucleolytic properties of CSDNase are essentially the same as those of other well-characterized acid DNases, and that CSDNase is a member of the acid DNase family. To our knowledge, this is the first report of an acid DNase in a fungus.
J Biochem Mol Biol 2004 Jul 31
PMID:Purification and characterization of an acid deoxyribonuclease from the cultured mycelia of Cordyceps sinensis. 1546 35

Genes encoding DNA-methyltransferases which recognize the same sequence 5'-GCATC-3' from SfaNI and Bst19I restriction-modification systems have been cloned and primary structures of these have been determined. It has been revealed that restriction-modification system Bst19I contains two DNA-methyltransferases M1.Bst19I and M2.Bst19I, whereas RM system SfaNI include only one DNA-methyltransferase M.SfaNI, N- and C-domain of which are homologous of M2.Bst19I and M1.Bst19I, respectively. M1.Bst19I and M2.Bst19I as well as both domains of M.SfaNI contain conservative elements in an order that is typical for N6-adenine DNA-methyltransferases alpha class. SfaNI and Bst19I DNA-methyltransferases share high homology level with methylases of FokI and BstF5I RM systems. Probably this reflects presence of the common DNA sequence 5'-GATG-3' in the recognition sites of all these RM systems. Basing on primary structures homology of methylases, highly conserved amino acid residues on known spatial model of DNA-methyltransferase M.DpnIIA have been determined.
Mol Biol (Mosk)
PMID:[Cloning, primary structure determination and comparative analysis of DNA-methyltransferases from SfaNI and Bst19I restriction-modification systems]. 1561 85

Promiscuous mutant EcoRI endonucleases produce lethal to sublethal effects because they cleave Escherichia coli DNA despite the presence of the EcoRI methylase. Three promiscuous mutant forms, Ala138Thr, Glu192Lys and His114Tyr, have been characterized with respect to their binding affinities and first-order cleavage rate constants towards the three classes of DNA sites: specific, miscognate (EcoRI*) and non-specific. We have made the unanticipated and counterintuitive observations that the mutant restriction endonucleases that exhibit relaxed specificity in vivo nevertheless bind more tightly than the wild-type enzyme to the specific recognition sequence in vitro, and show even greater preference for binding to the cognate GAATTC site over miscognate sites. Binding preference for EcoRI* over non-specific DNA is also improved. The first-order cleavage rate constants of the mutant enzymes are normal for the cognate site GAATTC, but are greater than those of the wild-type enzyme at EcoRI* sites. Thus, the mutant enzymes use two mechanisms to partially bypass the multiple fail-safe mechanisms that protect against cleavage of genomic DNA in cells carrying the wild-type EcoRI restriction-modification system: (a) binding to EcoRI* sites is more probable than for wild-type enzyme because non-specific DNA is less effective as a competitive inhibitor; (b) the combination of increased affinity and elevated cleavage rate constants at EcoRI* sites makes double-strand cleavage of these sites a more probable outcome than it is for the wild-type enzyme. Semi-quantitative estimates of rates of EcoRI* site cleavage in vivo, predicted using the binding and cleavage constants measured in vitro, are in accord with the observed lethal phenotypes associated with the three mutations.
J Mol Biol 2005 Apr 29
PMID:Thermodynamic and kinetic basis for the relaxed DNA sequence specificity of "promiscuous" mutant EcoRI endonucleases. 1581 70

Two GATC specific methylases together with Sau3AI isoschizomeric restriction endonuclease were partially characterized in Mitsuokella multiacida 46/5. This is the first report on the presence of solitary Dam methyltransferase alongside GATC specific restriction-modification system resulting in the unusual two-fold methylation of the GATC motifs.
Mol Biol Rep 2005 Dec
PMID:A unique pair of GATC specific DNA methyltransferases in Mitsuokella multiacida. 1632 90

A deoxyribonuclease (DNase) was isolated from viscera of the cold-adapted marine bivalve Icelandic scallop. The 42 kDa DNase was shown to be a single polypeptide which catalyses DNA hydrolysis in the absence of divalent cations. The isolated enzyme showed maximal activity at pH 6 and no activity above pH 7.2 against native DNA. The scallop DNase was slightly more susceptible to heat denaturation than porcine DNase II and makes double-strand breaks in circular DNA substrate as the porcine enzyme. The N-terminal sequence of the scallop DNase was shown to be closely similar to DNase II (EC 3.1.22.1) proteins from other organisms. The scallop DNase is in addition to plancitoxin I from A. planci, the only DNase II enzyme isolated from marine invertebrates.
Comp Biochem Physiol B Biochem Mol Biol 2006 Mar
PMID:Deoxyribonuclease II from the Icelandic scallop (Chlamys islandica): isolation and partial characterization. 1642 84


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