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

Intact and fast-sedimenting nucleoids of Bacillus licheniformis were isolated under low-salt conditions and without addition of detergents, polyamines or Mg2+. These nucleoids were partially unfolded by treatment with RNase and completely unfolded by treatments that disrupt protein-DNA interactions, like incubation with proteinase K, 0.1% sodium dodecyl sulphate and high ionic strength. Ethidium bromide intercalation studies on RNase-treated, proteinase-K-treated and non-treated nucleoids in combination with sedimentation analysis of DNase-I-treated nucleoids revealed that DNA is organized in independent, negatively supertwisted domains. In contrast to the DNA organization in bacterial nucleoids, isolated under high-salt conditions and in the presence of detergents (Stonington & Pettijohn, 1971; Worcel & Burgi, 1972), the domains of supertwisted DNA in the low-salt-isolated nucleoids studied here are restrained by protein-DNA interactions. A major role for nascent RNA in restraining supertwisted DNA was not observed. The superhelix density of B. licheniformis nucleoids calculated from the change of the sedimentation coefficient upon ethidium bromide intercalation, was of the same order of magnitude as that of other bacterial nucleoids and eukaryotic chromosomes, isolated under high-salt conditions: namely, -0.150 (corrected to standard conditions: 0.2 M-NaCl, 37 degrees C; Bauer, 1978). Electron microscopy of spread nucleoids showed relaxed DNA and regions of condensed DNA. Spreading in the presence of 100 micrograms ethidium bromide per ml revealed only condensed structures, indicating that nucleoids are intact. From spreadings of proteinase-K-treated nucleoids we infer that supertwisted DNA and the protein-DNA interactions, responsible for restraining the superhelical DNA conformation, are localized in the regions of condensed DNA.
J Mol Biol 1983 Jan 15
PMID:Folding of prokaryotic DNA. Isolation and characterization of nucleoids from Bacillus licheniformis. 618 37

A fragment of the E. coli chromosome including the recC gene has been cloned by in vitro recombinant DNA techniques into a phage lambda vector to give the recombinant phage lambda drecC. This was used to derive the phage lambda drecBC by in vivo recombination. On lysogenisation of recB and recC mutants with lambda drecBC wild levels of UV-resistance and RecBC DNase activity were restored. Infection of E coli with lambda drecBC led to the synthesis of phage-coded proteins of 125 kilodaltons (kd) and 135 kd that were not synthesised on infection with the original lambda vector, whereas a 125 kd protein but not a 135 kd protein was synthesised in similar experiments with lambda drecC. The recombinant phages, which are unable to form plaques, presumably due to the deletion of essential phage genes during their construction, provided useful starting points from which to subclone the recB, recC, and the neighbouring thyA and argA genes individually into multiple copy plasmid vectors.
Mol Gen Genet 1982
PMID:Construction of recombinant lambda phages that carry the E. coli recB and recC genes. 621 90

The sequence of almost 700 nucleotides encompassing the gene for ribosomal protein S20 of Escherichia coli has been determined using the chemical technique of Maxam and Gilbert (Maxam, A. M., and Gilbert, W. (1977) Proc. Natl. Acad. Sci. U. S. A. 74, 560-564). Comparison of this sequence with that of known bacterial promoters reveals two promoter-like sequences whose putative initiation sites for transcription lie 132 ("site 1") and 42 ("site 2") base pairs to the 5' side of the coding sequence. Partial digestion experiments demonstrate that RNA polymerase is capable of binding to and protecting both of these sites from DNase 1 digestion, consistent with their functioning as promoters. Interestingly, site 1 is more compact that any previously described bacterial promoter. A second feature of the sequence is the presence of UUG as the translational initiation codon. Finally, the nucleotide sequence supports the hypothesis (Jue, R. A., Woodbury, N. W., and Doolittle, R. F. (1980) J. Mol. Evol. 15, 129-148) that S20 is composed of three tandemly repeated domains.
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PMID:Nucleotide sequence of the gene for ribosomal protein S20 and its flanking regions. 626 39

DNase treatment used to convert supercoiled DNAs to the open form is suitable for molecular weight determination by electron microscopy or agarose gel electrophoresis. The comparative experiments showed that S1-nuclease has advantages over DNase for molecular weight determination in preparations containing many species of plasmids of significantly differing sizes, isolated from multiplasmid E. coli strains.
Mol Biol (Mosk)
PMID:[Nuclease S1 for DNA molecular weight determination in multiplasmid Escherichia coli strains]. 628 79

The binding of purified simian virus 40 (SV40) large T antigen (T) from monkey cells infected with wild-type SV40 virus to viral replication origin-containing DNA fragments was studied by DNase footprinting and restriction endonuclease protection methods. A strong affinity binding site (site 1) of 30 base-pairs and a second, adjacent 40 base-pair lower affinity binding site (site 2), which includes the origin of replication, were detected in these assays. These sites appear identical to those previously noted in similar assays performed with the Ad2 + D2 (D2) T protein. Heating T prior to incubation with DNA significantly increased the binding to these two sites, and the order of binding did not change. Moreover, protection of sequences was observed on both strands in these two sites suggesting that both strands can participate in binding of T to these two sites. Studies with DNAs from two internal site 2 deletion mutants as well as with a DNA fragment lacking the distal 13 base-pairs of site 2 revealed that sequences in the "early" portion of site 2 are sufficient for T binding to the intact site. Furthermore, use of a new assay that measures protection of DNA sequences from specific restriction enzyme cleavage revealed that site 2 can be subdivided into two subsites, 2A and 2B, where 2A corresponds to the above-noted early segment of this locus. In titration experiments, the affinity of 2A for T was greater than that of 2B. Hence, binding to a major portion of the replication initiation sequence (i.e. site 2) is the product of at least two interactions. Finally, analyses performed with DNA from a site 1 deletion mutant, cs1085, revealed that prior binding of T to this locus did not facilitate its binding to site 2. The opposite effect was observed when D2T was employed in these assays. Thus, although similar in many respects, these proteins display a detectable difference in their DNA binding mechanisms.
J Mol Biol 1983 Aug 25
PMID:Binding of simian virus 40 large T antigen from virus-infected monkey cells to wild-type and mutant viral replication origins. 631 Jan 27

The promoter for the gene encoding the low affinity L-arabinose uptake protein in Escherichia coli was studied. The promoter was cloned, sequenced, its transcription start site determined by S1 nuclease mapping, the proteins required for in vitro transcription were determined, and the regulatory protein binding sites located by DNase footprinting. The araE promoter shows no evidence of an operator site upstream from the CRP binding site, but otherwise it is similar to the araBAD promoter.
J Mol Biol 1983 Dec 25
PMID:The araE low affinity L-arabinose transport promoter. Cloning, sequence, transcription start site and DNA binding sites of regulatory proteins. 631 8

Isolated nuclei derived from simian virus 40 (SV40)-infected cells and incubated with [alpha-32P]UTP can elongate the in vivo preinitiated SV40 late RNA, synthesizing a viral RNA species 94 nucleotides long (attenuator RNA) as well as longer RNA molecules. In contrast to newly synthesized SV40 RNA, the attenuator RNA is not associated with the nuclear matrix. Pretreating the cells with 5,6-dichloro-1-beta-ribofuranosylbenzimidazole before the incubation of isolated nuclei in vitro, enhances the accumulation of the attenuator RNA, but again it is removed from nuclei by DNase and high salt. In contrast, pretreating the cells with proflavine, an intercalating drug that interferes with RNA secondary structure, prevents the accumulation of the attenuator RNA and increases the amount of the long RNA molecules. These RNA molecules become associated with the nuclear matrix. Isolated nuclear matrices from SV40-infected cells are highly enriched in transcriptionally active ternary complexes. Thus, isolated nuclear matrices that contain from 2 to 6% of SV40 DNA are capable of synthesizing at least 35% of the viral RNA synthesized in isolated nuclei after 2 to 15 minutes incubation with [alpha-32P]UTP. The RNA synthesized in vitro on purified nuclear matrices and isolated nuclei is derived from the same regions of the viral genome, suggesting that there is an association between transcribed DNA sequences and the nuclear matrix. The results suggest a major role for the nuclear matrix in controlling SV40 gene expression.
J Mol Biol 1984 Feb 05
PMID:Control of late simian virus 40 transcription by the attenuation mechanism and transcriptionally active ternary complexes are associated with the nuclear matrix. 631 19

Escherichia coli K-12 cells carrying the high copy number plasmid ColE2-P9 and a sfiA-lacZ gene fusion exhibit abnormally high levels of SOS-regulated phi sfiA-lacZ expression. Increased sfiA-lacZ expression is caused by the action of colicin E2, which is a DNase, rather than by the presence of multiple copies of a binding site for LexA protein, the repressor for the sfiA and colicin E2 genes. Expression of sfiA-lacZ was reduced to normal levels if the ColE2+ strain lacked the outer membrane colicin E2 receptor protein (BtuB) or if they carried an increased number of colicin E2 immunity genes. The results suggest that cultures of ColE2+ strains contain a small number of cells which produce colicin which can then enter other, non-producing cells in the culture and cause sufficient damage to the DNA to induce the SOS system. The levels of colicin E2 immunity in the producing cells is presumably sufficient to prevent extensive lethal effects of the colicin, but insufficient to prevent limited endonuclease activity. An important consequence of this phenomenon is that the DNase action of colicin E2 can stimulate its own production.
Mol Gen Genet 1983
PMID:Autoinduced synthesis of colicin E2. 634 78

RNA polymerase binds very tightly at a site called Brex in the lambda immunity region, to the left of the rex gene and about 600 nucleotides to the right of PL. The complex formed is resistant to 1 M NaCl in the absence of nucleotide triphosphate. While in vitro little or no transcription is observed from Brex, in vivo, when inserted in a plasmid vector which allows detection of its activity, it acts as an efficient promoter. We have mapped the site protected by RNA polymerase against DNase and determined its sequence which is abnormal compared that of an average promoter.
Mol Gen Genet 1980
PMID:An unusual RNA polymerase binding site in the immunity region of phage lambda. 645 Aug 73

Bacteriophage Mud (Casadaban and Cohen 1979) was used to bring the transcription of the gene for beta galactosidase (lacZ) under the control of the promoter of the structural gene for colicin Ib (cia(Ib)) on a derivative of the Col plasmid Col-Ib.P9. Transcription of this fusion operon was stimulated by agents which damaged cellular DNA (mitomycin C, bleomycin and colicin E2). Increased transcription of the cia-lacZ operon could be detected within 13 min of the addition of these agents. In a strain bearing the tif-1 (recA441) mutation, constitutive expression of the SOS DNA repair system at 42 degree C also increased transcription of the cia-lacZ operon. Transcription of the cia-lacZ operon was also stimulated by inhibition of DNA gyrase activity with nalidixic acid but not with novobiocin. Transitory inhibition of protein synthesis with chloramphenicol or by proline starvation of a proline auxotroph did not stimulate cia-lacZ transcription. Transcription of the cia-lacZ operon was substantially reduced in the presence of a recA mutation, but was largely unaffected by a mutation in recB affecting the RecBC DNase or by catabolite repression. Control experiments in which the production of colicin Ib was measured confirmed that the experiments with the fusion operon gave an accurate indication as to the activity of the wild type cia gene except for the effect of catabolite repression, where we observed up to 99% reduction in colicin Ib production in strains carrying mutant crp or cya alleles. The overall results confirm previous suggestions that there was considerable similarity between the regulatory systems controlling production of colicins and the repressor-dependent regulation of lambdoid prophage induction.
Mol Gen Genet 1981
PMID:Transcription regulation of colicin Ib synthesis. 646 Sep 13


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