scholarly journals Dissociation of the AT-specific bifunctional intercalator [N-MeCys3,N-MeCys7]TANDEM from TpA sites in DNA

1995 ◽  
Vol 306 (1) ◽  
pp. 15-19 ◽  
Author(s):  
M C Fletcher ◽  
R K Olsen ◽  
K R Fox
Keyword(s):  
Dissociation Rate ◽  
Dnase I ◽  
Time Dependent ◽  
Base Pairs ◽  
Unusual Structure ◽  
Dnase I Footprinting ◽  
Calf Thymus ◽  
Stability Of Complexes ◽  
Dna Fragment ◽  
The Stability

We have examined the dissociation of [N-MeCys3,N-MeCys7]TANDEM, an AT-selective bifunctional intercalator, from TpA sites in mixed-sequence DNAs by a modification of the footprinting technique. Dissociation of complexes between the ligand and radiolabelled DNA fragments was initiated by adding a vast excess of unlabelled calf thymus DNA. Portions of this mixture were subjected to DNAse I footprinting at various times after adding the competitor DNA. Dissociation of the ligand from each site was seen by the time-dependent disappearance of the footprinting pattern. Within a natural DNA fragment (tyrT) the ligand dissociates from TTAT faster than from ATAT. We found that the stability of complexes with isolated TpA steps decreases in the order ATAT > TTAA > TATA. Dissociation from each of these sites is much faster than from longer regions of (AT)n. These results confirm the requirement for A and T base-pairs surrounding the TpA step and suggest that the interaction is strongest with regions of alternating AT, possibly as a result of its unusual structure. The ligand dissociates more slowly from the centre of (AT)n tracts than from the edges, suggesting that variations in dissociation rate arise from sequence-dependent variations in local DNA structure.

Regulatory elements controlling pituitary-specific expression of the human prolactin gene

1990 ◽  
Vol 10 (9) ◽  
pp. 4690-4700
Author(s):  
B Peers ◽  
M L Voz ◽  
P Monget ◽  
M Mathy-Hartert ◽  
M Berwaer ◽  
...  
Keyword(s):  
Regulatory Elements ◽  
Distal Region ◽  
Dnase I ◽  
Proximal Region ◽  
Base Pairs ◽  
Specific Expression ◽  
Positive Region ◽  
Dnase I Footprinting ◽  
Human Prolactin ◽  
Flanking Sequences

We have performed transfection and DNase I footprinting experiments to investigate pituitary-specific expression of the human prolactin (hPRL) gene. When fused to the chloramphenicol acetyltransferase (CAT) reporter gene, 5,000 base pairs of the 5'-flanking sequences of the hPRL gene were able to drive high cat gene expression in prolactin-expressing GH3B6 cells specifically. Deletion analysis indicated that this pituitary-specific expression was controlled by three main positive regulatory regions. The first was located just upstream from the TATA box between coordinates -40 and -250 (proximal region). We have previously shown that three motifs of this region bind the pituitary-specific Pit-1 factor. The second positive region was located in the vicinity of coordinates -1300 to -1750 (distal region). DNase I footprinting assays revealed that eight DNA motifs of this distal region bound protein Pit-1 and that two other motifs were recognized by ubiquitous factors, one of which seems to belong to the AP-1 (jun) family. The third positive region was located further upstream, between -3500 and -5000 (superdistal region). This region appears to enhance transcription only in the presence of the distal region.

scholarly journals Interaction of bleomycin with a bent DNA fragment

1992 ◽  
Vol 284 (3) ◽  
pp. 929-934 ◽  
Author(s):  
K P Nightingale ◽  
K R Fox
Keyword(s):  
Binding Sites ◽  
Dna Bending ◽  
Cobalt Complex ◽  
Dnase I ◽  
Cleavage Sites ◽  
Kinetoplast Dna ◽  
Bent Dna ◽  
Diethyl Pyrocarbonate ◽  
Dnase I Footprinting ◽  
Dna Fragment

The interaction of bleomycin with a kinetoplast DNA fragment has been examined using various footprinting techniques. This DNA adopts a bent structure and displays an unusually low gel mobility on account of its phased runs of adenines. The bleomycin-cobalt complex increases the mobility of this DNA fragment, in contrast with other DNAs which show a decreased rate of gel migration, suggesting that the antibiotic removes DNA bending, possibly via an unwinding mechanism. Removal of the bending is confirmed by hydroxy-radical footprinting which produces a more even ladder of bands in the presence of the ligand. Cleavage by bleomycin is at the sequence G-pyrimidine, though not all such sites are affected to the same extent and some cutting is found at GA and GG. DNase I footprinting confirms the antibiotic-binding sites but reveals that some strong cleavage sites do not yield footprints. Bleomycin renders adenines on the 3′ side of its cleavage sites (GT, GC and GA) hyper-reactive to diethyl pyrocarbonate.

scholarly journals Alternate-strand DNA triple-helix formation using short acridine-linked oligonucleotides

1994 ◽  
Vol 301 (2) ◽  
pp. 569-575 ◽  
Author(s):  
E Washbrook ◽  
K R Fox
Keyword(s):  
Metal Ion ◽  
Dimethyl Sulphate ◽  
Dnase I ◽  
Target Sequence ◽  
Base Pairs ◽  
Helix Formation ◽  
Dnase I Footprinting ◽  
Triple Helix Formation ◽  
Triple Helices ◽  
Dna Triple Helix

We have used DNAse I footprinting to examine the formation of intermolecular DNA triple helices at sequences containing adjacent blocks of purines and pyrimidines. The target sites G6T6.A6C6 and T6G6.C6A6 were cloned into longer DNA fragments and used as substrates for DNAse I footprinting, which examined the binding of the acridine (Acr)-linked oligonucleotides Acr-T5G5 and Acr-G5T5 respectively. These third strands were designed to incorporate both G.GC triplets, with antiparallel Gn strands held together by reverse Hoogsteen base pairs, and T.AT triplets, with the two T-containing strands arranged antiparallel to each other. We find that Acr-T5G5 binds to the target sequence G6T6.-A6C6, in the presence of magnesium at pH 7.0, generating clear DNAse I footprints. In this structure the central guanine is not recognized by the third strand and is accessible to modification by dimethyl sulphate. Under these conditions no footprint was observed with Acr-G5T5 and T6G6.C6A6, though this triplex was evident in the presence of manganese chloride. Manganese also facilitated the binding of Acr-T5G5 to a second site in the fragment containing the sequence T6G6.C6A6. This represents interaction with the sequence G4ATCT6, located at the boundary between the synthetic insert and the remainder of the fragment, and suggests that this bivalent metal ion may stabilize triplexes that contain one or two mismatches. Manganese did not affect the interaction of either oligonucleotide with G6T6.A6C6.

scholarly journals Regulatory elements controlling pituitary-specific expression of the human prolactin gene.

1990 ◽  
Vol 10 (9) ◽  
pp. 4690-4700 ◽  
Author(s):  
B Peers ◽  
M L Voz ◽  
P Monget ◽  
M Mathy-Hartert ◽  
M Berwaer ◽  
...  
Keyword(s):  
Regulatory Elements ◽  
Distal Region ◽  
Dnase I ◽  
Proximal Region ◽  
Base Pairs ◽  
Specific Expression ◽  
Positive Region ◽  
Dnase I Footprinting ◽  
Human Prolactin ◽  
Flanking Sequences

We have performed transfection and DNase I footprinting experiments to investigate pituitary-specific expression of the human prolactin (hPRL) gene. When fused to the chloramphenicol acetyltransferase (CAT) reporter gene, 5,000 base pairs of the 5'-flanking sequences of the hPRL gene were able to drive high cat gene expression in prolactin-expressing GH3B6 cells specifically. Deletion analysis indicated that this pituitary-specific expression was controlled by three main positive regulatory regions. The first was located just upstream from the TATA box between coordinates -40 and -250 (proximal region). We have previously shown that three motifs of this region bind the pituitary-specific Pit-1 factor. The second positive region was located in the vicinity of coordinates -1300 to -1750 (distal region). DNase I footprinting assays revealed that eight DNA motifs of this distal region bound protein Pit-1 and that two other motifs were recognized by ubiquitous factors, one of which seems to belong to the AP-1 (jun) family. The third positive region was located further upstream, between -3500 and -5000 (superdistal region). This region appears to enhance transcription only in the presence of the distal region.

Symmetrizable Difference Dchemes

2005 ◽  
Vol 5 (1) ◽  
pp. 3-50 ◽  
Author(s):  
Alexei A. Gulin
Keyword(s):  
Initial Data ◽  
Difference Scheme ◽  
Difference Schemes ◽  
Time Dependent ◽  
Stability Boundaries ◽  
Typical Representative ◽  
Variable Weight ◽  
The Stability ◽  
Conditions Of Stability ◽  
The Right

AbstractA review of the stability theory of symmetrizable time-dependent difference schemes is represented. The notion of the operator-difference scheme is introduced and general ideas about stability in the sense of the initial data and in the sense of the right hand side are formulated. Further, the so-called symmetrizable difference schemes are considered in detail for which we manage to formulate the unimprovable necessary and su±cient conditions of stability in the sense of the initial data. The schemes with variable weight multipliers are a typical representative of symmetrizable difference schemes. For such schemes a numerical algorithm is proposed and realized for constructing stability boundaries.

scholarly journals High-Speed Video Analysis of the Process Stability in Plasma Spraying

2021 ◽  
Author(s):  
K. Bobzin ◽  
M. Öte ◽  
M. A. Knoch ◽  
I. Alkhasli ◽  
H. Heinemann
Keyword(s):  
Plasma Spraying ◽  
High Speed ◽  
Plasma Jet ◽  
Plasma Jets ◽  
Time Dependent ◽  
Acquisition Time ◽  
Fast Fourier Transformation ◽  
Frequency Spectra ◽  
Dependent Signal ◽  
The Stability

AbstractIn plasma spraying, instabilities and fluctuations of the plasma jet have a significant influence on the particle in-flight temperatures and velocities, thus affecting the coating properties. This work introduces a new method to analyze the stability of plasma jets using high-speed videography. An approach is presented, which digitally examines the images to determine the size of the plasma jet core. By correlating this jet size with the acquisition time, a time-dependent signal of the plasma jet size is generated. In order to evaluate the stability of the plasma jet, this signal is analyzed by calculating its coefficient of variation cv. The method is validated by measuring the known difference in stability between a single-cathode and a cascaded multi-cathode plasma generator. For this purpose, a design of experiment, covering a variety of parameters, is conducted. To identify the cause of the plasma jet fluctuations, the frequency spectra are obtained and subsequently interpreted by means of the fast Fourier transformation. To quantify the significance of the fluctuations on the particle in-flight properties, a new single numerical parameter is introduced. This parameter is based on the fraction of the time-dependent signal of the plasma jet in the relevant frequency range.

scholarly journals Computational Studies of 4-Formylpyridinethiosemicarbazone and Structural and Biological Studies of Its Ni(II) and Cu(II) Complexes

2019 ◽  
Vol 2019 ◽  
pp. 1-10
Author(s):  
Mydhili P. Sripathi ◽  
Sireesha Berely ◽  
Chittireddy Venkata Ramana Reddy
Keyword(s):  
Elemental Analysis ◽  
Radical Scavenging ◽  
Molar Conductance ◽  
Computational Studies ◽  
Hydrolytic Cleavage ◽  
Bacterial Strains ◽  
Intercalative Mode ◽  
Biological Studies ◽  
Calf Thymus ◽  
The Stability

To understand the stability, chelation behaviour, and biological activity of 4-Formylpyridinethiosemicarbazone (H4FPT), it is important to recognize its interactive geometry. Hence, computational studies on geometrically optimized structures of thione and thiol forms of H4FPT were performed. Binary metal complexes of the ligand, H4FPT (L) with the Ni(II) and Cu(II) metal ions (M), were synthesized and characterized by various spectroanalytical techniques as elemental analysis, molar conductance, magnetic susceptibility measurements, LC-MS, TGA, IR, UV-Visible, ESR, and powder XRD. Elemental analysis, LC-MS, and TGA studies indicate 1:2 (ML2) composition for mononuclear Ni(II) complex and 1:1 (ML) composition for dinuclear Cu(II) complex. Electronic absorption titrations, fluorescence quenching studies, and viscosity measurements suggest intercalative mode of binding of the complexes with calf thymus DNA (CT-DNA). These complexes also promote hydrolytic cleavage of plasmid pBR322. The ligand (H4FPT) and its complexes showed moderate-to-good activity against Gram-positive and Gram-negative bacterial strains. The DPPH radical scavenging studies showed antioxidant nature of both complexes.

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