The Use of Diethyl Pyrocarbonate and Potassium Permanganate as Probes for Strand Separation and Structural Distortions in DNA

Kahl, Brenda F.; Paule, Marvin R.

doi:10.1007/978-1-60327-015-1_6

Cited by 10 publications

(6 citation statements)

References 51 publications

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“…KMnO 4 , on the other hand, reacts with exposed C5-6 position (double bond) of thymines (Ts) if they are not within a duplex or base-stacked, i.e. in a loop (50,51). Subsequent treatment with a base cleaves the backbone of modified Gs or Ts, the bands of which can be resolved by denaturing PAGE (52).…”

Section: Resultsmentioning

confidence: 99%

Alternative DNA structure formation in the mutagenic human c-MYC promoter

Mundo¹,

Zewail-Foote²,

Kerwin³

et al. 2017

Nucleic Acids Research

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Mutation ‘hotspot’ regions in the genome are susceptible to genetic instability, implicating them in diseases. These hotspots are not random and often co-localize with DNA sequences potentially capable of adopting alternative DNA structures (non-B DNA, e.g. H-DNA and G4-DNA), which have been identified as endogenous sources of genomic instability. There are regions that contain overlapping sequences that may form more than one non-B DNA structure. The extent to which one structure impacts the formation/stability of another, within the sequence, is not fully understood. To address this issue, we investigated the folding preferences of oligonucleotides from a chromosomal breakpoint hotspot in the human c-MYC oncogene containing both potential G4-forming and H-DNA-forming elements. We characterized the structures formed in the presence of G4-DNA-stabilizing K+ ions or H-DNA-stabilizing Mg2+ ions using multiple techniques. We found that under conditions favorable for H-DNA formation, a stable intramolecular triplex DNA structure predominated; whereas, under K+-rich, G4-DNA-forming conditions, a plurality of unfolded and folded species were present. Thus, within a limited region containing sequences with the potential to adopt multiple structures, only one structure predominates under a given condition. The predominance of H-DNA implicates this structure in the instability associated with the human c-MYC oncogene.

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Section: Resultsmentioning

confidence: 99%

Alternative DNA structure formation in the mutagenic human c-MYC promoter

Mundo¹,

Zewail-Foote²,

Kerwin³

et al. 2017

Nucleic Acids Research

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“…Partial DNA modification with dimethyl sulfate (DMS) and diethyl pyrocarbonate (DEPC) were carried out following standard procedures . Prior to addition of the modifying chemicals, DNA solutions were incubated at 0 °C for 1 h in the appropriate buffered solution: buffer M (50 mM Tris, pH 7.4, 1 mM MgCl 2 ), supplemented with either 100 mM LiCl (lithium buffer) or 100 mM KCl (potassium buffer).…”

Section: Methodsmentioning

confidence: 99%

A Contractile Electronic Switch Made of DNA

Huang

Sen

2010

J. Am. Chem. Soc.

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Double-helical DNA has been shown to conduct both electrons and electron holes, the latter over distances of >20 nm. DNA is thus a material of significant interest for the bottom-up construction of nanocircuitry. Here, we describe a contractile DNA nanoswitch, which can toggle between a structurally extended "off" state and a contracted "on" state, with a 40-fold conductivity difference between the two. To turn on, two short motifs of guanine-guanine mismatches in an otherwise standard double helix synapse to form a conductive G-quadruplex, bypassing an insulating element within the helix. This switch can be turned repeatedly on by treatment with millimolar concentrations of K(+) and turned off by sequestration of the K(+) by a crown ether. Circular dichroism and thymine-thymine photocross-linking experiments reveal that strand orientations within the on state G-quadruplex are wholly antiparallel and that the two conductive double-helices interface with the same face of the quadruplex. Although this DNA nanoswitch is chemically gated, it should be adaptable to other kinds of gating and thus serve as a prototype for increasingly sophisticated and complex electronic devices made of DNA.

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“…These constraints suppress the formation of G‐quadruplexes or, in other words, the integrity of the double helix needs to be disrupted for a G‐quadruplex to form. To obtain structural insights, we analyzed the structural status of the partially transcribed DNAs using potassium permanganate (KMnO 4 ) footprinting in which thymine residues were preferentially cleaved in melted or distorted DNA duplexes 13. During transcription, a RNAP denatures a DNA duplex and maintains a small transcription bubble that moves along with the enzyme to accommodate nascent RNA synthesis.…”

mentioning

confidence: 99%

Strand‐Biased Formation of G‐Quadruplexes in DNA Duplexes Transcribed with T7 RNA Polymerase

et al. 2015

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G-quadruplex-forming sequences are enriched near transcription start sites (TSSs) in animal genes. They readily form G-quadruplexes in transcription, which in turn regulate transcription. Therefore, the control of G-quadruplex formation is important for their functionality. It is now shown that G-quadruplexes form efficiently on the non-template, but hardly on the template DNA strand in the downstream vicinity of TSSs in DNA duplexes when they are transcribed by the T7 RNA polymerase (RNAP). Structural analysis reveals that the T7 RNAP causes distortion in a DNA duplex both inside and in front of the enzyme. This structural distortion leads to strand-biased G-quadruplex formation when a G-quadruplex-forming sequence is partially fed into the T7 RNAP to a position about seven nucleotides away from the front of RNA synthesis. Based on these facts, we propose a model for the strand-biased formation of G-quadruplexes in transcribed DNA duplexes.

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The Use of Diethyl Pyrocarbonate and Potassium Permanganate as Probes for Strand Separation and Structural Distortions in DNA

Cited by 10 publications

References 51 publications

Alternative DNA structure formation in the mutagenic human c-MYC promoter

Alternative DNA structure formation in the mutagenic human c-MYC promoter

A Contractile Electronic Switch Made of DNA

Strand‐Biased Formation of G‐Quadruplexes in DNA Duplexes Transcribed with T7 RNA Polymerase

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