T<sub>4</sub> polynucleotide ligase catalyzed joining on triple-stranded nucleic acids

Raae, Arnt J.; Kleppe, Kjell

doi:10.1021/bi00607a037

Cited by 5 publications

(4 citation statements)

References 20 publications

(42 reference statements)

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“…Effective peptide concentrations for stabilization were in the range of 10-5 to 10-2 M. Pentalysine was the most effective triplex stabilizer of the three basic peptides studied. It is effective at concentrations lower than those required for stabilization by other peptides and similar to the effective concentrations of spermine which is a well-known triplex stabilizer (Glaser & Gabbay, 1968;Raae & Kleppe, 1978;Hampel et al, 1991;Hanvey et al, 1991;Singleton & Dervan, 1993;Thomas & Thomas, 1993). Significant triplex formation was observed at pH 7 in the presence of pentalysine.…”

Section: Stabilization Of Intermodular Triplex Dna By Basicmentioning

confidence: 89%

“…Polyamines are known to stabilize nucleic acids against thermal denaturation (Tabor, 1962;Glaser & Gabbay, 1968;Thomas & Bloomfield, 1984), facilitate DNA condensation (Gosule & Schellman, 1978), promote the B to A transition (Minyat et al, 1978), and induce left-handed Z-DNA (Wang et al, 1979;Behe & Felsenfeld, 1981). Both divalent metal cations and polyamines have been shown to stabilize triple-helical nucleic acids Glaser & Gabbay, 1968;Raae & Kleppe, 1978; Kohwi & Kohwi-Shigematsu, 1978;Hampel et al, 1991;Hanvey et al, 1991;Maikov et al, 1993;Singleton & Dervan, 1993;Thomas & Thomas, 1993). The interaction of nucleic acids with lysine-and arginine-rich polypeptides as reasonable models for basic proteins has been extensively studied [Helene and Maurizot (1981) for a review].…”

Section: Stabilization Of Intermodular Triplex Dna By Basicmentioning

confidence: 99%

“…999902-133 to R.R.S. concentrations of polyamines, whose distributed charges allow them to interact simultaneously with different nucleic acid strands, are effective in the stabilization of triple-helical nucleic acids in vitro (Glaser & Gabbay, 1968;Raae & Kleppe, 1978;Hampel et al, 1991;Hanvey et al, 1991;Singleton & Dervan, 1993;Thomas & Thomas, 1993). As high as 1 mM concentrations of polyamines can be detected in the cell (Tabor & Tabor, 1976;Sarhan & Seiler, 1989), where they are predominantly bound to macromolecules, including nucleic acids and phospholipids (Davis et al, 1992).…”

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confidence: 99%

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Stabilization of Triple-Helical Nucleic Acids by Basic Oligopeptides

Potaman

Sinden²

1995

Biochemistry

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Intermolecular triplex DNA is stabilized by metal cations and polyamines which reduce repulsion between the negatively charged phosphates of the three nucleic acid strands. We use a quantitative chemical-probing assay involving protection of duplex guanines in a homopyrimidine.homopurine (Py.Pu) sequence from dimethyl sulfate modification to study effects of basic oligopeptides on the stability of triplex DNA. An intermolecular protonated pyrimidine.purine.pyrimidine (Py.Pu*Py) triplex formed readily between a duplex DNA region and a 14-mer pyrimidine triplex-forming oligonucleotide (TFO) at pH 5. The triplex was stabilized at pH by the addition of magnesium ions. In the presence of spermine and lysine-rich peptides, the intermolecular triplex was stabilized up to pH 6.5-7.0. The effective peptide concentration required for stabilization was 10(-5)-10(-2) M. Of the basic peptides studied, pentalysine (Lys-Lys-Lys-Lys-Lys) was the most effective triplex stabilizer. It was effective at concentrations which are lower than those required for Lys-Gly-Lys-Gly-Lys and Lys-Ala-Lys-Ala-Lys and are similar to active concentrations of spermine. Basic peptides were more effective at stabilizing a Py.Pu*Py triplex than a pyrimidine.purine.purine (Py.Pu*Pu) triplex. At 1 mM, Lys-Lys-Lys-Lys-Lys stabilized the Py.Pu*Pu triplex at a level comparable to stabilization by Mn2+ and spermine, whereas Lys-Gly-Lys-Gly-Lys and Lys-Ala-Lys-Ala-Lys resulted in weaker TFO binding. The concentration of TFOs required to form triplex DNA were significantly reduced in the presence of peptides.(ABSTRACT TRUNCATED AT 250 WORDS)

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Section: Stabilization Of Intermodular Triplex Dna By Basicmentioning

confidence: 89%

Section: Stabilization Of Intermodular Triplex Dna By Basicmentioning

confidence: 99%

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confidence: 99%

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Stabilization of Triple-Helical Nucleic Acids by Basic Oligopeptides

Potaman

Sinden²

1995

Biochemistry

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“…It is able to join two dsDNAs (blunt‐end or sticky‐end ligation), or seal a break between two ssDNA fragments annealed on the complementary DNA strand (nick‐ligation). It can join phosphodiester linkages on triple‐stranded nucleic acids [2], seal single‐stranded 1–5‐nucleotide gaps [3], and act as a lyase, removing apurininc/apyrimidinic (AP) sites in DNA [4]. The enzyme requires a bivalent metal cation such as Mg 2+ or Mn 2+ , and joins DNA using ATP as a coenzyme.…”

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confidence: 99%

Kinetics and thermodynamics of nick sealing by T4 DNA ligase

Cherepanov¹,

Vries

2003

European Journal of Biochemistry

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T4 DNA ligase is an Mg 2+ -dependent and ATP-dependent enzyme that seals DNA nicks in three steps: it covalently binds AMP, transadenylates the nick phosphate, and catalyses formation of the phosphodiester bond releasing AMP. In this kinetic study, we further detail the reaction mechanism, showing that the overall ligation reaction is a superimposition of two parallel processes: a ÔprocessiveÕ ligation, in which the enzyme transadenylates and seals the nick without dissociating from dsDNA, and a ÔnonprocessiveÕ ligation, in which the enzyme takes part in the abortive adenylation cycle (covalent binding of AMP, transadenylation of the nick, and dissociation). At low concentrations of ATP (< 10 lM) and when the DNA nick is sealed with mismatching base pairs (e.g. five adjacent), this superimposition resolves into two kinetic phases, a burst ligation (% 0.2 min )1 ) and a subsequent slow ligation (% 2 · 10 )3 min )1 ). The relative rate and extent of each phase depend on the concentrations of ATP and Mg 2+ . The activation energies of self-adenylation (16.2 kcalAEmol )1 ), transadenylation of the nick (0.9 kcalAEmol )1 ), and nicksealing (16.3-18.8 kcalAEmol )1 ) were determined for several DNA substrates. The low activation energy of transadenylation implies that the transfer of AMP to the terminal DNA phosphate is a spontaneous reaction, and that the T4 DNA ligase-AMP complex is a high-energy intermediate. To summarize current findings in the DNA ligation field, we delineate a kinetic mechanism of T4 DNA ligase catalysis.

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Probing DNA triple helix structure by chemical ligation

1991

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A series of oligomeric double and triple helical DNAs with irregular sequences of homo?urine and homopyrimidine strands were prepared. DNA triplexes were identified by CD spectroscopy and thermal denaturation profiles (biphaslc helix-coil transition). Condensation of ohgonucleotides on single and double-stranded DNA templates was performed using water-soluble carbodiimide, phosphodiester and pyrophosphate internucleotide bonds being newly formed. Such chemical ligation proved to be a sensitive monitor of changes in the sugar-phosphate backbone resulting from conversion of double to triple helix and of third-strand binding.DNA triplex; Chemical ligation of nucleic acids 1, INTRODUCTIONRenewed interest in DNA triple helices has recently intensified because of their potential role in the regulation of the eukaryotic genome [ 1,2]. There is also interest in generating nucleic acid ligands that recognize double-stranded targets. Such probes could serve as artificial gene repressors [3] and as DNA cleaving agents in sequencing procedures or in chromosome mapping [4]. Triple helices are believed to adopt an A-form structure, the purine strand and one pyrimidine strand being Watson-Crick paired and the other pyrimidine strand being Hoogsteen base-paired parallel to the purine strand along the major grooves [5-g]. In addition to physical methods [5-71, chemical approaches such as chemical and enzymatic cleavage-protection assays [8] and sequence-specific cIeavage of double helical DNA [9] have been used for studying the molecular structure of triplexes. Recently, it was also reported that double-stranded DNA can serve as a template to promote BrCN-induced condensation of homopyrimidine Hoogsteen paired third strands [lo], Using a different condensing agent, the present study demonstrates the potentialities of chemical ligation for monitoring changes in the sugar-phosphate backbone on converting a DNA duplex to triplex and for following third-strand binding. Because the two homopyrimidine strands of a triplex are antiparallel, they cannot replace each other in the triplex structure when irregular sequences are used. This makes it possible to indepen- MATERIALS AND METHODS Oligodeoxyribonucleotideswere synthesized by the phosphoramidite method on a Cyclon (Biosearch) DNA synthesizer. TTCCCTC and TCCTCTCTA were phosphorylated at the 5' end with polynucleotide kinase and [r-"P]ATP.ACTCCCTTCp was obtained from ACTCCCTTCrU as described in Ill]. CD1 and MES were from Merck. 2.1, Optical tneasurenientsEquimolar solutions of oligonucleotidcs l-7 (Scheme 1) were mixed to give comptexcs I-VI (Scheme 2). e260 VdUeS for oligomers 3-5 were assumed to be 8200, 13300 and 9000 (per monomer), respectively. iMelting curves of oligomer mixtures in 0.05 M MES, pH 6.0,0.02 M MgClz (buffer A) were determined on a Cary 219 spectrophotometer. The data were processed on a'tiewlett Packard 9825A computer. Circular dichroism measurements were made using a Jasco J-500C spectrometer. Cliet~lical ligation reactiorrOligomer mixtures (0.5 x IO-' M (...

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T₄ polynucleotide ligase catalyzed joining on triple-stranded nucleic acids

Cited by 5 publications

References 20 publications

Stabilization of Triple-Helical Nucleic Acids by Basic Oligopeptides

Stabilization of Triple-Helical Nucleic Acids by Basic Oligopeptides

Kinetics and thermodynamics of nick sealing by T4 DNA ligase

Probing DNA triple helix structure by chemical ligation

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T4 polynucleotide ligase catalyzed joining on triple-stranded nucleic acids

Cited by 5 publications

References 20 publications

Stabilization of Triple-Helical Nucleic Acids by Basic Oligopeptides

Stabilization of Triple-Helical Nucleic Acids by Basic Oligopeptides

Kinetics and thermodynamics of nick sealing by T4 DNA ligase

Probing DNA triple helix structure by chemical ligation

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Product

Resources

About

T₄ polynucleotide ligase catalyzed joining on triple-stranded nucleic acids