Slow relaxational processes in the melting of linear biopolymers: A theory and its application to nucleic acids

Anshelevich, V. V.; Vologodskii, Alexander; Lukashin, A. V.; Frank‐Kamenetskii, Maxim D.

doi:10.1002/bip.360230105

Cited by 121 publications

(72 citation statements)

References 32 publications

(13 reference statements)

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“…In (3) the melting of this DNA was shown to be equilibrium at a high ionic strength (lxSSC). Figure 3 shows that the position of peak 3 in lxSSC This is exactly what was predicted by the theory of slow relaxation processes in DNA melting (1). Figure 3 shows that the amplitude and width of the fourth and largest peak also change with changing heating rate at the high ionic strength.…”

Section: Resultssupporting

confidence: 75%

The nonequilibrium character of DNA melting: effects of the heating rate on the fine structure of melting curves

Kozyavkin

Lyubchenko

1984

Nucl Acids Res

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SUMMARYTo investigate the effects of heating rate on the DNA melting profile and to test the predictions of the theory of slow relaxation processes in DNA melting (1) concerning these effects, we obtained differential melting curves for the Bsp I Cl fragment of T7 DNA (1461 bp) and the Sma I-Eco RI fragment of Col El DNA (1291 bp) at heating Sates of 0.05 and 0.5 deg/min. At low ionic strength (0.02 M Na ) the heating rate has been shown to affect the position of the third peak in melting curve for Cl fragment.According to the melting maps (2), this peak corresponds to the unwinding of the section between the end of the molecule and Vhe region already melted.At high ionic strength (0.2 M Na ), when the melting of this DNA is reversible (3), the position of the peaks does not depend on the heating rate.In the case of the Col El DNA fragment the heating rate affects, as might be expected from the melting maps (4), only the last peak, as the melting of the last section is always nonequilibrium. The results of the study are in good qualitative agreement and in satisfactory quantitative agreement with the theoretical predictions (1).

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Section: Resultssupporting

confidence: 75%

The nonequilibrium character of DNA melting: effects of the heating rate on the fine structure of melting curves

Kozyavkin

Lyubchenko

1984

Nucl Acids Res

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“…This situation is analogous to the cases of the dissociation of linear DNA duplexes (34) or PNA-DNA triplex invasion complexes (35,36). Indeed, the energetics of the elementary steps of these processes look very similar-dissociation of two PNA-DNA base pairs of the double-duplex invasion complex results in the restoration of one DNA base pair.…”

Section: Mechanism Of the Double-duplex Invasionmentioning

confidence: 77%

Kinetics and mechanism of the DNA double helix invasion by pseudocomplementary peptide nucleic acids

Demidov

Protozanova

Izvolsky

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

102

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If adenines and thymines in two mutually complementary mixedbase peptide nucleic acid (PNA) oligomers are substituted with diaminopurines and thiouracils, respectively, so-called pseudocomplementary PNAs (pcPNAs) are created. Pairs of pcPNAs have recently demonstrated an ability to highly selectively target essentially any designated site on double-stranded DNA (dsDNA) by forming very stable PNA-DNA strand-displacement complexes via double duplex invasion (helix invasion). These properties of pcPNAs make them unique and very promising ligands capable of denying the access of DNA-binding proteins to dsDNA. To elucidate the sequence-unrestricted mechanism of sequence-specific dsDNA recognition by pcPNAs, we have studied the kinetics of formation of corresponding PNA-DNA complexes at various temperatures by the gel-shift assay. In parallel, the conditions for possible selfhybridization of pcPNA oligomers have been assayed by mixing curve (Job plot) and thermal melting experiments. The data indicate that, at physiological temperatures (Ϸ37°C), the equilibrium is shifted toward the pairing of corresponding pcPNAs with each other. This finding explains a linear concentration dependence, within the submicromolar range, of the pcPNA invasion rate into dsDNA at 37°C. At elevated temperatures (>50°C), the rather unstable pcPNA duplexes dissociate, yielding the expected quadratic dependence for the rate of pcPNA invasion on the PNA concentration. The polycationic character of pcPNA pairs, carrying the duplicated number of protonated terminal PNA residues commonly used to increase the PNA solubility and binding affinity, also explains the self-inhibition of pcPNA invasion observed at higher PNA concentrations. Melting of pcPNA duplexes occurs with the integral transition enthalpies ranged from ؊235 to ؊280 kJ⅐mol ؊1 , contributing to an anomalously high activation energy of Ϸ150 kJ⅐mol ؊1 found for the helix invasion of pcPNAs carrying four different nucleobases. A simplified kinetic model for pcPNAs helix invasion is proposed that interprets all unusual features of pcPNAs binding to dsDNA. Our findings have important implications for rational use of pcPNAs.double-stranded DNA ͉ pseudocomplementary PNA ͉ sequence-selective recognition P eptide nucleic acids (PNAs) and their derivatives are of significant biomedical and biotechnological interest as prospective biomolecular tools for highly selective manipulation of nucleic acids (1-8). Recently, a new modification of PNAs has been introduced for sequence-unrestricted targeting of doublestranded DNA (dsDNA). Along with ordinary guanines and cytosines, these PNAs, dubbed pseudocomplementary PNAs (pcPNAs; refs. 9 and 10), carry 2,6-diaminopurines (D) and 2-thiouracils ( s U) instead of adenines and thymines, respectively.Model building revealed a steric clash between the bulky thio group of s U (or similarly modified thymine) and one of the two amino groups of D within the s U-D pair (9, 11). This clashing effect must severely destabilize the pcPNA-pcPNA duplexes whereas bindin...

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“…Of course, the final stage is also reversible but the reverse kinetic constant is very small when the conditions favor PNA2/DNA triplex formation. Under such conditions the kinetic constant of dissociation of the complex is known to depend exponentially on the length of the complex (18,19). This explains why the "incorrect" triplexes are predominantly short-lived whereas the "correct" triplex is very long-lived.…”

Section: Resultsmentioning

confidence: 99%

Kinetics and mechanism of polyamide ("peptide") nucleic acid binding to duplex DNA.

Demidov

Yavnilovich

Belotserkovskii

et al. 1995

Proc. Natl. Acad. Sci. U.S.A.

195

147

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To elucidate the mechanism of recognition of double-stranded DNA (dsDNA) by homopyrimidine polyamide ("peptide") nucleic acid (PNA) leading to the stranddisplacement, the kinetics of the sequence-specific PNA/DNA binding have been studied. The binding was monitored with time by the gel retardation and nuclease S1 cleavage assays. The experimental kinetic curves obey pseudo-first-order kinetics and the dependence of the pseudo-first-order rate constant, kps, on PNA concentration, P, obeys a power law kps the proposed kinetic scheme is performed. The interpretation of our experimental data in the framework of the proposed kinetic scheme leads to the following conclusions. The sequence specificity of the recognition is essentially provided at the "search" step of the process, which consists in the highly reversible transient formation of duplex between one PNA molecule and the complementary strand of duplex DNA while the other DNA strand is displaced. This search step is followed by virtually irreversible "locking" step via PNA2/DNA triplex formation. The proposed mechanism explains how the binding of homopyrimidine PNA to dsDNA meets two apparently mutually contradictory features: high sequence specificity of binding and remarkable stability of both correct and mismatched PNA/DNA complexes.A new type of DNA analogue, polyamide ("peptide") nucleic acid (PNA), was described in 1991 (1). This sequence-specific DNA binding reagent is believed to be a very promising drug (2, 3) with numerous potential applications (4). For homopyrimidine PNAs a unique type of duplex DNA/drug interaction is observed. It consists of PNA binding to one of the DNA strands through formation of stable PNA2/DNA triplex while the noncomplementary DNA strand is left in single-stranded state (1, 5, 6) thus forming a structure that we call the P loop. P-loop formation leads to selective inhibition of protein binding to DNA (7,8), results in transcription elongation arrest (2,7,9,10), creates an artificial transcription promoter (11), makes it possible to convert single-strand-specific nucleases into sequence-selective cutters (12), and, if PNA is biotinylated, to place electron-microscopy markers on doublestranded DNA (dsDNA) (13).For biomedical and molecular biological applications of PNA it is essential to understand the factors controlling PNA/ DNA binding and its sequence specificity. The data indicate that under conditions in which the PNA/DNA complexes are normally studied, the binding is virtually irreversible (5), thus implying a crucial role of kinetic factors in the stranddisplacement reaction. Here we present a kinetic study of homopyrimidine PNAs binding to dsDNA and propose a kinetic model for PNA/DNA sequence-specific recognition.

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Slow relaxational processes in the melting of linear biopolymers: A theory and its application to nucleic acids

Cited by 121 publications

References 32 publications

The nonequilibrium character of DNA melting: effects of the heating rate on the fine structure of melting curves

The nonequilibrium character of DNA melting: effects of the heating rate on the fine structure of melting curves

Kinetics and mechanism of the DNA double helix invasion by pseudocomplementary peptide nucleic acids

Kinetics and mechanism of polyamide ("peptide") nucleic acid binding to duplex DNA.

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