Peculiarities of DNA-proflavine binding under different concentration ratios

Phys. Chem. Chem. Phys.

2013

DNA intercalation is a clinically relevant biophysical process due to its potential to inhibit the growth and survival of tumor cells and microbes through the arrest of the transcription and replication processes. Extensive kinetic and thermodynamic studies have followed since the discovery of the intercalative binding mode. However, the molecular mechanism and the origin of the thermodynamic and kinetic profile of the process are still not clear. Here we have constructed the free energy landscape of intercalation, de-intercalation and dissociation from both the major and minor grooves of DNA using extensive all-atom metadynamics simulations, capturing both the free energy barriers and stability in close agreement with fluorescence kinetic experiments. In the intercalated state, an alternate orientation of proflavine is found with an almost equal stability compared to the crystal orientation, however, separated by a 5.0 kcal mol(-1) barrier that decreases as the drug approaches the groove edges. This study provides a comprehensive picture in comparison with experiments, which indicates that the intercalation and de-intercalation of proflavine happen through the major groove side, although the effective intercalation barrier increases because the path of intercalation goes through the stable (abortive) minor groove bound state, making the process a millisecond long one in excellent agreement with the experiments. The molecular origin of the higher barrier for the intercalation from the minor groove side is attributed to the desolvation energy of DNA and the loss of entropy, while the barrier from the major groove, in the absence of desolvation energy, is primarily entropic.

Section: Introductionmentioning

confidence: 99%

Intercalation and de-intercalation pathway of proflavine through the minor and major grooves of DNA: roles of water and entropy

Phys. Chem. Chem. Phys.

2013

“…Fluorescence properties of the intercalating drugs enabled exhaustive thermodynamic and kinetic studies, − while structural investigations were carried out using NMR, crystallographic studies, molecular modeling, − force spectroscopic methods, and so forth. Most of the kinetic studies on intercalation indicate that the process comprises at least two steps: a fast bimolecular outside binding, followed by a slow intercalation.…”

Section: Introductionmentioning

confidence: 99%

Molecular Mechanism of Direct Proflavine–DNA Intercalation: Evidence for Drug-Induced Minimum Base-Stacking Penalty Pathway

2012

J. Phys. Chem. B

DNA intercalation, a biophysical process of enormous clinical significance, has surprisingly eluded molecular understanding for several decades. With appropriate configurational restraint (to prevent dissociation) in all-atom metadynamics simulations, we capture the free energy surface of direct intercalation from minor groove-bound state for the first time using an anticancer agent proflavine. Mechanism along the minimum free energy path reveals that intercalation happens through a minimum base stacking penalty pathway where nonstacking parameters (Twist→Slide/Shift) change first, followed by base stacking parameters (Buckle/Roll→Rise). This mechanism defies the natural fluctuation hypothesis and provides molecular evidence for the drug-induced cavity formation hypothesis. The thermodynamic origin of the barrier is found to be a combination of entropy and desolvation energy.

“…At this concentration, no external bound proflavine is seen experimentally. 7,22 Therefore, we attempt here to directly probe the association using molecular dynamics simulation with a higher concentration of proflavine molecules around DNA (P/D = 1) at which outside bound states were predicted experimentally. 18 Moreover, we performed this study at four different ionic concentrations (0.0 M, 0.15 M, 0.5 M and 1.0 M NaCl) to understand the effect of ions on the drug-DNA association.…”

Section: (C) Proflavine Aggregation In the Presence Of Dnamentioning

confidence: 99%

“…Only recently, it was shown that a low P/D ratio makes the outside-bound state stable. 7 The outside bound state is also crystallized using modified DNA. 25 It was proposed that the stacking of proflavine would occur along the DNA axis because of the presence of phosphate groups, which would attract the cationic proflavine molecules.…”

Section: (D) Structure Of the Outside-bound Statementioning

confidence: 99%

“…5 A similar trend in timescales is also obtained for a modified DNA (halogenated DNA) where the faster time constant is 0.24 ms and the slower time constant is 1.3 ms. 6 Of the two states, the intercalated state is more stable. Thus, the nature and structure of it was captured by several experimental studies including spectroscopic, 3,5,7 NMR, 8 crystallographic, 9 computational, and theoretical studies. [10][11][12][13][14][15][16][17] The outside bound state being a metastable state remained elusive and received relatively less attention.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Structure and dynamics of proflavine association around DNA

Phys. Chem. Chem. Phys.

2016

Proflavine is a small molecule that intercalates into DNA and, thereby, acts as an anticancer agent. Intercalation of proflavine is shown to be a two-step process in which the first step is believed to be the formation of a pre-intercalative outside bound state. Experimental studies so far have been unable to capture the nature of the outside bound state. However, the sub-millisecond timescale observed in fluorescence kinetic experiments is often attributed to the binding of proflavine outside of DNA. Here, we have performed molecular dynamics simulations with multiple proflavine molecules to study the structure and dynamics of the formation of the outside bound state of DNA at different ion concentrations. We observed that the timescale of the outside bound state formation is, at least, five orders of magnitude faster (in nanoseconds) than the experimentally reported timescale (sub-milliseconds) attributed to binding outside DNA. Moreover, we also observed the stacked arrangement of proflavine all around DNA, which is different from the experimentally predicted stacking arrangement perpendicular to the helical axis of DNA in the close vicinity of the phosphate groups. This study, therefore, provides insight into the molecular structure and dynamics of the pre-intercalative outside bound state and will help in understanding the overall intercalation mechanism.