1. Determination of Free Energy Landscapes via Computer Simulations and its Application to the DNA i-motif Vasileios A. Tatsis, Raghvendra P. Singh and Andreas Heuer Institute of Physical Chemsitry, WWU, Münster, Germany
Simulation system definitions
MD Engine: GROMACS with PLUMED
Unfolding simulations temperatures: 300 and 500 K
Electro-neutral system: 21 Na+ (for backbone charges)
Water model : TIP3P(≈ 6000 water molecules)
Simulation Box: 6.7 nm3.
Force-field: Amber03-bsc0 (for nucleic acids)
3D Periodic Boundary Conditions
PME for electrostatic interactions
Simulations times up to 1μs (variable)
Acknowledgements
Molecular Dynamic Simulations
Figure 2: Upper panel (L to R): Free energy landscapes resulted from: multiple unbiased MD simulations, at 300K; (A and B ) Two well-tempered metadynamics simulations. The distance between the center of mass for C1 and T22 to A11d[C1/T22)-A11] and the distance between C1 and T22 d[C1-T22] were used as reaction coordinates. Units: Energy in kJ/mol., time length in 1μs. Lower panel (L to R): Normalized histograms of RMSD between the modeled i-motif structure and the crystal structure (pdb code: 1ELN) and Heat map describing the average RMSD value using the two assisting collective variables. Data extracted from the first well-tempered metadynamics simulations of 700 ns.
Reaction coordinates were chosen using COM of rC1/T22, rC1,T22/A11.
Metadynamics shows deeper minimum while system seems stuck at starting coordinates in MD.
The RMSD shows that metadynamics explored phase space efficiently.
i-motif
An atypical DNA formed at low pH and a high temperature, C-rich telomeric regions fold in a unique conformation1.
The formation is facilitated by the N3 protonation of cytosine residues. i-motif DNA can used in many bio-nanotechnological applications2 and recently has been shown its significance in cancer treatment3.
We present microsecond range of MD simulations of a single stranded deprotonated DNA i-motif in ambient temperature and in high temperature, focusing primarily on thermodynamic and energetically favorable conformations.
Figure1 (L to R): Showing the ribbon and cartoon representation of folded structure of 22mer single stranded DNA and the observation of cytosine pairing in acidic solution. 11 22 1
References
[1] Guéron, M., Leroy, J.L, Current Opinion in Structural Biology 10(3), (2000)
[2] Liu, D., Balasubramian, S., Angewandte Chemie International 42(46),  (2003)
[3] Ren, J., Qu, X., Trent, J.O., Chaires, J.B. Nucleic Acids Research 30, 2307–2315 (2002)
Published results
[4] Smiatek, J.,Chen, C., Liu, D., Heuer, A. The Journal of Physical Chemistry B, 115, (2011)
[5] Smiatek, J., Liu, D., Heuer, Current Physical Chemistry, 2, (2012)
[6] Smiatek, J., Janssen-Mueller, D., Friederich, R., Heuer, A. Physica A, 394, (2014)
Work in progress
Currently, we are applying Bias-exchange metadynamics with the flavor of replica-exchange to study the i-motif unfolding.
Computational pulling experiments via Steered-MD in 1D and 2D free energy landscapes of unfolding mechanism.
Free energy landscape for crowed assemblies of DNA i-motif.
Studying the stability of DNA i-motif using nucleotide defects by mutating the sequence as well as the effects of hairpin length on structural stability and conformations.
Effects of sequence defects on i-motif
The probability matrix provides the vital information regarding the occurrence of i-motif, hairpin and/or extended hairpin conformation from the free energy landscapes of different defects of i-motif.
The tendency is the same but the effect in the high pH-regime is much weaker (if we calculate probability based on the ΔG values under acidic conditions, the effect would be much stronger) because then the i-motif is further stabilized by the H-bonds.
3C-2G
3C-1A
3C-1T
3C-2T
3C-7A
Figure 3 : Comparison of free energy landscapes to show the effect of induced alteration on i-motif structure. The defects were induced using the COM of rC1/T22, rC1,T22/A11
Probability
sequence
i-motif
hairpin
Ext hairpin
ΔG (kcal/mol)
0.09
0.07
0.84
-4.9
3C-7G
0.10
0.11
0.78
-2.2
3C-2G
0.05
0.86
-0.6
3C-1pDA
0.06
0.83 --
3C-1pDT
0.03
0.90