Unfolding mechanism and the free energy landscape of a single stranded DNA i-motif

arXiv:1105.3894v1 [physics.bio-ph] 19 May 2011 Unfolding mechanism and the free energy landscape of a single stranded DNA i-motif Jens Smiatek1, Chun Chen2, Dongsheng Liu2, and Andreas Heuer1 1Institute of Physical Chemistry, University of Muenster, D-48149 M¨unster, Germany 2Department of Chemistry, Tsinghua University, Beijing 100190, P. R. China (Dated: November 8, 2018) We present Molecular Dynamics simulations of a single stranded unprotonated DNA i-motif in explicit solvent. Our results indicate that the native structure in non-acidic solution at 300 K is unstable and completely vanishes on a time scale up to 10 ns. Two unfolding mechanisms with decreasing connectivity between the initially interacting nucleobases can be identified where one pathway is characterized as entropically more favorable. The entropic preference can be mainly explained by strong water ordering effects due to hydrogen bonds for several occurring structures along the pathways. Finally we are able to indicate via free energy calculations the most stable configurations belonging to distinct hairpin structures in good agreement to experimental results. Keywords: DNA i-motif, Molecular Dynamics simulations, unfolding mechanisms, free energy landscape INTRODUCTION The appearance of non Watson-Crick like structures in DNA has been reported two decades ago [1]. Since this time a lot of effort has been spent to investigate these conformations and possible applications in detail [2–5]. Experiments lead to the conclusion that these structures are the only known DNA configurations that involve sys- tematic base intercalation [2]. Prominent representatives are the G-quadruplex structures and the i-motif [5] where the first one is formed by guanine (G) rich sequences [3] while the latter is present in more cytosine (C) rich strands of DNA [2]. The stabilizing mechanism for these at a first glance frag- ile structures is realized by a proton mediated cytosine binding between different strands or regions of the se- quence resulting in a stable C-CH+ pairing [1, 2, 4, 5]. Due to an acidic environment, this is achieved by hemi- protonated cytosines which mimick an ordinary C-G binding as it is present in double helix DNA. Hence it becomes clear that these structures are only occurring at slightly acidic to neutral conditions resulting in pH values from 4.8 to 7.0 [1, 2, 6]. I-motifs show a remarkable sta- bility [6] and have been found as tetrameric and dimeric complexes although their existence has also been proven for single stranded DNA [2]. A sketch of the C-CH+ complex where the additional proton mediates a hydro- gen bond between the nitrogens of the cytosine groups and the corresponding single stranded i-motif with its sequence is shown in Fig. 1. Due to its biological appearance in centromeric and telomeric DNA, the distinct i-motif conformations have been discussed as a new class of possible biological targets for cancers and other diseases [7, 8]. However, a detailed investigation of the function in the human cell is still missing. Despite this lack of knowledge, the application of this special configuration in modern biotechnology has experienced an enormous growth over the last years [4]. Since the i-motif becomes unstable at pH values larger than 7, a systematic decrease and increase of protons in the solution by changing the pH value results in a re- versible folding and unfolding mechanism. It has been shown that this process occurs on a timescale of seconds [4, 6]. Technological applications for this mechanism are given by molecular nanomachines [4, 9], switchable nanocon- tainers [10], pH sensors to detect the pH value inside liv- ing cells [11], building materials for logic gate devices [12] and sensors for distinguishing single walled and multi- walled carbon nanotube systems [13]. Recently it has been reported [14], that the grafting density massively influences the structure of an i-motif layer in nanode- vices due to steric hindrance. Regarding these examples it becomes clear that a detailed investigation of the un- folding pathway of the i-motif is of prior importance. In this paper we present the results of Molecular Dy- namics simulations concerning the unfolding mechanism of a maximum unstable single stranded DNA i-motif structure without hemi-protonated cytosines. Our re- sults indicate a fast initial decay of the i-motif leading to hairpin structures on a timescale up to 10 ns which dominate the unfolded regime in contrast to a fully ex- tended strand. The numerical findings are validated by experimental Circular Dichroism (CD) spectropolarime- try data. By distinct investigation of the unfolding path- ways, two main mechanisms can be identified which sig- nificantly differ in their entropic properties. We are able to separate the contributions of the solvent and the chain configurational entropy explicitly to determine the influ- ence on the unfolding pathways. The experimental re- sults can be explained by a temperature dependen

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