Molecular Dynamics of a kB DNA Element: Base Flipping via Cross-strand Intercalative Stacking in a Microsecond-scale Simulation

📝 Abstract

The sequence-dependent structural variability and conformational dynamics of DNA play pivotal roles in many biological milieus, such as in the site-specific binding of transcription factors to target regulatory elements. To better understand DNA structure, function, and dynamics in general, and protein-DNA recognition in the ‘kB’ family of genetic regulatory elements in particular, we performed molecular dynamics simulations of a 20-base pair DNA encompassing a cognate kB site recognized by the proto-oncogenic ‘c-Rel’ subfamily of NF-kB transcription factors. Simulations of the kB DNA in explicit water were extended to microsecond duration, providing a broad, atomically-detailed glimpse into the structural and dynamical behavior of double helical DNA over many timescales. Of particular note, novel (and structurally plausible) conformations of DNA developed only at the long times sampled in this simulation – including a peculiar state arising at ~ 0.7 us and characterized by cross-strand intercalative stacking of nucleotides within a longitudinally-sheared base pair, followed (at ~ 1 us) by spontaneous base flipping of a neighboring thymine within the A-rich duplex. Results and predictions from the us-scale simulation include implications for a dynamical NF-kB recognition motif, and are amenable to testing and further exploration via specific experimental approaches that are suggested herein.

💡 Analysis

The sequence-dependent structural variability and conformational dynamics of DNA play pivotal roles in many biological milieus, such as in the site-specific binding of transcription factors to target regulatory elements. To better understand DNA structure, function, and dynamics in general, and protein-DNA recognition in the ‘kB’ family of genetic regulatory elements in particular, we performed molecular dynamics simulations of a 20-base pair DNA encompassing a cognate kB site recognized by the proto-oncogenic ‘c-Rel’ subfamily of NF-kB transcription factors. Simulations of the kB DNA in explicit water were extended to microsecond duration, providing a broad, atomically-detailed glimpse into the structural and dynamical behavior of double helical DNA over many timescales. Of particular note, novel (and structurally plausible) conformations of DNA developed only at the long times sampled in this simulation – including a peculiar state arising at ~ 0.7 us and characterized by cross-strand intercalative stacking of nucleotides within a longitudinally-sheared base pair, followed (at ~ 1 us) by spontaneous base flipping of a neighboring thymine within the A-rich duplex. Results and predictions from the us-scale simulation include implications for a dynamical NF-kB recognition motif, and are amenable to testing and further exploration via specific experimental approaches that are suggested herein.

📄 Content

Molecular dynamics of a κB DNA element: Base flipping via cross-strand intercalative stacking in a microsecond- scale simulation

Cameron Mura1,*,† & J. Andrew McCammon1,2

Author affiliations & correspondence: 1 Department of Chemistry & Biochemistry and Center for Theoretical Biological Physics; University of California, San Diego; La Jolla, CA 92093-0365 2 Howard Hughes Medical Institute and Department of Pharmacology;
University of California, San Diego; La Jolla, CA 92093-0636

• To whom correspondence should be addressed: Tel: 1.434.924.7824 Fax: 1.434.924.3710 Email: cmura@virginia.edu † Current address: Department of Chemistry; University of Virginia; Charlottesville, VA 22904-4319

Manuscript information: Synopsis: Microsecond-scale molecular dynamics simulations of DNA reveal unanticipated conformational features, with implications for site-specific binding of NF-κB transcription factors to regulatory κB DNA elements, as well as spontaneous base flipping via a mechanism of cross-strand intercalative stacking of nucleotides. Format: Nucleic Acids Research article (in press) Running title: Molecular dynamics of κB DNA Last modified: 02 July 2008 Length estimate: 12.5 printed pages (using ‘words/925 + figures/2.8’; excluding references) Additional notes: This manuscript is accompanied by nine figures and thirteen items of supplementary information (nine figures and four video animations).

Molecular dynamics of κB DNA Mura & McCammon

2

Abstract

The sequence-dependent structural variability and conformational dynamics of DNA play pivotal roles in many biological milieus, such as in the site-specific binding of transcription factors to target regulatory elements. To better understand DNA structure, function, and dynamics in general, and protein···DNA recognition in the “κB” family of genetic regulatory elements in particular, we performed molecular dynamics simulations of a 20-base pair DNA encompassing a cognate κB site recognized by the proto-oncogenic “c-Rel” subfamily of NF-κB transcription factors. Simulations of the κB DNA in explicit water were extended to microsecond duration, providing a broad, atomically-detailed glimpse into the structural and dynamical behavior of double helical DNA over many timescales. Of particular note, novel (and structurally plausible) conformations of DNA developed only at the long times sampled in this simulation – including a peculiar state arising at ≈ 0.7 μs and characterized by cross-strand intercalative stacking of nucleotides within a longitudinally-sheared base pair, followed (at ≈ 1 μs) by spontaneous base flipping of a neighboring thymine within the A-rich duplex. Results and predictions from the μs- scale simulation include implications for a dynamical NF-κB recognition motif, and are amenable to testing and further exploration via specific experimental approaches that are suggested herein.

Introduction

DNA is often viewed as a relatively rigid biological macromolecule (1-3), with RNA and proteins thought of as exhibiting broader ranges of both intrinsic three-dimensional structural variability as well as dynamical flexibility. This perspective of a locally-rigid, globally-flexible biopolymer is consistent with the rather passive biological role of DNA as the repository of genetic information – the genome is read- out by the process of transcription. Links between structure and potential biological functions (both normal and aberrant) have been explored for conformations that deviate from the standard B-form double helix – including such varieties as multi-stranded triplexes, quadruplex structures found in telomeric G- rich tracts, cruciforms adopted by inverted repeats, hairpins and slipped structures, and so on (4). However, beyond these alternative secondary structures, it is also becoming increasingly apparent that the structure and dynamics of the canonical Watson-Crick DNA double helix on a very local (base pair) level play pivotal roles in specific biological functions, such as the site-specific binding of transcription factors to target DNA elements. This idea of a functional role for sequence-specific DNA fine structure and dynamics is embodied in the concept of “indirect readout,” (5,6) wherein features of protein···DNA recognition are dictated by subtle conformational and dynamical properties of DNA beyond the stereochemical code provided by the specific linear array of chemical functionalities that line the major and minor grooves for a given nucleotide sequence.

Despite the vast literature dedicated to DNA structural biology since the first atomic-resolution crystal structures of both left- (7) and right-handed (8) double helices, many aspects of DNA structure and dynamics remain unclear – including the intrinsic coupling between structure and conformational dynamics that is the basis of indirect readout. For instance, controversy surrounds the

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