Analyzing the relationship between single base flipping and strand slippage near DNA duplex termini

Insertion-deletion (indel) mutations are caused by strand slippage between pairing primer and template strands during nucleic acid strand extension. A possible causative factor for such strand slippage is base flipping in the primer strand or template strand, for insertion or deletion mutations, respectively. A simple mechanistic description is that the "hole" in the nucleic acid duplex left behind by a flipping base is occupied by a neighboring base on the same strand, resulting in slippage with respect to its paired strand. The extent of single base flipping required for occupation of its former place in the double helix by a neighboring base is not fully understood. The present study uses restrained molecular dynamics (MD) simulations along a pseudohedihedral base flipping parameter to construct two-dimensional free energy profiles along base flipping and strand slippage geometric parameters. These profiles, generated for both cytosine and guanine single base flipping in a short repetitive indel mutation hot-spot DNA sequence, illustrate the extent of single base flipping that can allow strand slippage by one base position. Relatively minor base flipping into both the major and minor grooves can result in strand slippage. Deconstruction of the collective variable strand slippage geometric parameter into its component distances illustrates the details of how strand slippage can accompany base flipping. The trans Watson-Crick:sugar edge interaction that stabilizes cytosine flipping in this hot-spot sequence is also characterized energetically. The impact of these results on understanding sequence dependence of indel errors in nucleic acid strand extension is discussed, along with a suggestion for future studies that can generalize the present findings to all nearest-neighbor sequence contexts.