The Effects of Ionic Strength on the Hydrodynamic Properties of I-Motif Folding

Abstract

Cytosine-rich nucleic acid sequences found in human DNA can adopt multiple intramolecular structures identified as i-motifs that are dependent on physiochemical solution conditions. The focus of this study is a four-stranded structure from the promoter of the human c-MYC gene. This compact, stable, and monomeric structure forms upon a decrease in pH (<5.0) causing hemi-protonation of a cytosine that results in a stable C-C+ hydrogen bond. Sedimentation velocity experiments were performed with the analytical ultracentrifuge in order to determine the hydrodynamic properties of the folded i-motif. The sedimentation velocity experiments were carried out in buffer conditions that differed in pH (4.5-8.0), salt type (NaCl and KCl), and salt concentration (up to 400 mM). Experiments were run at different pH values in order to observe the linkage between the pKa of the Hoogsteen base pair and i-motif folding. High salt concentration was used to avoid non-ideality (primary charge effect) observed when nucleic acids sediment at low salt concentration. The data indicates that the S20,w value increases when the primary charge effect is overcome at a higher salt concentration. The experimental S20,w values are compared with values obtained by bead model simulations using SOMO as implemented in Ultrascan 3. (Supported by UMC AUC Facility.)