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Influence of divalent cations on the conformation of phosphorothioate oligodeoxynucleotides: a circular dichroism
1Department of Pharmaceutical Sciences, The University of Connecticut, Storrs, CT 06269-2092, USA.
Abstract:
Phosphorothioate oligodeoxynucleotides (ODNs) have been extensively investigated in vivo and in vitro for antisense control of gene expression. It has been shown that cellular uptake of phosphorothioate ODNs in some in vitro cell systems increases in the presence of divalent cations. In this work, we analyze the conformation of phosphorothioate ODNs and specific changes induced in it by various divalent cations using circular dichroism (CD) spectroscopy. CD data were obtained with several phosphorothioate ODNs in the absence and presence of the divalent cations Mg(2+), Ca(2+), Sr(2+), Ba(2+) and Mn(2+). All CD spectra indicated stable conformations of the ODNs in solution. The spectra were strongly dependent on ODN sequence and composition. Some ODNs such as T(23) and another with 'random' distribution of bases showed CD spectra characteristic of B-form DNA. Other ODNs which had at least three consecutive guanines in their sequences exhibited spectra characteristic of parallel G-tetraplexes. CD spectra of antisense ODNs exhibited specific responses to divalent cations. Changes in the conformation were not simply due to ionic strength effects. Mn(2+) diminished secondary structure in some ODNs. Group II divalent ions stabilized the parallel G-tetraplexes, and Mg(2+) generally had the weakest stabilizing efficiency. Each sequence/ion combination had a specific response so these effects cannot be generalized. These sequence-dependent, divalent ion-sensitive, and structurally unique solution conformations may be related to ion-mediated ODN uptake.
Insights
Phosphorothioate oligodeoxynucleotides (ODNs) adopt stable conformations influenced by their sequence and divalent cations. These cations, like Mn(2+) and Mg(2+), specifically alter ODN structures, potentially affecting cellular uptake.
Area of Science:
- Biochemistry
- Molecular Biology
- Spectroscopy
Background:
- Phosphorothioate oligodeoxynucleotides (ODNs) are crucial for antisense gene expression control.
- Cellular uptake of these ODNs can be enhanced by divalent cations in vitro.
- Understanding ODN conformation is key to optimizing their therapeutic applications.
Purpose of the Study:
- To investigate the solution conformations of phosphorothioate ODNs.
- To analyze how various divalent cations (Mg(2+), Ca(2+), Sr(2+), Ba(2+), Mn(2+)) induce conformational changes.
- To explore the relationship between ODN structure, cation interaction, and potential effects on cellular uptake.
Main Methods:
- Circular dichroism (CD) spectroscopy was employed to study ODN conformations.
- CD spectra were recorded for multiple phosphorothioate ODNs in the presence and absence of different divalent cations.
- Analysis focused on sequence-dependent structural changes and cation-specific effects.
Main Results:
- ODN conformations were stable in solution and highly dependent on sequence and composition.
- Specific sequences adopted B-form DNA or parallel G-tetraplex structures.
- Divalent cations induced specific conformational changes, not solely due to ionic strength; Mn(2+) destabilized some structures, while Group II ions stabilized G-tetraplexes, with Mg(2+) showing weaker stabilization.
Conclusions:
- Phosphorothioate ODNs exhibit diverse, sequence-specific conformations in solution.
- Divalent cations modulate these conformations in a sequence-dependent manner.
- These unique, ion-sensitive structures may play a role in ion-mediated ODN cellular uptake.