Analysis of Drug-DNA Interactions On Solid Supports

Analysis of Drug-DNA Interactions on Solid Supports.

Due to the increasing interest in anticancer, antibiotic and antiviral drug design, nucleic acid-drug interactions present the focus of studies of many researchers. Observing such interactions on the surface provides the benefit of material and cost-efficient technology that is feasible for high-throughput assays. Electrochemical approaches moreover provide easy integration with microelectronic technology. The experimental setup is simple and affordable compared to equipment that would be necessary for comparably high-throughput methods based on solution protocols.

In this research electrochemical techniques were applied for analysis of drug-DNA interactions via measuring effect of drug binding on the equilibrium melting transition of double-stranded DNA tethered to a surface. First, single-stranded DNA “probes” were immobilized on the surface via self-assembly. Next, complementary "DNA target" species, labeled with an electrochemical tag with reversible redox activity, were added to solution in which the probe layer is immersed. The ensuing target-probe hybridization can be monitored through appearance of the label oxidation/reduction current when the probe layer is electrochemically interrogated with cyclic voltammetry. Moreover, by quantitatively analyzing charge passed due to cycling of the label's oxidation state, the coverage of duplexes on the surface can be precisely determined. When such measurements are performed as a function of temperature the DNA duplex hybridization and melting transitions are traced out, corresponding to electrochemical acquisition of equilibrium DNA surface melting profiles.

Upon addition of the minor groove-binding drug netropsin, the melting curves are observed to shift to higher temperatures, indicating stabilization of the duplex state by this small molecule, oligopeptide ligand. These changes in the melting transition can be analyzed to derive thermodynamic parameters of netropsin binding to its cognate DNA site, and to evaluate the effects of ionic strength and the oligonucleotide sequence on the drug binding affinity.