Understanding the interaction between drug molecules and nucleic acids is an important factor in the design of new drugs. The thermodynamics of this interaction, such as binding affinity, binding free energy and enthalpic/entropic changes, have been determined to date using bulk methods including calorimetry techniques. In addition, single-molecule approaches have also been devised, which are of particular interest as they can provide detailed information on the mechanical and elastic properties of double-stranded DNA molecules bound to small molecules.

Now, Ian Hsu and colleagues at the National Tsing Hua University in Taiwan1 have developed a new single-molecule approach that can accurately predict the binding affinity constant, and hence determine the free-energy change, of a model drug–DNA interaction.

Fig. 1: Schematic illustration of the single-molecule approach, showing the technetium-doped intercalated molecule that can insert itself between Watson–Crick base pairs. The two-trap optical tweezers system has sensors at either side that determine the tension of the DNA molecule.

The method uses a two-trap optical tweezers system in which a DNA strand with a bead at each end is suspended between the traps (Fig. 1). A mono-intercalating molecule (termed APMED) based on a pyrene structure was used as the model drug molecule. The pyrene unit inserted itself between Watson–Crick base pairs and, on intercalation, changed the tension, and hence length, of the DNA strand. This change in length was measured by the optical traps and by taking measurements at different concentrations of APMED and using a wormlike chain model, the researchers were able to determine the binding affinity constant and free-energy change.

“Compared with the existing single-molecule approach, which uses extrapolation from nonzero force data, we provide a direct zero-force measurement of binding energetics and the results are more consistent with other existing data,” says Hsu.

The pyrene-based molecule can be adapted to incorporate a technetium radionuclide (99mTc) — an Auger electron emitter. In future studies, Hsu envisages that this method will be able to answer some fundamental questions in targeted radiotherapy2. “For example, we could determine how many Auger electron emitters are attached to the target DNA and how rapidly the DNA double-strand breaks occur as a function of Auger electron emitter radioactivity,” says Hsu. “This knowledge should allow us to propose a better benchmark measuring method to extract DNA double-strand breaks in the presence of 99mTc.”