LAUSR

| 7 Abstract As a chemotherapeutic agent heavily used since 1969, Doxorubicin’s (Doxo) mission is well known and carried out, in a variety of cancers. Doxo halts cancerous cell replication and causes cancerous cells to die through inducing cellular apoptosis. The necessary concentration for this to happen and the way in which these interactions are carried out at molecular level is still under debate. We use a dual beam optical tweezers to trap and isolate a single DNA molecule so that we can explore the binding of Doxo at the single molecule level. Stretching the DNA molecule in the presence of various concentrations of Doxo allow us to quantify the binding and better understand this complex interaction. Traditional stretch and release experiments in the presence of Doxo displayed that Doxo binding was not in equilibrium. To obtain the equilibrium binding properties, the DNA was stretched and held at a constant force, allowing the drug ample time to bind to the DNA. The equilibrium extensions of DNA upon binding to various concentrations of Doxo was obtained at four different constant forces. One dimensional lattice binding model, McGhee von Hippel Model, was used obtain the binding affinity at each force and extrapolated to obtain the binding affinity in the absence of force, Kd (0) = 1087 ± 187 nM. This very first characterization of Doxo binding to DNA at the single molecule level also yields the extension of the DNA upon a single intercalation event, ∆xeq = 0.40 ± 0.02 nm. The time scale of in which Doxo reaches binding equilibrium with DNA and the extension obtained upon single binding event challenges the well believed notion that Doxo is a simple classical intercalator. Understanding the binding mechanisms of Doxo to DNA at the single molecule level plays a key role to design new and improved cancer drugs for the future.