Investigating Enzyme Kinetics Using Nanofluidic Devices

We have developed a nanofluidic device consisting of two parallel electrodes in a nanometer-scale cavity with openings at both ends. Electrochemically active molecules that enter the cavity transfer charge between the electrodes when a potential difference is applied. Typically, electrochemical reactions only involve one or a few electrons per molecule, rendering direct detection of a single molecule in solution virtually impossible. This limitation is overcome, in this situation, by redox cycling, where each molecule can transfer, on average, thousands of electrons by repeatedly traveling between the electrodes before escaping back out into the bulk. Further, the devices are created using standard microfabrication technology, and can therefore be produced reliably in large quantities.

Numerous sensing applications can be envisioned with such nanofluidic devices, ranging from water purity to drug screening. This talk will focus on employing these devices for determining enzyme kinetics. The reaction kinetics can be monitored directly by the device if the enzyme is capable of converting an electrochemically inactive substrate into an active product. Since the nanochannel is open to the bulk, the composition of the solution inside the device is a representative sample of the bulk solution. Therefore, as the concentration of product in the bulk solution increases, the average number of product molecules in the nanochannel also goes up leading to a larger signal. Our model enzyme is tyrosinase, which converts electrochemically inactive monophenols into active diphenols and quinones.