The International Research Fellowship Program enables U.S. scientists and engineers to conduct nine to twenty-four months of research abroad. The program's awards provide opportunities for joint research, and the use of unique or complementary facilities, expertise and experimental conditions abroad.
This award will support a twenty-four month research fellowship by Dr. Edgar D. Goluch to work with Dr. Serge Lemay at Delft University in the Netherlands.
The goal of the proposal is to develop a new technique that electrochemically detects the activity of a single enzyme in real time by providing a signal each time a product molecule is generated. The approach demonstrates a new generation of electrochemical biosensors with single molecule detection sensitivity that will be employed to study the internal dynamics of individual enzyme molecules. This technique uses a microfabricated device that consists of two electrodes in a glass channel that are separated by about 10 nanometers. The electrodes detect electrochemically active molecules that are located in the liquid-filled volume between them. Catalysis is studied by placing a single enzyme that can convert inactive substrates into redox-active products between the electrodes. A reversibly redox-active molecule can rapidly gain and lose electrons when exposed to an electrical potential. While a single electron transfer event is very difficult to detect, the nanoscale gap allows millions of electrons to be transported by a redox-active molecule every second, thus producing a measurable amount of electrical current. The dramatic amplification of the electrochemical signals provided by this detection scheme allows the construction of electrochemical biosensors with unprecedented sensitivity and selectivity. Tyrosinase, which catalyzes the conversion of phenols into redox-active quinone molecules, is used as the model system. Of particular interest are the structural changes that the enzyme undergoes over time and their effect on enzyme activity, as well as the effects of external factors, such as pH, temperature, and buffer concentration. Finally, the influence of dramatic fluctuations in substrate concentration generated by the minute fluid volume of the system is being investigated.
