A Biomolecular Photodiode for Imaging of Cell Membrane Potential

Abstract

Despite the recognized importance of electrical signals in certain biological systems, there has been limited success in the creation of a reliable voltage sensor for imaging of such activity. Using standard molecular biology techniques, we have created a biomolecular photodiode consisting of a membrane-bound cytochrome c protein fused with a GFP (green fluorescent protein) variant. A similar photodiode assembly has been shown to produce unidirectional photocurrent in vitro with the cytochrome acting as an acceptor of excited state electrons from the FP donor upon excitation with visible light. Electron transfer between the cytochrome and the FP is a highly voltage dependent process. By embedding this assembly in the plasma membrane of living cells, it is subjected to the same electric potential as the membrane. As the membrane potential of the cell changes, as in an action potential, the extent of electron transfer is expected to vary significantly, resulting in a change in fluorescence intensity of the FP donor. As this is a very fast process with high sensitivity to changes in electric potential, the biophotodiode has potential to form a robust sensor of electrical activity in cells. The feasibility of the sensor is investigated in several ways, including modeling, electrophysiology, and direct application of current to purified membrane fragments.