(558b) Genetically-Encoded Biosensors of Cellular Metabolites in Plants
The discovery of fluorescent proteins, such as GFP or BFP, gave researchers the ability to visualize organelles and proteins within cells. Genetically-encoded biosensors, some based on GFP-like proteins, allow the non-invasive, non-destructive monitoring of cellular metabolites at different points in the cell cycle. They offer the opportunity to better understand cell signaling pathways and to monitor in vivo responses to various external forces. Additionally, since sensing occurs upon binding of a ligand, the sensitivity and specificity of sensors can be modulated via mutation.
This talk will focus on two types of protein-based biosensors. FRET-based sensors are a three protein fusion with two fluorescent proteins flanking a ligand binding domain. Upon binding of the ligand, a confirmational change in the receptor region brings the fluorescent proteins into the appropriate configuration for energy transfer, leading to a signal. Alternatively, a binding region can be directly inserted into a single fluorescent protein, as is the case with cameleon GFP calcium sensors. We have explored the use of periplasmic binding proteins, such as maltose binding protein, estrogen receptors, and the auxin binding protein in both FRET and cameleon designs. The advantages and disadvantages of each type of protein-based sensor will be discussed in the context of monitoring metabolite concentrations in living plants.