Team:Slovenia/Mechanosensing/CaDependent mediator
nbsp;Calcium-depended mediator
A new split calcium sensing/reporting system based on split firefly luciferase linked to M13 and calmodulin was designed that is able to report the increase of the cytosolic calcium ions induced by mechanoreceptor stimulation by emitted light.
Motivation
While calcium influx could be detected by exogenous fluorescent dyes such as the FuraRed and Fluo-4, we needed a genetically encoded calcium sensor that would couple a change in the calcium concentration to a biologically relevant output, such as the luciferase reporter or reconstitution of a split protease the initiation of the signaling pathway. The ideal calcium sensor should be inactive at intracellular concentration of calcium and have a high response to calcium concentrations above physiological levels and should be detected by a quick and easy readout. For our intended application, the calcium sensor should also have the potential to act as the reconstitution mechanism for split proteins so that a conformational rearrangement in the presence of calcium would bring the two split protein fragments together and reconstitute the protein’s activity.
