Ricerc@Sapienza

Channelrhodopsin (ChR) is a fundamental transmebrane protein (TMP) that can act as light-sensitive ion channel through the cell membrane. This protein performs its function through a series of conformational changes triggered by the absorption of visible light. In the last decade, ChR has been widely studied for its possible applications in the field of optogenetics but the functional mechanisms related to its function is still partly unknown.
Mid-Infrared (IR) spectroscopy is considered a powerful label-free tool to investigate the modifications of light-sensitive TMPs structure. However, the sensitivity of mid-IR spectroscopy, in terms of probed molecules, requires to be applied to large amount of purified proteins in order to probe the subtle light-induced modifications of protein IR absorption spectrum. In order to study the more biologically interesting case of proteins embedded in native cell membranes, which are intrinsically heterogeneous, it is necessary to push IR spectroscopy capabilities at the nanoscale exploiting plasmonic approaches, such as in the novel IR platform based on an Atomic Force Microscopy (AFM) coupled to a quantum cascade laser. This technique allows one to combine spectroscopic information with nanometric resolution provided by the scanning probe approach (AFM).
Despite IR nanospectroscopy has been largely applied to protein samples, the first application of this approach to the investigation of the light-sensitive TMPs has been demonstrated only recently with experiments carried out during my Master degree and first year of PhD project. I have investigated the light-induced activity of Bacteriorhodopsin, a prototype for light-sensitive TMPs, at the single cell membrane monolayer. These results pave the way towards the investigation at the nanoscale of more interesting light-sensitive optogenetic gates, such as ChR, in order to better understand the functional mechanisms of these proteins when embedded in their native cell membrane.