Gaoxiang Mei Defense:Spectroscopy Study of Optogenetic Rhodopsins- Improved Opsin-Based Transmembrane Voltage Sensors

Optogenetics is a technique used by researchers to control or monitor living cells like neurons with light. The most commonly used optogenetic rhodopsin in monitoring of neuronal activity were opsin transmembrane voltage sensors(OTVS) like QuasAr1, QuasAr2 and NovArch. The optical properties of these proteins like fluorescence could be altered by the membrane potential. Therefore, researchers could map the neuronal action potentials into fluorescence. This method successfully enables us to monitor the membrane potential with sub-millisecond temporal resolution and sub-cellular spatial resolution. Despite these advances, effective in vivo optogenetic monitoring and control of neural activity using microbial rhodopsins is in large part severely limited because: 1) biological tissues strongly absorb and scatter visible light, and 2) until recently the absorbance bands of all known microbial rhodopsins and mutants did not extend into the NIR region. Thus one of the major focuses of our research in the Molecular Biophysics Lab (MBL) at Boston University is to understand on an atomic level the molecular mechanisms of OTVSs and what determines the analog retinal redshifts. To achieve this goal, we use UV/Vis spectroscopy, fluorescence spectroscopy, resonance Raman spectroscopy (RRS), and Fourier-transform Raman spectroscopy(FT-Raman) in addition to biochemical techniques, such as site-directed mutagenesis and analog retinal substitution. In my defense today, a review will be presented with special focus on earlier work by our group on the molecular mechanism of GPR and AR3 mutants.