Resonance Raman spectroscopy of Sensory Rhodopsin II from Natronobacterium Pharaonis

Sensory rhodopsin II (pSRII), the photophobic receptor from Natronobacterium pharaonis, has been studied by time-resolved resonance Raman (RR) spectroscopy using the rotating cell technique. Upon excitation with low laser power, the RR spectra largely reflect the parent state pSRII500 whereas an increase of the laser power leads to a substantial accumulation of long-lived intermediates contributing to the RR spectra. All RR spectra could consistently be analysed in terms of four component spectra which were assigned to the parent state pSRII500 and the long-lived intermediates M400, N485 and O535 based on the correlation between the CNC stretching frequency and the absorption maximum. The parent state and the intermediates N485 and O535 exhibit a protonated Schiff base. The C=N stretching frequencies and the H/D isotopic shifts indicate strong hydrogen bonding interactions of the Schiff base in pSRII500 and O535 whereas these interactions are most likely very weak in N485