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 stale pSRII(500) 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 pSRII(500) and the long-lived intermediates M-400, N-485 and O-535 based on the correlation between the C=C stretching frequency and the absorption maximum, The parent state and the intermediates N-485 and O-535 exhibit a protonated Schiff base. The C=N stretching frequencies and the MID isotopic shifts indicate strong hydrogen bonding interactions of the Schiff base in pSRII(500) and O-535 whereas these interactions are most likely vera weak in N-485 (C) 2000 Federation of European Biochemical Societies.