Resonance Raman spectroscopy of sensory rhodopsin II from Natronobacteriumpharaonis

Sensory rhodopsin II (pSRII), the photophobic receptor from Natronobacteriu m pharaonis, has been studied by time-resolved resonance Raman (RR) spectro scopy using the rotating cell technique. Upon excitation with low laser pow er, the RR spectra largely reflect the parent stale pSRII(500) whereas an i ncrease of the laser power leads to a substantial accumulation of long-live d intermediates contributing to the RR spectra, All RR spectra could consis tently be analysed in terms of four component spectra which were assigned t o 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 an d 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 th e Schiff base in pSRII(500) and O-535 whereas these interactions are most l ikely vera weak in N-485 (C) 2000 Federation of European Biochemical Societ ies.