Primary events in bacteriorhodopsin probed by subpicosecond spectroscopy

Abstract

The photochemical cycle of light-adapted bacteriorhodopsin at room temperature was initiated by 0.6-ps single pulses at 615 nm. In the spectral region from 410 to 750 nm the absorption difference spectra were measured. Three photoinduced states have been observed during the first 10 ps after excitation. The difference spectrum between the latest one and bacteriorhodopsin is similar to the difference spectrum between the earlier observed K intermediate (with nanosecond lifetime) and bacteriorhodopsin. The K intermediate is preceded by the primary photoproduct with 3-ps lifetime (the J intermediate). Both J and K have red-shifted spectra compared with the initial spectrum of bacteriorhodopsin. The earliest state has a lifetime of 0.7 ps, and a blue-shifted spectrum. We have designated this state as I 460 in accordance with its maximum in difference spectrum, and considered that I 460 is the electronically excited state of bacteriorhodopsin. The photoinduced dichroism measurements made are in good agreement with this consideration. The complete sequence of the primary photochemical events in bacteriorhodopsin can be written as: bacteriorhodopsin → hv bacteriorhodopsin∗(I 460) → 0.7 ps J → 3 ps K