PICOSECOND TIME-RESOLVED FLUORESCENCE SPECTROSCOPY OF 13-DEMETHYLRETINAL BACTERIORHODOPSIN

The time-resolved fluorescence spectroscopy of the 13-demethylretinal artificial pigment of bacteriorhodopsin (13-dm BR) is measured with 6 ps time resolution over the initial 100 ns of its photocycle. Following 6 ps pump laser excitation (568-573 nm) to initiate the photocycle, fluorescence is generated with a 6 ps probe laser pulse (590-610 nm) appearing at time delays as long as 100 ns after the pumping. Fluorescence can be assigned to the excited electronic states of both 13-dm BR-560 and to the K-like photocycle intermediate (13-dm K) formed within 100 ps of 568-573 nm excitation. The intensity maximum of the fluorescence spectrum for 13-dm K is shifted to the blue by almost-equal-to 8 nm from that of 13-dm BR-560 and has a spectral profile which remains unchanged throughout the initial 10 ps-100 ns interval of the photocycle. The fluorescence intensity (I(f)) from 13-dm BR-560 is almost-equal-to 2 times larger than that from native BR-570 while I(f) from 13-dm K is > 6.5 times larger than the corresponding I(f) from K-590 in the native BR photocycle. In addition, approximately 26 ns is required for the fluorescence signal from 13-dm K to reach its maximum value which is almost-equal-to 9 times larger than the fluorescence signal from 13-dm BR-560. The significance of the transient fluorescence in 13-dm BR is discussed with respect to the molecular mechanism underlying the 13-dm BR photocycle.