NANOSECOND TIME-RESOLVED INFRARED-SPECTROSCOPY DISTINGUISHES 2 K-SPECIES IN THE BACTERIORHODOPSIN PHOTOCYCLE

The photochemical reaction process of bacteriorhodopsin in the nanosec ond time range (-120 similar to 860 ns) was measured in the 1400 simil ar to 900 cm(-1) region with an improved time resolved dispersive-type infrared spectrometer. The system is equipped with a newly developed detection unit whose instrumental response to a 5-ns laser pulse has a full width of the half-maximum of 60 ns. It provides highly accurate data that enabled us to extract a kinetic process one order of magnitu de faster than the instrumental response. The spectral changes in the 1400 similar to 900 cm(-1) region were analyzed by singular value deco mposition and resolved into three components. These components were se parated by fitting with 10- and 1000-ns exponential functions and a st ep function, which were convoluted with the instrumental response func tion. The components with decay time constants of 10 and 1000 ns are n amed K and KL, respectively, on the basis of previous visible spectros copy. The spectral shapes of K and KL are distinguishable by their hyd rogen-out-of-plane (HOOP) modes, at 958 and 984 cm(-1), respectively. The former corresponds to the K intermediate recorded at 77 K and the latter to a K-like photoproduct at 135 K. On the basis of published da ta, these bands are assigned to the 15-HOOP mode, indicating that the K and KL differ in a twist around the C-14-C-15 bond.