Protein Dynamics in the Bacteriorhodopsin Photocycle: A Nanosecond Step-Scan FTIR Investigation of the KL to L Transition

A time-resolved step-scan FTIR spectrometer with a time resolution of 20 ns was developed and used to investigate the KL to L transition in the photocycle of bacteriorhodopsin in the time range from −60 to 940 ns. Broadband FTIR absorbance difference spectra with a spectral range of 850−2050 cm-1 and a spectral resolution of 4 cm-1 have been obtained. Our data show that there are two sets of BR photoproduct difference bands exhibiting different kinetics. The intensity changes of bands attributed to structural changes of the carboxyl groups Asp-96 and Asp-115 as well as of bands assigned to CC and CC stretching modes of the chromophore show single exponential behavior with a time constant of about 400 ns, implying that these two processes are coupled. The time-dependent intensity changes of bands attributed to structural changes of the chromophore region near the Schiff base exhibit slower kinetics with a time constant of about 2 μs. We interpret our data in terms of a process during the KL to L transition...