Abstract
To obtain high quality time-resolved ultraviolet resonance Raman (UVRR) spectra of transient intermediates in the bacteriorhodopsin (BR) photocycle, we developed a new UVRR spectrometer. A home-made F=3.5 prism prefilter was used in front of a 50 cm CCD detected spectrograph to give high throughput, wide tunability, and excellent stray light rejection along with low dispersion. Using this system, we obtained 239.5 nm excited time-resolved UVRR spectra of BR which revealed small but significant features associated with the formation of the KL-intermediate at 10 ns delays. This difference spectrum exhibits intensity decreases at 1624, 1561, 1012 and 763cm-1 due to an altered environment of one or more Trp residues and a frequency shift of the Tyr ʋ8b band at 1602 cm-1. These signals show that the photoisomerization of retinal from all-trans to 13-cis induces significant changes in the structure and environment of aromatic residues that line the retinal binding pocket in only 10 ns.