Absorption spectra of intermediates of bacteriorhodopsin measured by laser photolysis at room temperatures |
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Authors: | Yoshinori Shichida Sinzi Matuoka Yusuke Hidaka Toru Yoshizawa |
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Institution: | Department of Biophysics, Faculty of Science, Kyoto University, Kyoto Japan |
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Abstract: | Picosecond laser spectroscopic analysis was applied to determine how many intermediates existed in the primary photochemical process of trans-bacteriorhodopsin (light-adapted bacteriorhodopsin) at room temperature (18°C) and to calculate their absorption spectra. Irradiation of bacteriorhodopsin with a laser pulse (wavelength, 532 nm; pulse width, 25 ps) yielded the K intermediate (K) which was produced through a precursor, having an absorption maximum (λmax) longer than that of K. K was stable during a picosecond time range (50–900 ps). The λmax was located at 610 nm and the extinction coefficient (?max) was 0.92-times that of bacteriorhodopsin. The same K intermediate was produced from bacteriorhodopsin even when it was excited with a high-energy pulse by which a saturation effect was induced. A transient difference spectrum measured at 150 ns after the excitation of bacteriorhodopsin was different in shape from that of the K intermediate, suggesting that an intermediate was formed by thermal decay of K. This intermediate, tentatively called the KL intermediate (KL), had a λmax at 596 nm and an ?max 0.80-times that of bacteriorhodopsin. KL decayed to the L intermediate (L) with a time constant of 2.2 μs. L has a λmax at 543 nm and an ?max 0.66-times that of bacteriorhodopsin. |
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Keywords: | Bacteriorhodopsin Absorption spectrum Laser photolysis (Purple membrane) |
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