Kandori H, Shichida Y, Yoshizawa T
Department of Biophysics, Faculty of Science, Kyoto University, Japan.
Biophys J. 1989 Sep;56(3):453-7. doi: 10.1016/S0006-3495(89)82692-X.
Picosecond laser photolysis of rhodopsin in 15% polyacrylamide gel was performed for estimating absolute absorption spectra of the primary intermediates of cattle rhodopsin (bathorhodopsin and photorhodopsin). Using a rhodopsin digitonin extract embedded in 15% polyacrylamide gel, a precise percentage of bleaching of rhodopsin after excitation of a picosecond laser pulse was measured. Using this value, the absolute absorption spectrum of bathorhodopsin was calculated from the spectral change before and 1 ns after the picosecond laser excitation (corresponding to the difference spectrum between rhodopsin and bathorhodopsin). The absorption spectrum of bathorhodopsin thus obtained displayed a lambda max at 535 nm, which was shorter than that at low temperature (543 nm) and a half band-width broader than that measured at low temperature. The oscillator strength of bathorhodopsin at room temperature was smaller than that at low temperature. The absolute absorption spectrum of photorhodopsin was also estimated from the difference spectrum measured at 15 ps after the excitation of rhodopsin (Shichida, Y., S. Matuoka, and T. Yoshizawa. 1984. Photobiochem. Photobiophys. 7:221-228), assuming a sequential conversion of photorhodopsin to bathorhodopsin. Its lambda max was located at approximately 570 nm, and the oscillator strength was smaller than those of rhodopsin and bathorhodopsin.
为了估算牛视紫红质（视紫红质中间体和光视紫红质）初级中间体的绝对吸收光谱，对15%聚丙烯酰胺凝胶中的视紫红质进行了皮秒激光光解。使用嵌入15%聚丙烯酰胺凝胶中的视紫红质洋地黄皂苷提取物，测量了皮秒激光脉冲激发后视紫红质漂白的精确百分比。利用该值，根据皮秒激光激发前和激发后1纳秒的光谱变化（对应于视紫红质和视紫红质中间体之间的差光谱）计算出视紫红质中间体的绝对吸收光谱。由此获得的视紫红质中间体的吸收光谱在535纳米处显示出最大吸收波长，该波长比低温下的波长（543纳米）短，半峰宽比低温下测量的宽。室温下视紫红质中间体的振子强度比低温下小。假设光视紫红质依次转化为视紫红质中间体，还根据视紫红质激发后15皮秒测量的差光谱估算了光视紫红质的绝对吸收光谱（Shichida, Y., S. Matuoka, and T. Yoshizawa. 1984. Photobiochem. Photobiophys. 7:221 - 228）。其最大吸收波长位于约570纳米处，振子强度比视紫红质和视紫红质中间体的小。
