Retinal isomer ratio in dark-adapted purple membrane and bacteriorhodopsin monomers.
Retinal isomer ratio in dark-adapted purple membrane and bacteriorhodopsin monomers.
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DOI：
10.1021/bi00428a063
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发表时间：
1989-01
期刊：
影响因子：
2.9
通讯作者：
P. Scherrer;M. Mathew;W. Sperling;W. Stoeckenius
中科院分区：
文献类型：
--
作者：
P. Scherrer;M. Mathew;W. Sperling;W. Stoeckenius
根据对视网膜提取物进行光谱分析和色谱分析得到的数据，人们一致认为暗适应紫膜（pm）含有等量的13-顺式和全反式视网膜，而光适应紫膜只含有全反式视网膜。我们已经开发了一种改进的提取技术，可以在4分钟内提取高达70%的视网膜。在室温下适应黑暗的pm提取物中，我们一致发现66-67%的13-顺式视网膜和33-34%的全反式视网膜，并且在适应光的样品中发现超过98.5%的全反式异构体。用13-顺式视网膜在2℃（以减少异构化）下重构菌青素得到的光谱显示，在554 nm处吸光度最大，与我们的提取数据根据暗适应和光适应bR光谱计算的13-顺式成分的光谱吻合得很好。暗适应pm中13-顺式：全反式异构体的比例为2:1，在0 ~ 38℃之间基本保持不变，但在40℃以上开始明显下降，在70℃附近下降得更快，在90℃时达到0.75。40℃以上，异构体比例的范霍夫图显示出非线性的温度依赖关系，表明该体系比简单的13-顺式/全反式异构体热平衡更为复杂。我们将可见光吸收带随温度升高的增宽、吸光度降低和蓝移归因于至少一个、可能两个或三个新的发色团的出现，这些发色团主要或完全含有全反式异构体。（摘要删节250字）
On the basis of data obtained by spectroscopic analysis and chromatography of retinal extracts, a consensus has been adopted that dark-adapted purple membrane (pm) contains 13-cis- and all-trans-retinal in equal amounts, whereas the light-adapted membrane contains all-trans-retinal only. We have developed an improved extraction technique which extracts up to 70% of the retinal in pm within 4 min. In the extracts from dark-adapted pm at room temperature, we consistently find 66-67% 13-cis-retinal and 33-34% all-trans-retinal, and more than 98.5% all-trans isomer in light-adapted samples. The spectrum obtained by reconstitution of bacterioopsin with 13-cis-retinal at 2 degrees C (to minimize isomerization) shows an absorbance maximum at 554 nm and agrees well with the spectrum for the 13-cis component calculated from the dark-adapted and light-adapted bR spectra with our extraction data. The ratio of 13-cis:all-trans isomer in dark-adapted pm is 2:1 and nearly constant between 0 and 38 degrees C but begins to decrease distinctly above 40 degrees C, and more rapidly near 70 degrees C, reaching 0.75 at 90 degrees C. The van't Hoff plot of the isomer ratio shows a nonlinear temperature dependence above 40 degrees C, indicating a more complex system than a simple thermal 13-cis/all-trans isomer equilibrium. We attribute the broadening, absorbance decrease, and blut shift of the visible absorption band with increasing temperature to the appearance of at least one and possibly two or three new chromophores which contain, mainly or exclusively, the all-trans isomer.(ABSTRACT TRUNCATED AT 250 WORDS)