Rousso I, Khachatryan E, Gat Y, Brodsky I, Ottolenghi M, Sheves M, Lewis A
Department of Organic Chemistry, The Weizmann Institute of Science, Rehovot 76100, Israel.
Proc Natl Acad Sci U S A. 1997 Jul 22;94(15):7937-41. doi: 10.1073/pnas.94.15.7937.
In this paper a new atomic force sensing technique is presented for dynamically probing conformational changes in proteins. The method is applied to the light-induced changes in the membrane-bound proton pump bacteriorhodopsin (bR). The microsecond time-resolution of the method, as presently implemented, covers many of the intermediates of the bR photocycle which is well characterized by spectroscopical methods. In addition to the native pigment, we have studied bR proteins substituted with chemically modified retinal chromophores. These synthetic chromophores were designed to restrict their ability to isomerize, while maintaining the basic characteristic of a large light-induced charge redistribution in the vertically excited Franck-Condon state. An analysis of the atomic force sensing signals lead us to conclude that protein conformational changes in bR can be initiated as a result of a light-triggered redistribution of electronic charge in the retinal chromophore, even when isomerization cannot take place. Although the coupling mechanism of such changes to the light-induced proton pump is still not established, our data question the current working hypothesis which attributes all primary events in retinal proteins to an initial trans<==>cis isomerization.
本文提出了一种用于动态探测蛋白质构象变化的新型原子力传感技术。该方法应用于膜结合质子泵细菌视紫红质（bR）的光诱导变化。目前所实现的该方法的微秒级时间分辨率涵盖了bR光循环的许多中间体，这些中间体已通过光谱方法得到了很好的表征。除了天然色素外，我们还研究了用化学修饰的视黄醛发色团取代的bR蛋白。这些合成发色团旨在限制其异构化能力，同时保持在垂直激发的弗兰克 - 康登态中光诱导的大电荷重新分布的基本特征。对原子力传感信号的分析使我们得出结论，即使异构化无法发生，bR中的蛋白质构象变化也可由视黄醛发色团中光触发的电荷重新分布引发。尽管这种变化与光诱导质子泵的耦合机制尚未确立，但我们的数据对当前将视网膜蛋白中的所有初级事件归因于初始反式⇌顺式异构化的工作假设提出了质疑。
