Pande C, Callender R, Henderson R, Pande A
Physics Department, City College of City University of New York, New York 10031.
Biochemistry. 1989 Jul 11;28(14):5971-8. doi: 10.1021/bi00440a038.
In an effort to understand the nature of chromophore-protein interactions in bacteriorhodopsin (bR), we have reinvestigated dimethyl sulfoxide (DMSO)-induced changes in bR [Oesterhelt et al. (1973) Eur. J. Biochem. 40, 453-463]. We observe that dark-adapted bR (bR560) in aqueous DMSO undergoes reversible transformation to a species absorbing maximally at 480 nm (bR480). Beginning at 40% DMSO, this change results in complete conversion to bR480 at 60% DMSO. The kinetics of the reaction reveal that this transformation takes place predominantly through the all-trans isomeric form of the pigment. Thermal isomerization of the 13-cis chromophore to the all-trans form is, therefore, the rate-limiting step in the formation of bR480 from the dark-adapted bR. As in native bR, the chromophore in bR480 is linked to the protein via a protonated Schiff base, and its isomeric composition is predominantly all-trans. The formation of bR480 is associated with minor changes in the protein secondary structure, and the membrane retains crystallinity. These changes in the protein structure result in a diminished chromophore-protein interaction near the Schiff base region in bR480. Thus, we attribute the observed spectroscopic changes in bR in DMSO to structural alteration of the protein. The 13-cis chromophoric pigment appears to be resistant to this solvent-induced change. The changes in the protein structure need not be very large; displacement of the protein counterion(s) to the Schiff base, resulting from minor changes in the protein structure, can produce the observed spectral shift.
为了了解细菌视紫红质（bR）中发色团与蛋白质相互作用的本质，我们重新研究了二甲基亚砜（DMSO）诱导的bR变化[奥斯特黑尔特等人（1973年），《欧洲生物化学杂志》40卷，453 - 463页]。我们观察到，在含水DMSO中暗适应的bR（bR560）可逆地转变为在480 nm处吸收最大的物种（bR480）。从40% DMSO开始，这种变化在60% DMSO时导致完全转化为bR480。反应动力学表明，这种转变主要通过色素的全反式异构体形式发生。因此，13 - 顺式发色团向全反式形式的热异构化是从暗适应的bR形成bR480的限速步骤。与天然bR一样，bR480中的发色团通过质子化席夫碱与蛋白质相连，其异构体组成主要是全反式。bR480的形成与蛋白质二级结构的微小变化相关，并且膜保持结晶性。蛋白质结构的这些变化导致bR480中席夫碱区域附近的发色团 - 蛋白质相互作用减弱。因此，我们将在DMSO中观察到的bR光谱变化归因于蛋白质的结构改变。13 - 顺式发色团色素似乎对这种溶剂诱导的变化具有抗性。蛋白质结构的变化不一定非常大；蛋白质结构的微小变化导致蛋白质抗衡离子向席夫碱的位移，可以产生观察到的光谱位移。
