Pande C, Lanyi J K, Callender R H
Physics Department, City College of New York, New York 10031.
Biophys J. 1989 Mar;55(3):425-31. doi: 10.1016/S0006-3495(89)82836-X.
Resonance Raman experiments were conducted to probe and understand the effect of various anions on halorhodopsin. These included monoatomic anions Cl- and Br-, which bind to the so-called halorhodopsin binding sites I and II, and polyatomic anions NO3- and ClO4-, which bind to site I only. The two types of ions clearly show different effects on the vibrational spectrum of the chromophore. The differences are not localized to the Schiff base region of the molecule, but extend to the chromophore structure-sensitive fingerprint region as well. We find that the protonated Schiff base frequency is at 1,633 cm-1 for Cl- and Br- ions, as reported previously for Cl-. However, we find that two Schiff base frequencies characterize halorhodopsin upon binding of the polyatomic anions. One frequency lies at the same location as that found for the monoatomic anions and the other is at 1,645 cm-1. Halorhodopsin with bound NO3- and ClO4- thus may consist of two heterogeneous structures in equilibrium. This heterogeneity does not seem to correlate with a retinal isomeric heterogeneity, which we can also demonstrate in these samples. The results suggest that anions binding to site I do not bind to the Schiff base directly, but can influence chromophore and/or protein conformational states.
进行了共振拉曼实验，以探究和理解各种阴离子对嗜盐视紫红质的影响。这些阴离子包括单原子阴离子Cl-和Br-，它们与所谓的嗜盐视紫红质结合位点I和II结合，以及多原子阴离子NO3-和ClO4-，它们仅与位点I结合。这两种离子对发色团的振动光谱表现出明显不同的影响。差异不仅局限于分子的席夫碱区域，还延伸到对发色团结构敏感的指纹区域。我们发现，对于Cl-和Br-离子，质子化席夫碱频率为1633 cm-1，这与之前报道的Cl-情况相同。然而，我们发现，在多原子阴离子结合后，嗜盐视紫红质有两个席夫碱频率特征。一个频率与单原子阴离子的频率位置相同，另一个在1645 cm-1。因此，结合了NO3-和ClO4-的嗜盐视紫红质可能由处于平衡状态的两种异质结构组成。这种异质性似乎与视网膜异构体异质性无关，我们也能在这些样品中证明这一点。结果表明，与位点I结合的阴离子并不直接与席夫碱结合，但可以影响发色团和/或蛋白质的构象状态。
