Furutani Yuji, Shichida Yoshinori, Kandori Hideki
Department of Applied Chemistry, Nagoya Institute of Technology, Showa-ku, Nagoya 466-8555, Japan.
Biochemistry. 2003 Aug 19;42(32):9619-25. doi: 10.1021/bi034592k.
Internal water molecules of rhodopsins play an important role in stabilizing the crucial ion pair comprised by the protonated retinal Schiff base and its counterion. Previous low-temperature FTIR spectroscopy of archaeal rhodopsins observed water O-D stretching vibrations at 2400-2100 cm(-1) in D(2)O, corresponding to strong hydrogen bonds. Since a water molecule bridges the protonated Schiff base and an aspartate in archaeal rhodopsins, the observed water molecules presumably hydrate the negative charges in the Schiff base region. In contrast, the FTIR spectroscopy data of bovine rhodopsin presented here revealed that there are no spectral changes of water molecules under strongly hydrogen-bonding conditions (in the range <2400 cm(-1) for O-D stretch) during the photoactivation processes. The only observed water bands were located in the >2500 cm(-1) region that corresponds to weak hydrogen bonding. These results imply that the ion pair state in vertebrate visual rhodopsins is stabilized in a manner different from that in archaeal rhodopsins. In addition, the internal water molecules that hydrate the negative charges do not play important role in the photoactivation processes of rhodopsin that involve proton transfer from the Schiff base to Glu113 upon formation of Meta II. Structural changes of the H-D exchangeable peptide amide of a beta-sheet are observed upon formation of metarhodopsin II, suggesting that motion of a beta-sheet is coupled to the proton transfer reaction from the Schiff base to its counterion.
视紫红质的内部水分子在稳定由质子化视黄醛席夫碱及其抗衡离子组成的关键离子对方面发挥着重要作用。先前对古细菌视紫红质进行的低温傅里叶变换红外光谱研究在重水（D₂O）中观察到水分子的O-D伸缩振动位于2400 - 2100 cm⁻¹处，这对应于强氢键。由于在古细菌视紫红质中一个水分子连接着质子化席夫碱和一个天冬氨酸，所以观察到的这些水分子可能使席夫碱区域的负电荷水合。相比之下，此处展示的牛视紫红质的傅里叶变换红外光谱数据表明，在光激活过程中，在强氢键条件下（O-D伸缩振动范围<2400 cm⁻¹）水分子没有光谱变化。唯一观察到的水吸收带位于>2500 cm⁻¹区域，这对应于弱氢键。这些结果意味着脊椎动物视觉视紫红质中的离子对状态是以一种不同于古细菌视紫红质的方式稳定的。此外，使负电荷水合的内部水分子在视紫红质的光激活过程中并不起重要作用，视紫红质的光激活过程涉及在形成变视紫红质II时质子从席夫碱转移到谷氨酸113。在形成变视紫红质II时观察到β折叠的H-D可交换肽酰胺的结构变化，这表明β折叠的运动与质子从席夫碱转移到其抗衡离子的反应相耦合。
