Imai H, Hirano T, Kandori H, Terakita A, Shichida Y
Department of Biophysics, Graduate School of Science, Kyoto University, Kyoto 606-8502, Japan.
Biochemistry. 2001 Mar 6;40(9):2879-86. doi: 10.1021/bi002227c.
To investigate the local structure that causes the differences in molecular properties between rod and cone visual pigments, we have measured the difference infrared spectra between chicken green and its photoproduct at 77 K and compared them with those from bovine and chicken rhodopsins. In contrast to the similarity of the vibrational bands of the chromophore, those of the protein part were notably different between chicken green and the rhodopsins. Like the rhodopsins, chicken green has an aspartic acid at position 83 (D83) but exhibited no signals due to the protonated carboxyl of D83 in the C=O stretching region, suggesting that the molecular contact between D83 and G120 through water molecule evidenced in bovine rhodopsin is absent in chicken green. A pair of positive and negative bands due to the peptide backbone (amide I) was prominent in chicken green, while the rhodopsins exhibited only small bands in this region. Furthermore, chicken green exhibited characteristic paired bands around 1480 cm(-1), which were identified as the imide bands of P189 using site-directed mutagenesis. P189, situated in the putative second extracellular loop, is conserved in all the known cone visual pigments but not in rhodopsins. Thus, some region of the second extracellular loop including P189 is situated near the chromophore and changes its environment upon formation of the batho-intermediate. The results noted above indicate that differences in the protein parts between chicken green and the rhodopsins alter the changes seen in the protein upon photoisomerization of the chromophore. Some of these changes appear to be the pathway from the chromophore to cytoplasmic surface of the pigment and thus could affect the activation process of transducin.
为了研究导致视杆和视锥视觉色素分子特性差异的局部结构，我们测量了鸡绿色视色素及其光产物在77K下的红外光谱差异，并将其与牛视紫红质和鸡视紫红质的光谱进行了比较。与发色团振动带的相似性不同，鸡绿色视色素和视紫红质的蛋白质部分的振动带明显不同。与视紫红质一样，鸡绿色视色素在第83位有一个天冬氨酸（D83），但在C=O伸缩区域没有因D83质子化羧基产生的信号，这表明在鸡绿色视色素中不存在牛视紫红质中所证实的D83与G120之间通过水分子的分子接触。鸡绿色视色素中由于肽主链（酰胺I）产生的一对正负带很突出，而视紫红质在该区域仅显示出小带。此外，鸡绿色视色素在1480 cm(-1)左右呈现出特征性的成对带，通过定点诱变将其鉴定为P189的酰亚胺带。位于假定的第二个细胞外环中的P189在所有已知的视锥视觉色素中保守，但在视紫红质中不保守。因此，包括P189在内的第二个细胞外环的某些区域位于发色团附近，并在形成嗜热中间体时改变其环境。上述结果表明，鸡绿色视色素和视紫红质在蛋白质部分的差异改变了发色团光异构化时蛋白质中所观察到的变化。其中一些变化似乎是从发色团到色素细胞质表面的途径，因此可能影响转导素的激活过程。
