Unbleachable rhodopsin with an 11-cis-locked eight-membered ring retinal: the visual transduction process.
Unbleachable rhodopsin with an 11-cis-locked eight-membered ring retinal: the visual transduction process.
复制标题
具有 11-顺式锁八元环视网膜的不可漂白视紫红质：视觉转导过程。
DOI：
10.1021/bi00168a004
复制
发表时间：
1994
期刊：
影响因子：
2.9
通讯作者：
Chen,AH
中科院分区：
文献类型：
--
作者：
Hu,S;Franklin,PJ;Wang,J;RuizSilva,BE;Derguini,F;Nakanishi,K;Chen,AH
摘要：视觉传导通过光视紫红质发生，光视紫红质是视紫红质的主要光产物，它松弛为视紫红质和一系列其他中间产物，直到它达到变视紫红质II阶段，在此阶段，导致视觉的酶级联被激活。尽管在与视觉传导相关的领域取得了进展，但触发过程本身，视紫红质化学中的一个关键问题，仍然没有解决。为了阐明发色团激发态与11-顺式到反式异构化的参与程度，并作为过去对11-m-锁七元环视紫红质（Rh 7）的研究的延伸，分别合成了11-顺式八元环和九元环视紫红质类似物，ret 8和ret 9。ret 8中的四亚甲基桥的庞大性导致了许多意想不到的障碍，试图重建一个包含ret 8的视紫红质（Rh 8）嵌入脂质双层膜。通过使用甲基化视紫红质解决了这些障碍，甲基化视紫红质产生含有11-m-ret 8作为其发色团的MeRh 8。MeRh 8表现出与天然视紫红质（Rh）非常相似的紫外-可见光谱和CD光谱;此外，光视紫红质形成的量子效率与Rh相当。Rh 8和其它环类似物的闪光光解研究[Mizukami，T.，Kandori，H.，Shichida，Y.，Chen，A.- H、Derguini，F.，考德威尔，CG，比格，C. F.、Nakanishi，K.，& Yoshizawa，T.等人（1993）Proc. Acad. Sci. USA 90，4072-4076]结合本发明的用MeRh 8和一系列二氢视紫红质的酶促研究，得出以下结论：（i）激发态的电荷易位确实发生;然而，（ii）有效转导发生需要涉及整个多烯部分的围绕11-烯的完全顺式-反式异构化。视色素视紫红质是G蛋白偶联的膜结合蛋白中研究最充分的一类。研究得最好的牛视紫红质（Rh），Xmax 500 nm，是由348个氨基酸折叠成7个跨膜α-螺旋组成的单一多肽;其发色团，全反式-rtxmz的11-c1 'j异构体2。1通过质子化席夫碱（SBH+）3与Lys-296结合（见表I）。通过发色团吸收光子导致11-cij全反式异构化，其引发Rh中的一系列构象变化。Rh激发的主要光产物现在被认为是Shichida等人发现的光视紫红质，Xmax 580 nm。（1984）;它在小于13 fs的时间内形成（Schoenlein等人，1991），并且被认为具有高度扭曲的11-反式双键。光-Rh热衰变为bathorhodopsin，Xmax 543 nm，1，其也具有扭曲的反式烯，但比光-Rh中的扭曲小。经过几个中间体的热弛豫后，Rh转化为关键中间体meta-Rh II，Xmax 380 nm（0 C），一种未质子化的席夫碱。在这个阶段，Meta II与G蛋白超家族成员transducin结合，从而激活导致视觉转导的酶级联反应。单光子吸收-*·转导素激活-* 磷酸二酯酶（PDE）激活的步骤导致105个cGMP分子水解为GMP，
Revised Manuscript Received October 27, 1993® abstract: Visual transduction occurs through photorhodopsin, the primary photoproduct of rhodopsin, which relaxes to bathorhodopsin and a series of otherintermediates until it reaches the metarhodopsin II stage, upon which the enzymatic cascade leading to vision is activated. Despite advances in areas related to visual tramduction, the triggering process itself, a key problem in the chemistry of rhodopsin, has remained unsolved. In order to clarify the extent of involvement of the chromophoric excited state versus the 11-cis to trans isomerization, and as an extension of past studies with 11-m-locked seven-membered ring rhodopsin (Rh7), 1\-cis eight-and nine-membered ring retinal analogs, ret8 and ret9, respectively, have been synthesized. The bulkiness of the tetramethylene bridge in ret8 led to numerous unexpected obstacles in attempts to reconstitute a ret8-containing rhodopsin (Rh8) embedded in lipid bilayer membranes. These obstacles were solved by using methylated rhodopsin which gave MeRh8 containing 11-m-ret8 as its chromophore. MeRh8 exhibited UV-vis and CD spectra very similar to those of native rhodopsin (Rh); furthermore, the quantum efficiency of photorhodopsin formation was comparable to that of Rh. Flash photolytic studies of Rh8 and other ring analogs [Mizukami, T., Kandori, H., Shichida, Y., Chen, A.-H., Derguini, F., Caldwell, CG, Bigge, C. F., Nakanishi, K., & Yoshizawa, T.(1993) Proc. Natl. Acad. Sci. USA 90, 4072-4076] coupled with the present enzymatic studies with MeRh8 and a series of dihydro-rhodopsins have led to the conclusion that (i) charge translocation in the excited state does occur; however,(ii) full cis-trans isomerization around 11-ene involving the entire polyene moiety is required for efficient transduction to occur. Repeated attempts to incorporate ret9 into opsin have as yet not been successful.The visual pigment rhodopsins are one of the most fully investigated group of G-protein-coupled membrane-bound proteins. The best studied bovine rhodopsin (Rh), Xmax 500 nm, is a single polypeptide consisting of 348 amino acids folded into seven membrane-spanning a-helices; its chromophore, the 1 1-ci'j isomer 2 of all-trans-rtxmz.\1, is bound to Lys-296 via a protonated Schiff base (SBH+) 3 (see Table I). Absorption of a photon by the chromophore leads to 11-cij to all-trans isomerization which initiates a series of conformational changes in Rh. The primary photoproduct of Rh excitation is now considered to be photorhodopsin, Xmax 580 nm, discovered by Shichida et al.(1984); it is formed in less than 13 fs (Schoenlein et al., 1991) and is considered to have a highly distorted 11-trans double bond. Photo-Rh thermally decays to bathorhodopsin, Xmax 543 nm, 1 which also has a distorted trans-ene but less distorted than in photo-Rh. After thermal relaxation through several further intermediates, Rh is converted into the key intermediate meta-Rh II, Xmax 380 nm (0 C), an unprotonated Schiff base. At this stage, meta II binds to transducin, a member of the G-protein superfamily, thus activating the enzymatic cascade leading to visual transduction. The steps of single photon absorption—*• transducin activation-* phosphodiesterase (PDE) activation results in hydrolysis of 105 molecules of cGMP to GMP, upon