Spectroscopic evidence for the formation of an N-intermediate during the photocycle of sensory rhodopsin (phoborhodopsin) from Natromonaspharaonis
Spectroscopic evidence for the formation of an N-intermediate during the photocycle of sensory rhodopsin (phoborhodopsin) from Natromonaspharaonis
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Natromonaspharaonis 感觉视紫红质 (phoborhodopsin) 光循环过程中形成 N-中间体的光谱证据
DOI：
10.1021/bi1019572
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发表时间：
2011
期刊：
影响因子：
2.9
通讯作者：
Unno Masashi
中科院分区：
文献类型：
--
作者：
Tateishi Yusuke;Abe Takayuki;Tamogami Jun;Nakao Yutaka;Kikukawa Takashi;KamoNaoki ;Unno Masashi
感觉视紫红质 II 是一种七跨膜螺旋状视网膜蛋白，在嗜盐古细菌的负趋光性中充当光感受器蛋白。来自 Natronomonas pharaonis (NpSRII) 的感觉视紫红质 II 在各种条件下都很稳定，并且可以在大肠杆菌细胞膜中功能性表达。来自微生物的视紫红质，称为微生物视紫红质，表现出光循环，这些分子的光照射会产生高能中间体，该中间体在经过几个中间体后热弛豫为原始色素。对于细菌视紫红质 (BR)（一种光驱动质子泵），光循环被建立为 BR → K → L → M → N → O → BR。 NpSRII 的光循环与 BR 相似，除了 N，即 M 热衰变成 O，而 N 在光循环中尚未得到很好的表征。因此，我们在这里检查了 NpSRII 中光循环的后半部分，在目前的瞬态吸收研究中，我们发现形成了一种新的光中间体，其最大吸收波长为 ∼500 nm。该中间体在叠氮化物存在下变得明显，从而加速了 M 的衰变。进一步应用瞬态共振拉曼光谱来证明该中间体含有 13-顺视黄醛质子化席夫碱。然而，对瞬态吸收数据的详细分析表明，M 衰变并不直接产生 N，而是产生与 N 平衡的 O。这些观察结果使我们能够提出涉及 N 的光循环的结构模型。
Sensory rhodopsin II is a seven transmembrane helical retinal protein and functions as a photoreceptor protein in negative phototaxis of halophilic archaea. Sensory rhodopsin II fromNatronomonas pharaonis(NpSRII) is stable under various conditions and can be expressed functionally inEscherichia colicell membranes. Rhodopsins from microorganisms, known as microbial rhodopsins, exhibit a photocycle, and light irradiation of these molecules leads to a high-energy intermediate, which relaxes thermally to the original pigment after passing through several intermediates. For bacteriorhodopsin (BR), a light-driven proton pump, the photocycle is established as BR → K → L → M → N → O → BR. The photocycle of NpSRII is similar to that of BR except for N, i.e., M thermally decays into the O, and N has not been well characterized in the photocycle. Thus we here examined the second half of the photocycle in NpSRII, and in the present transient absorption study we found the formation of a new photointermediate whose absorption maximum is ∼500 nm. This intermediate becomes pronounced in the presence of azide, which accelerates the decay of M. Transient resonance Raman spectroscopy was further applied to demonstrate that this intermediate contains a 13-cisretinal protonated Schiff base. However, detailed analysis of the transient absorption data indicated that M-decay does not directly produce N but rather produces O that is in equilibrium with N. These observations allowed us to propose a structural model for a photocycle that involves N.