Department of Organic Chemistry, Weizmann Institute of Science , Rehovot 76100, Israel.
J Phys Chem B. 2015 Jan 15;119(2):456-64. doi: 10.1021/jp510534s. Epub 2014 Dec 30.
Xanthorhodopsin (xR) is a retinal protein that contains, in addition to the retinal moiety, a salinixanthin chromophore absorbing at 456, 486, and 520 nm [Balashov, S. P.; Science 2005, 309, 2061]. The CD spectrum of xR is very unique with a "conservative" character, containing negative and positive lobes and resembling the first derivative of the absorption spectrum [Balashov, S. P.; Biochemistry 2006, 45, 10998]. It was suggested that the CD spectrum is likely to be composed of several components and that the salinixanthin interacts closely with the retinal chromophore [Balashov, S. P.; Biochemistry 2006, 45, 10998; Imasheva, E. S.; Photochem. Photobiol. 2008, 84, 977; Lanyi, J. K.; Acta Bioenerg. 2008, 1777, 684; Smolensky, E.; Biochemistry 2009, 48, 8179; Smolensky Koganov, E.; Biochemistry 2013, 52, 1290]. In this work, we aim to further explore the nature and origin of the unique CD spectrum of xR. We follow the absorption and CD spectra at different pHs of wild-type (wt) xR and of artificial xR pigments, characterized by a shifted absorption maximum of the retinal chromophore, as well as their corresponding reduced retinal protonated Schiff base pigments. Our results revealed a protein residue (other than the protonated Schiff base counterion), for which protonation affects the CD spectrum by decreasing the negative lobe at ∼530 nm and the positive lobes at 478 and 455 nm, which might be due to elimination of excitonic coupling between the salinixanthin chromophores, although other possibilities cannot be completely excluded. This spectrum change occurs by the pH decreasing, even in artificial pigment where the absorption of the retinal pigment is significantly shifted from 570 to about 450 nm. The possible excitonic coupling between the salinixanthin chromophores and its contribution to the CD spectrum of xR were supported by a good fitting of the CD spectrum to conservative (excitonic) bands [Zsila, F.; Tetrahedron: Asymmetry 2001, 12, 3125; Zsila, F.; Tetrahedron: Asymmetry 2002, 13, 273]. We propose that the CD spectrum of xR consists of contributions from an excitonic coupling interaction between the salinixanthins chromophores located in different subunits of the 3D structure of xR, the chiral conformation of the salinixanthin within its binding site, and the contribution of the retinal chromophore to the negative lobe at around 550 nm.
黄蛋白（xR）是一种视网膜蛋白，除了含有视黄醛部分外，还含有吸收波长为 456、486 和 520nm 的盐藻黄素发色团[Balashov, S. P.; Science 2005, 309, 2061]。xR 的 CD 谱非常独特，具有“保守”特征，包含负峰和正峰，类似于吸收光谱的一阶导数[Balashov, S. P.; Biochemistry 2006, 45, 10998]。有人提出，CD 谱可能由几个组成部分组成，盐藻黄素与视黄醛发色团密切相互作用[Balashov, S. P.; Biochemistry 2006, 45, 10998; Imasheva, E. S.; Photochem. Photobiol. 2008, 84, 977; Lanyi, J. K.; Acta Bioenerg. 2008, 1777, 684; Smolensky, E.; Biochemistry 2009, 48, 8179; Smolensky Koganov, E.; Biochemistry 2013, 52, 1290]。在这项工作中，我们旨在进一步探索 xR 独特 CD 谱的性质和起源。我们研究了野生型（wt）xR 和人工 xR 色素在不同 pH 值下的吸收和 CD 光谱，这些色素的特征是视黄醛发色团的吸收最大值发生了位移，以及它们相应的还原视黄醛质子化席夫碱色素。我们的结果揭示了一个蛋白质残基（除了质子化席夫碱抗衡离子），其质子化通过降低约 530nm 的负峰和 478nm 和 455nm 的正峰来影响 CD 谱，这可能是由于盐藻黄素发色团之间的激子耦合消除所致，尽管不能完全排除其他可能性。这种光谱变化是通过 pH 值降低引起的，即使在人工色素中，视黄醛色素的吸收也明显从 570nm 左右移动到约 450nm。盐藻黄素发色团之间的可能激子耦合及其对视黄蛋白 xR 的 CD 谱的贡献得到了保守（激子）带的良好拟合的支持[Zsila, F.; Tetrahedron: Asymmetry 2001, 12, 3125; Zsila, F.; Tetrahedron: Asymmetry 2002, 13, 273]。我们提出，xR 的 CD 谱由位于 xR 三维结构不同亚基中的盐藻黄素发色团之间的激子耦合相互作用、盐藻黄素在其结合位点中的手性构象以及视黄醛发色团对约 550nm 处负峰的贡献组成。
