Function of extracellular loop 2 in rhodopsin: glutamic acid 181 modulates stability and absorption wavelength of metarhodopsin II.
Function of extracellular loop 2 in rhodopsin: glutamic acid 181 modulates stability and absorption wavelength of metarhodopsin II.
复制标题
视紫质细胞外环 2 的功能：谷氨酸 181 调节变视紫红质 II 的稳定性和吸收波长。
DOI：
10.1021/bi0160011
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发表时间：
2002
期刊：
影响因子：
2.9
通讯作者：
Sakmar,ThomasP
中科院分区：
文献类型：
--
作者：
Yan,ElsaCY;Kazmi,ManijaA;De,Soma;Chang,BelindaSW;Seibert,Christoph;Marin,EthanP;Mathies,RichardA;Sakmar,ThomasP
第二个视紫红质的胞外环折叠回受体的膜嵌入区域，形成11-顺式视黄醛发色团的结合袋的一部分。这个环的羧酸侧链Glu181指向视网膜多烯链的中心。我们通过描述一组位点定向突变体来研究Glu181在牛视紫红质中的作用。19个单位点突变体中的16个表达并结合11-顺式视网膜形成色素。突变体色素E181Q在没有NaCl的情况下，λ最大值出现了明显的红移，达到508 nm。其他替代对突变体色素在黑暗中的光谱特征没有显著影响。因此，Glu181对紫红质基态的光谱调节没有显著贡献。对这些数据最可能的解释是，Glu181在视紫质的黑暗状态下被质子化且不带电。在黑暗中，Glu181突变体对羟胺的反应性显著增强。突变体在光照下形成后视紫红质ii样光产物，但许多光产物的λ最大值发生偏移。此外，突变体色素的后视紫红质ii样光产物的衰变速率也发生了显著变化。突变体对羟胺的反应性增强支持了第二个细胞外环阻止溶剂进入发色团结合袋的观点。此外，Glu181强烈影响活性后视紫红质II光产物中视黄醛席夫碱的环境。
The second extracellular loop of rhodopsin folds back into the membrane-embedded domain of the receptor to form part of the binding pocket for the 11-cis-retinylidene chromophore. A carboxylic acid side chain from this loop, Glu181, points toward the center of the retinal polyene chain. We studied the role of Glu181 in bovine rhodopsin by characterizing a set of site-directed mutants. Sixteen of the 19 single-site mutants expressed and bound 11-cis-retinal to form pigments. The λmaxvalue of mutant pigment E181Q showed a significant spectral red shift to 508 nm only in the absence of NaCl. Other substitutions did not significantly affect the spectral features of the mutant pigments in the dark. Thus, Glu181 does not contribute significantly to spectral tuning of the ground state of rhodopsin. The most likely interpretation of these data is that Glu181 is protonated and uncharged in the dark state of rhodopsin. The Glu181 mutants displayed significantly increased reactivity toward hydroxylamine in the dark. The mutants formed metarhodopsin II-like photoproducts upon illumination but many of the photoproducts displayed shifted λmaxvalues. In addition, the metarhodopsin II-like photoproducts of the mutant pigments had significant alterations in their decay rates. The increased reactivity of the mutants to hydroxylamine supports the notion that the second extracellular loop prevents solvent access to the chromophore-binding pocket. In addition, Glu181 strongly affects the environment of the retinylidene Schiff base in the active metarhodopsin II photoproduct.