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Dimeric Structure of the Blue Light Sensor Protein Photozipper in the Active State
Biochemistry ( IF 2.9 ) Pub Date : 2017-12-21 00:00:00 , DOI: 10.1021/acs.biochem.7b01045 Kohei Ozeki 1 , Hiroyuki Tsukuno 1 , Hiroki Nagashima 1 , Osamu Hisatomi 2 , Hiroyuki Mino 1
Biochemistry ( IF 2.9 ) Pub Date : 2017-12-21 00:00:00 , DOI: 10.1021/acs.biochem.7b01045 Kohei Ozeki 1 , Hiroyuki Tsukuno 1 , Hiroki Nagashima 1 , Osamu Hisatomi 2 , Hiroyuki Mino 1
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The light oxygen voltage-sensing (LOV) domain plays a crucial role in blue light (BL) sensing in plants and microorganisms. LOV domains are usually associated with the effector domains and regulate the activities of effector domains in a BL-dependent manner. Photozipper (PZ) is monomeric in the dark state. BL induces reversible dimerization of PZ and subsequently increases its affinity for the target DNA sequence. In this study, we report the analyses of PZ by pulsed electron–electron double resonance (PELDOR). The neutral flavin radical was formed by BL illumination in the presence of dithiothreitol in the LOV-C254S (without the bZIP domain) and PZ-C254S mutants, where the cysteine residue responsible for adduct formation was replaced with serine. The magnetic dipole interactions of 3 MHz between the neutral radicals were detected in both LOV-C254S and PZ-C254S, indicating that these mutants are dimeric in the radical state. The PELDOR simulation showed that the distance between the radical pair is close to that estimated from the dimeric crystal structure in the “light state” [Heintz, U., and Schlichting, I. (2016) eLife5, e11860], suggesting that in the radical state, LOV domains in PZ-C254S form a dimer similar to that of LOV-C254S, which lacks the bZIP domain.
中文翻译：
处于活动状态的蓝光传感器蛋白质光拉链的二聚体结构
轻氧电压感测（LOV）域在植物和微生物的蓝光（BL）感测中起着至关重要的作用。LOV结构域通常与效应子结构域相关，并以BL依赖性方式调节效应子结构域的活性。在黑暗状态下，光拉链（PZ）是单体的。BL诱导PZ可逆二聚化，并随后增加其对靶DNA序列的亲和力。在这项研究中，我们报告了脉冲电子-电子双共振（PELDOR）对PZ的分析。中性黄素自由基是通过在LOV-C254S（无bZIP域）和PZ-C254S突变体中二硫苏糖醇存在下通过BL照射形成的，其中负责加合物形成的半胱氨酸残基被丝氨酸取代。在LOV-C254S和PZ-C254S中均检测到中性自由基之间3 MHz的磁偶极相互作用，表明这些突变体在自由基状态下为二聚体。PELDOR模拟表明，自由基对之间的距离接近于在“轻态”下由二聚体晶体结构估计的距离[Heintz，U.和Schlichting，I.（2016）[eLife 5 [ e11860]]，这表明在自由基状态下，PZ-C254S中的LOV域形成类似于LOV-C254S中缺少bZIP域的二聚体。
更新日期：2017-12-21
中文翻译：
处于活动状态的蓝光传感器蛋白质光拉链的二聚体结构
轻氧电压感测（LOV）域在植物和微生物的蓝光（BL）感测中起着至关重要的作用。LOV结构域通常与效应子结构域相关，并以BL依赖性方式调节效应子结构域的活性。在黑暗状态下，光拉链（PZ）是单体的。BL诱导PZ可逆二聚化，并随后增加其对靶DNA序列的亲和力。在这项研究中，我们报告了脉冲电子-电子双共振（PELDOR）对PZ的分析。中性黄素自由基是通过在LOV-C254S（无bZIP域）和PZ-C254S突变体中二硫苏糖醇存在下通过BL照射形成的，其中负责加合物形成的半胱氨酸残基被丝氨酸取代。在LOV-C254S和PZ-C254S中均检测到中性自由基之间3 MHz的磁偶极相互作用，表明这些突变体在自由基状态下为二聚体。PELDOR模拟表明，自由基对之间的距离接近于在“轻态”下由二聚体晶体结构估计的距离[Heintz，U.和Schlichting，I.（2016）[eLife 5 [ e11860]]，这表明在自由基状态下，PZ-C254S中的LOV域形成类似于LOV-C254S中缺少bZIP域的二聚体。
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