Structural basis of the protochromic green/red photocycle of the chromatic acclimation sensor RcaE
Structural basis of the protochromic green/red photocycle of the chromatic acclimation sensor RcaE
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色适应传感器 RcaE 的原色绿/红光循环的结构基础
DOI：
10.1073/pnas.2024583118
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发表时间：
2021
期刊：
影响因子：
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通讯作者：
Masaki Mishima
中科院分区：
文献类型：
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作者：
Takayuki Nagae;Masashi Unno;Taiki Koizumi;Yohei Miyanoiri;Tomotsumi Fujisawa;Kento Masui;Takanari Kamo;Kei Wada;Toshihiko Eki;Yutaka Ito;Yuu Hirose;Masaki Mishima
蓝细菌色素（CBCRs）是光敏色素超家族的胆色素结合型光传感器，具有显著的光谱多样性。绿色/红色CBCR亚家族在蓝细菌光合天线的颜色适应调节中起重要作用，并在合成生物学中用于基因表达的光遗传学控制。这表明该亚家族的吸收变化是由胆色素C_（15）-Z/C_（15）-E光异构化和随后的胆色素质子化状态的变化引起的。然而，结构信息和直接证据的胆色素质子化状态是缺乏的。在这里，我们报告的高分辨率（1.63纳米）晶体结构的胆色素结合域的色驯化传感器RcaE在红色吸收的光产物状态。胆色素被埋在由疏水残基组成的“桶”内，其中胆色素构型/构象为C5-Z、syn/C10-Z、syn/C15-E、syn，其中A-至C-环共面并且D-环倾斜。A-至C-环的三个吡咯氮在α-面被影响胆色素pKa的Leu 249的疏水盖覆盖，而它们在β-面与Glu 217的羧基直接氢键结合。Glu 217还连接到在桶中形成孔的沃茨的簇，其在分子动力学模拟中与溶剂沃茨交换。我们建议，“漏桶”结构的功能作为一个质子出口/流入途径后，光转换。NMR分析表明，四个吡咯氮原子确实在红色吸收状态下完全质子化，但其中一个，最有可能是B环氮，在绿色吸收状态下去质子化。这些发现加深了我们对CBCR中存在的不同光谱调谐机制的理解。
Cyanobacteriochromes (CBCRs) are bilin-binding photosensors of the phytochrome superfamily that show remarkable spectral diversity. The green/red CBCR subfamily is important for regulating chromatic acclimation of photosynthetic antenna in cyanobacteria and is applied for optogenetic control of gene expression in synthetic biology. It is suggested that the absorption change of this subfamily is caused by the bilin C15-Z/C15-Ephotoisomerization and a subsequent change in the bilin protonation state. However, structural information and direct evidence of the bilin protonation state are lacking. Here, we report a high-resolution (1.63Å) crystal structure of the bilin-binding domain of the chromatic acclimation sensor RcaE in the red-absorbing photoproduct state. The bilin is buried within a “bucket” consisting of hydrophobic residues, in which the bilin configuration/conformation is C5-Z,syn/C10-Z,syn/C15-E,synwith the A- through C-rings coplanar and the D-ring tilted. Three pyrrole nitrogens of the A- through C-rings are covered in the α-face with a hydrophobic lid of Leu249 influencing the bilin pKa, whereas they are directly hydrogen bonded in the β-face with the carboxyl group of Glu217. Glu217 is further connected to a cluster of waters forming a hole in the bucket, which are in exchange with solvent waters in molecular dynamics simulation. We propose that the “leaky bucket” structure functions as a proton exit/influx pathway upon photoconversion. NMR analysis demonstrated that the four pyrrole nitrogen atoms are indeed fully protonated in the red-absorbing state, but one of them, most likely the B-ring nitrogen, is deprotonated in the green-absorbing state. These findings deepen our understanding of the diverse spectral tuning mechanisms present in CBCRs.