Evolution-inspired design of multicolored photoswitches from a single cyanobacteriochrome scaffold
Evolution-inspired design of multicolored photoswitches from a single cyanobacteriochrome scaffold
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DOI：
10.1073/pnas.2004273117
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发表时间：
2020-07-07
影响因子：
11.1
通讯作者：
Narikawa, Rei
中科院分区：
文献类型：
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作者：
Fushimi, Keiji;Hasegawa, Masumi;Narikawa, Rei
蓝细菌色素（Cyanobacteriochromes，CBCRs）是一种小的线性四吡咯（bilin）结合的光传感器，通常作为多结构域蓝细菌信号蛋白的模块组分而被发现。CBCR家族已被分为许多谱系，大致与其光谱多样性相关，但CBCR具有保守的DXCF基序被发现在多个谱系。DXCF CBCR通常具有两个保守的Cys残基：第一个Cys保持与胆色素发色团连接，第二个Cys存在于DXCF基序中。第二个半胱氨酸通常形成第二个硫醚键，提供了一种感知蓝光和紫光的机制。DXCF CBCR已被描述为具有蓝色/绿色、蓝色/橙子、蓝色/青色和绿色/青色光循环，并且已经很好地建立了这种光谱多样性的分子基础。我们在此描述了AM1_1499 g1的特征，这是一种非典型DXCF CBCR，缺乏第二个半胱氨酸残基，并显示出橙子/绿色光循环。基于CBCR光谱调谐的先前研究，我们已经成功地设计了七种AM 1_1499 g1变体，其表现出稳健的黄色/青色、绿色/青色、蓝色/青色、橙子/黄色、黄色/绿色、绿色/绿色和蓝色/绿色光循环。通过修饰单个CBCR产生的显着光谱多样性为在相同细胞背景下多路合成光生物学系统提供了良好的模板，从而绕过了具有不同蛋白质支架的多个探针所需的耗时的经验优化过程。
Cyanobacteriochromes (CBCRs) are small, bistable linear tetrapyrrole (bilin)-binding light sensors which are typically found as modular components in multidomain cyanobacterial signaling proteins. The CBCR family has been categorized into many lineages that roughly correlate with their spectral diversity, but CBCRs possessing a conserved DXCF motif are found in multiple lineages. DXCF CBCRs typically possess two conserved Cys residues: a first Cys that remains ligated to the bilin chromophore and a second Cys found in the DXCF motif. The second Cys often forms a second thioether linkage, providing a mechanism to sense blue and violet light. DXCF CBCRs have been described with blue/green, blue/orange, blue/teal, and green/teal photocycles, and the molecular basis for some of this spectral diversity has been well established. We here characterize AM1_1499g1, an atypical DXCF CBCR that lacks the second cysteine residue and exhibits an orange/green photocycle. Based on prior studies of CBCR spectral tuning, we have successfully engineered seven AM1_1499g1 variants that exhibit robust yellow/teal, green/teal, blue/teal, orange/yellow, yellow/green, green/green, and blue/green photocycles. The remarkable spectral diversity generated by modification of a single CBCR provides a good template for multiplexing synthetic photobiology systems within the same cellular context, thereby bypassing the timeconsuming empirical optimization process needed for multiple probes with different protein scaffolds.