rsCherryRev1.4 has been reported as one of the reversibly photoswitchable variants of mCherry, and is an improved version with a faster off-switching speed and lower switching fatigue at high light intensities than its precursor rsCherryRev. However, rsCherryRev1.4 still has some limitations such as a tendency to dimerize as well as complex photophysical properties. Here, the crystal structure of rsCherryRev1.4 was determined at a resolution of 2 Å and it was discovered that it forms a dimer that shows disulfide bonding between the protomers. Mutagenesis, gel electrophoresis and size-exclusion chromatography strongly implicate Cys24 in this process. Replacing Cys24 in rsCherryRev1.4 resulted in a much lower tendency towards dimerization, while introducing Cys24 into mCherry correspondingly increased its dimerization. In principle, this finding opens the possibility of developing redox sensors based on controlled dimerization via disulfide cross-linking in fluorescent proteins, even though the actual application of engineering such sensors still requires additional research.

中文翻译：

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荧光蛋白 rsCherryRev1.4 中不寻常的二硫键二聚化

rsCherryRev1.4 已被报道为 mCherry 的可逆光开关变体之一，并且是一个改进版本，与它的前身 rsCherryRev 相比，在高光强度下具有更快的关闭开关速度和更低的开关疲劳。然而，rsCherryRev1.4 仍然存在一些局限性，例如二聚化倾向以及复杂的光物理特性。在这里，rsCherryRev1.4 的晶体结构以 2 Å 的分辨率确定，并发现它形成了一个二聚体，显示了原聚体之间的二硫键。诱变、凝胶电泳和尺寸排阻色谱强烈暗示 Cys24 参与了该过程。在 rsCherryRev1.4 中替换 Cys24 导致二聚化的趋势低得多，而将 Cys24 引入 mCherry 相应地增加了它的二聚化。原则，