Structural insight into enhanced calcium indicator GCaMP3 and GCaMPJ to promote further improvement
Structural insight into enhanced calcium indicator GCaMP3 and GCaMPJ to promote further improvement
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DOI：
10.1007/s13238-013-2103-4
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发表时间：
2013-04-01
期刊：
影响因子：
21.1
通讯作者：
Ji, Guangju
中科院分区：
文献类型：
--
作者：
Chen, Yingxiao;Song, Xianqiang;Ji, Guangju
遗传编码Ca2+指标（GECI）是重要的钙+在体内的测量。GCaMP2是一种广泛使用的GECI，最近得到了迭代改进。在改进的变体中，GCaMP3表现出明显更好的荧光强度。在这项研究中，我们开发了一种新的GECI，称为GCaMPJ，并确定了GCaMP3和GCaMPJ的晶体结构。与GCaMP3相比，GCaMPJ的荧光增强1.5倍，钙亲和力增强1.3倍。在Ca2+结合后，GCaMP3表现为单体和二聚体形式。这两种形态的结构叠加揭示了Arg-376在改善单体性能方面的作用。然而，GCaMPJ很少在类似GCaMP3的条件下形成二聚体。对Tyr-380的结构和诱变研究证实了它在阻断cpEGFP β管孔中的重要性。我们的研究提出了一种有效的工具来绘制完整器官中的Ca2+信号，以促进GCaMP传感器的进一步改进。
Genetically encoded Ca2+ indicators (GECI) are important for the measurement of Ca2+ in vivo. GCaMP2, a widely-used GECI, has recently been iteratively improved. Among the improved variants, GCaMP3 exhibits significantly better fluorescent intensity. In this study, we developed a new GECI called GCaMPJ and determined the crystal structures of GCaMP3 and GCaMPJ. GCaMPJ has a 1.5-fold increase in fluorescence and 1.3-fold increase in calcium affinity over GCaMP3. Upon Ca2+ binding, GCaMP3 exhibits both monomeric and dimeric forms. The structural superposition of these two forms reveals the role of Arg-376 in improving monomer performance. However, GCaMPJ seldom forms dimers under conditions similar to GCaMP3. St ructural and mutagenesis studies on Tyr-380 confirmed its importance in blocking the cpEGFP beta-barrel holes. Our study proposes an efficient tool for mapping Ca2+ signals in intact organs to facilitate the further improvement of GCaMP sensors.