Ca²⁺ regulates several cellular functions, including signaling events, energy production, and cell survival. These cellular processes are mediated by Ca²⁺-binding proteins, such as EF-hand superfamily proteins. Among the EF-hand superfamily proteins, allograft inflammatory factor-1 (AIF-1) and swiprosin-1/EF-hand domain-containing protein 2 (EFhd2) are cytosolic actin-binding proteins. AIF-1 modulates the cytoskeleton and increases the migration of immune cells. EFhd2 is also a cytoskeletal protein implicated in immune cell activation and brain cell functions. EFhd1, a mitochondrial fraternal twin of EFhd2, mediates neuronal and pro-/pre-B cell differentiation and mitoflash activation. Although EFhd1 is important for maintaining mitochondrial morphology and energy synthesis, its mechanism of action remains unclear. Here, we report the crystal structure of the EFhd1 core domain comprising a C-terminus of a proline-rich region, two EF-hand domains, and a ligand mimic helix. Structural comparisons of EFhd1, EFhd2, and AIF-1 revealed similarities in their overall structures. In the structure of the EFhd1 core domain, two Zn²⁺ ions were observed at the interface of the crystal contact, suggesting the possibility of Zn²⁺-mediated multimerization. In addition, we found that EFhd1 has Ca²⁺-independent β-actin-binding and Ca²⁺-dependent β-actin-bundling activities. These findings suggest that EFhd1, an actin-binding and -bundling protein in the mitochondria, may contribute to the Ca²⁺-dependent regulation of mitochondrial morphology and energy synthesis.

Ca ²⁺调节几种细胞功能，包括信号传导事件，能量产生和细胞存活。这些细胞过程是由Ca ²⁺介导的-结合蛋白，例如EF-手超家族蛋白。在EF手超家族蛋白中，同种异体移植炎症因子1（AIF-1）和swiprosin-1 / EF手含域蛋白2（EFhd2）是胞质肌动蛋白结合蛋白。AIF-1调节细胞骨架并增加免疫细胞的迁移。EFhd2还是一种涉及免疫细胞激活和脑细胞功能的细胞骨架蛋白。EFhd1是EFhd2的线粒体异卵双胞胎，介导神经元和前B细胞的前/后细胞分化以及线粒体的激活。尽管EFhd1对于维持线粒体形态和能量合成很重要，但其作用机理仍不清楚。在这里，我们报告EFhd1核心域的晶体结构，包括富含脯氨酸的区域的C端，两个EF手域和一个配体模拟螺旋。EFhd1，EFhd2和AIF-1的结构比较表明，它们的整体结构相似。在EFhd1核心结构域的结构中，两个Zn在晶体接触的界面处观察到²⁺离子，表明Zn ²⁺介导的多聚反应的可能性。另外，我们发现EFhd1具有不依赖Ca ^2+的β-肌动蛋白结合和不依赖Ca ^2+的β-肌动蛋白捆绑活性。这些发现表明，EFhd1是线粒体中的一种肌动蛋白结合和捆绑蛋白，可能有助于线粒体形态和能量合成的Ca ²⁺依赖性调节。