Structural analysis of dynamins involved in mitochondrial morphology

除了动力蛋白，其他动力蛋白家族成员还参与了多种基本的细胞过程，包括线粒体的分裂和融合、抗病毒活性、植物细胞板的形成和叶绿体的生物发生。在这些蛋白质中，自组装和齐聚成有序结构是一个共同的特征，对大多数蛋白质来说，是其功能所必需的。虽然有大量关于动力素的信息，但对动力素相关蛋白质的结构性质知之甚少。近年来，我们研究了两个Dynamin家族成员，一个参与线粒体分裂(DNM1)，另一个参与线粒体融合，OPA1。我们正在检验一种假设，即Dynamin家族成员共享共同的作用机制。我们的工作提供了明确的证据，表明动力蛋白相关蛋白具有相似的组装和GTP酶特性。然而，结构上的限制仍然必须允许它们具有广泛多样的生物学功能。 Dynamin蛋白家族由独特的GTP酶组成，参与整个细胞的膜分裂和融合事件。我们的目标是了解这些蛋白质的动态结构特性，并将它们与不同的细胞功能联系起来。动力蛋白对细胞内吞和囊泡活动是必不可少的。更多的动力蛋白家族成员参与了多种基本的细胞过程，包括线粒体的分裂和融合、抗病毒活性、细胞板的形成和叶绿体的生物发生。在这些蛋白质中，自组装和齐聚成有序结构是一个共同的特征，对大多数蛋白质来说，是其功能所必需的。虽然有大量关于动力素的信息，但对动力素相关蛋白质的结构性质知之甚少。 为了确定动力蛋白家族成员之间是否存在共同的作用机制，我们研究了DNM1和OPA1的结构和功能。DNM1是参与线粒体分裂的酵母动力蛋白家族成员，OPA1是参与线粒体融合的人类动力蛋白。 此前，我们与加州理工大学的David Chan博士合作，通过负染和低温电子显微镜检查了OPA1的结构。OPA1(常染色体显性遗传性视神经萎缩)突变可导致遗传性视网膜神经节细胞病变。在细胞中，OPA1已被证明是线粒体膜内融合所必需的，但其作用机制尚不清楚。将OPA1添加到含有心磷脂的脂质体中可提高GTP的水解率，并促进OPA1在脂质周围自组装成螺旋阵列，形成蛋白质-脂质管。目前，我们正在通过螺旋和单粒子图像处理方法来解决OPA1的结构问题。OPA1在GTP水解过程中的构象状态也被检测到。

In addition to dynamin, other dynamin family members have been implicated in a variety of fundamental cellular processes, including mitochondrial fission and fusion, anti-viral activity, plant cell plate formation and chloroplast biogenesis. Among these proteins, self-assembly and oligomerization into ordered structures is a common characteristic and, for the majority, is essential for their function. Although there is a wealth of information regarding dynamin, little is known about the structural properties of dynamin-related proteins. In recent years, we have examined two dynamin family members, one involved in mitochondria fission (Dnm1) and the other involved in mitochondrial fusion, Opa1. We are testing the hypothesis that dynamin family members share a common mechanism of action. Our work provides clear evidence that dynamin related proteins have similar assembly and GTPase properties. However, structural constraints must still allow for their widely diverse biological functions. The dynamin family of proteins consists of unique GTPases involved in membrane fission and fusion events throughout the cell. Our goal is to understand the dynamic structural properties of these proteins and correlate them with their diverse cellular functions. Dynamin is essential for endocytosis and vesiculation events in the cell. Additional dynamin family members have been implicated in a variety of fundamental cellular processes, including mitochondrial fission and fusion, anti-viral activity, cell plate formation and chloroplast biogenesis. Among these proteins, self-assembly and oligomerization into ordered structures is a common characteristic and, for the majority, is essential for their function. Although there is a wealth of information regarding dynamin, little is known about the structural properties of dynamin-related proteins. To determine if a common mechanism of action exists among the dynamin family members, we examined the structure and function of Dnm1, a yeast dynamin family member involved in mitochondria fission and Opa1, a human dynamin involved in mitochondria fusion. Previously, in collaboration with Dr. David Chan from Cal Tech, we examined the structure of OPA1 by negative stain and cryo electron microscopy. Mutations in OPA1 (autosomal dominant optic atrophy) can lead to an inherited neuropathy of the retinal ganglion cells. In the cell, OPA1 has been shown to be essential for the fusion of the inner mitochondrial membranes, but its mechanism of action remains poorly understood. Addition of OPA1 to liposomes containing cardiolipin results in enhanced GTP hydrolysis rate and promotes OPA1 to self-assemble into helical arrays around the lipid, forming protein-lipid tubes. Currently we are solving the structure of OPA1 by helical and single particle image processing methods. The conformational state of OPA1 during GTP hydrolysis is also being examined.