Structures and functions of the human Josephin domain-containing proteins

含约瑟芬结构域的蛋白是脱泛素酶的五大家族之一。 (配音)。该家族的创始成员是ataxin-3，一种多谷氨酰胺蛋白，也是 最常见的遗传性共济失调，脊髓小脑性共济失调3型(SCA3，也称为Machado-Joseph 疾病)。阿泰素-3‘S配音活性位于该分子的N-末端，即所谓的约瑟芬结构域。 Aaxin-3与许多与维持蛋白质动态平衡相关的细胞过程有关，包括 错误折叠的蛋白质到侵袭体的运输和蛋白质通过内质网的流动- 相关降解途径(ERAD)。Aaxin-3还可以抑制扩展的重复序列的毒性 多聚谷氨酰胺结构域。所有这些功能都需要配音活性，从而确立了 Josephin结构域，并暗示Josephin Dub活性与 蛋白质错误折叠。 在人类基因组中编码了三种ataxin-3的同源物：ataxin-3样蛋白(AT3L)， 约瑟芬-1和约瑟芬-2。这三种催化剂都具有相似活性，但在催化效率和 专一性。含有Josephin结构域的同源物在大多数真核生物中都是保守的，包括植物和 原生动物，这意味着这个结构域和它所表达的DUB活动是有价值的生物学 功能。 我们建议分析含有人约瑟芬结构域的分子结构和功能 以了解它们如何识别和作用于底物和其他相互作用的蛋白质。我们 将使用X射线晶体学来确定不同约瑟芬结构域的结构，无论是单独的还是复杂的 与泛素底物结合；表征四个人约瑟芬结构域的DUB活性 蛋白质，评估它们如何结合和切割小泛素和大泛素底物；探索它们的泛素- 结合活性；以及探索ataxin-3与其细胞结合伙伴的相互作用。

The Josephin domain-containing proteins are one of the five major families of deubiquitinating enzymes (DUBs). The founding member of the family is ataxin-3, a polyglutamine protein and the causative agent of the most common inherited ataxia, spinal cerebellar ataxia type 3 (SCA3, also known as Machado-Joseph disease). Ataxin-3's DUB activity resides in the molecule's N-terminal half, the so-called Josephin domain. Ataxin-3 is linked to many cellular processes that are related to maintaining protein homeostasis, including the transport of misfolded proteins to aggresomes and the flux of proteins through the endoplasmic reticulum- associated degradation pathway (ERAD). Ataxin-3 can also suppress the toxicity of expanded repeat polyglutamine domains. All of these functions require DUB activity, establishing the biological significance of the Josephin domain and hinting at a link between Josephin DUB activity and neurodegeneration induced by protein misfolding. Three homologs of ataxin-3 are encoded in the human genome: The ataxin-3-like protein (AT3L), Josephin-1, and Josephin-2. All three possess DUB activity, but differ substantially in catalytic efficiency and specificity. Josephin domain-containing homologs are conserved in most eukaryotes, including plants and protozoans, implying that this domain and the DUB activity it expresses are responsible for valuable biological functions. We propose to analyze the molecular structure and function of the human Josephin domain-containing proteins in order to understand how they recognize and act upon substrates and other interacting proteins. We will use X-ray crystallography to determine structures for different Josephin domains, alone and in complex with ubiquitin substrates; characterize the DUB activities of the four human Josephin domain-containing proteins, assessing how they bind and cleave both small and large ubiquitin substrates; explore their ubiquitin- binding activities; and probe the interactions of ataxin-3 with its cellular binding partners.