Function and mechanism of ClpV, a unique Hsp100 protein of proteobacteria that interacts with eukaryotic cells

HSP100蛋白形成环状低聚物，以一种依赖于ATP的方式重塑目标底物，这一活性对细胞蛋白质质量控制至关重要。大多数Hsp100蛋白(如ClpA、ClpC、ClpX)与一种多肽酶(ClpP)相关，其功能广泛且受调节的蛋白分解。相反，ClpB不与ClpP相互作用，而是与DNAK伴侣系统合作重新激活聚集的蛋白质。这些主要的Hsp100蛋白的功能和机制已被详细研究，但对该蛋白家族的特殊成员知之甚少。我们最近发现了Hsp100蛋白家族中一个独特的成员ClpV，它几乎存在于病原性变形杆菌中，但也存在于共生变形杆菌中。ClpV被组织在一个保守的基因簇中，表现出与In型和IV型分泌系统相似的组织结构。ClpV以及ClpV相关基因簇(CVAC)的成员到目前为止还没有被鉴定。Hsp100蛋白在蛋白质分泌和毒力方面的潜在作用是该蛋白家族令人兴奋的一个全新方面。我们将利用遗传学和生物化学的方法来研究ClpV的功能和机制，主要集中在以下几个方面：1.分析ClpV及其相关基因簇的调控。研究CVAC作为一种潜在的分泌系统，并对ClpV的功能进行分析。分析了ClpV和CVAC在体内的功能，重点分析了它们在毒力中的可能作用。鉴定ClpV底物和伙伴蛋白，这将结合对这种特殊的Hsp100蛋白的详细结构功能分析来进行。

Hsp100 proteins form ring-shaped oligomers, which remodel target Substrates in an ATP dependent manner, an activity that is of central importance for cellular protein quality control. Most Hsp100 proteins (e.g. ClpA, ClpC, ClpX) associate with a peptidase (ClpP) and function in general and regulated proteolysis. In contrast, ClpB does not interact with ClpP but instead cooperates with the DnaK chaperone System in reactivating aggregated proteins. Functions and mechanisms of these major Hsp100 proteins have been studied in detail, however, little is known about specialized members of this protein family.We have recently identified a unique member of the Hsp100 protein family, ClpV, which is almost present in pathogenic but also in symbiotic proteobacteria. clpV is organized in a conserved gene cluster, exhibiting a similar organization as type in and type IV secretion Systems. ClpV, as well as members of the ClpV-associated gene cluster (CVAC) have not been characterized to date. The potential role of an Hsp100 protein in protein secretion and virulence is an exciting and an entirely new aspect for this protein family. We will use genetic and biochemical approaches to investigate ClpV function and mechanism with a major focus on the following aspects:1. Analysis of the regulation of clpV and its associated gene cluster.2. Investigation of the CVAC as a potential secretion System and analysis of ClpV function in this context.3. Analysis of the in vivo function of ClpV and the CVAC with a major focus on their putative roles in virulence.4. Identification of ClpV Substrates and partner proteins, which will be carried out in combination with a detailed structure-function analysis of this specialized Hsp100 protein.