Functional studies of the stressosome

应激体的功能研究

• 批准号: BB/F001533/1
• 负责人: Richard Lewis
• 金额: $ 44.31万
• 依托单位: Newcastle University
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2008
• 资助国家: 英国
• 起止时间: 2008 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FF001533%2F1
• 关键词: Functional; studies; stressosome

For survival, all organisms must respond to changes in their environment. One of the most important ways by which all organisms signal to themselves that a response is required to a change in growth conditions is mediated by protein phosphorylation. Here, the covalent modification of regulatory proteins by phosphorylation changes the properties of the target protein and hence its functions. Phosphorylation is performed by enzymes called kinases, which require ATP as a co-factor for the phosphorylation reaction. The stress response of the bacterium Bacillus subtilis and its close relatives is also regulated by phosphorylation. The response to stress provides the cell with a protective mechanism against a wide range of chemical and physical insults. Ultimately the stress signaling pathway controls the activity of the molecular machine, RNA polymerase, which is required to initiate the first step in the synthesis of the proteins that together act to provide the cell with its resistance against the imposed stress. Key to the information-processing pathway that indicates to the cell that its environment is deteriorating is a large structure called the 'stressosome'. The stressosome is composed of several proteins that together assembles the stressosome into a small virus-like structure. The proteins in this 'mini-virus' act in concert to trap a key kinase prior to stress and then to release it on stress to act in the signaling system that ultimately activates RNA polymerase. We are currently determining the structure of the stressosome in a variety of functional states and these on-going studies are challenging us to answer questions that the structures pose. These answers will only be obtained by functional studies using classical biochemical approaches in combination with bacterial cell biology techniques. For instance, we need to establish why the stressosome exists in the first place - perhaps its purpose is to provide a response to stress that is much greater in magnitude than the stress signal to guarantee that the health of the cell is maintained. We will aim to destroy the formation of the stressosome complex to see the impact on this disruption on the ability of the bacterial cell to respond appropriately to stress. Furthermore, there are missing components in the stress-signalling pathway, for instance how is stress actually perceived by a single-celled organism? Clues to the cellular location and identity of stress-receptors may be found by tagging the key signalling kinase with a protein that will colour the kinase green and enable us to track its location using a microscope during the cell cycle and during the response to stress. Finally, stressosome proteins are also found in many other bacteria and they are proposed to, and we will demonstrate that they do play roles regulating other important cellular processes.

为了生存，所有生物都必须对环境的变化做出反应。所有生物体通过蛋白质磷酸化介导的最重要的方式之一向自己发出信号，要求对生长条件的变化做出反应。在这里，通过磷酸化对调节蛋白的共价修饰改变了靶蛋白的性质，从而改变了其功能。磷酸化由称为激酶的酶进行，其需要ATP作为磷酸化反应的辅因子。枯草芽孢杆菌及其近亲的应激反应也受磷酸化调节。对压力的反应为细胞提供了一种保护机制，以抵御各种化学和物理伤害。最终，应激信号传导途径控制分子机器RNA聚合酶的活性，RNA聚合酶是启动蛋白质合成的第一步所必需的，这些蛋白质共同作用为细胞提供对施加的应激的抵抗力。信息处理途径的关键是向细胞表明其环境正在恶化，这是一个称为“应激体”的大型结构。应激体由几种蛋白质组成，这些蛋白质一起将应激体组装成小的病毒样结构。这种“微型病毒”中的蛋白质在应激前协同作用以捕获关键激酶，然后在应激时释放它，以作用于最终激活RNA聚合酶的信号系统。我们目前正在确定各种功能状态下应激体的结构，这些正在进行的研究正在挑战我们回答这些结构提出的问题。这些答案只能通过使用经典生物化学方法结合细菌细胞生物学技术的功能研究来获得。例如，我们首先需要确定为什么应激体存在--也许它的目的是提供一种比应激信号大得多的应激反应，以保证细胞的健康得到维持。我们的目标是破坏应激体复合物的形成，以观察这种破坏对细菌细胞适当应对应激的能力的影响。此外，压力信号通路中还缺少一些组件，例如，单细胞生物实际上是如何感知压力的？通过用一种蛋白质标记关键的信号激酶，可以发现应激受体的细胞定位和身份的线索，这种蛋白质将激酶染成绿色，使我们能够在细胞周期和应激反应期间使用显微镜跟踪其位置。最后，在许多其他细菌中也发现了应激体蛋白，我们将证明它们确实在调节其他重要的细胞过程中发挥作用。

项目成果

Crystallization and preliminary X-ray analysis of RsbS from Moorella thermoacetica at 2.5 A resolution.

来自热乙酸莫尔氏菌的 RsbS 的结晶和初步 X 射线分析，分辨率为 2.5 A。

• DOI: 10.1107/s1744309108003849
• 发表时间: 2008
• 期刊: Acta crystallographica. Section F, Structural biology and crystallization communications
• 作者: Quin M