Mechanism of Action of sHSP Chaperones in Protection from Oxidative Stress

描述（由申请人提供）：分子伴侣是由细胞应激激活并有助于防止错误折叠蛋白质聚集的蛋白质。小分子热休克蛋白伴侣蛋白家族（small heat shock protein chaperone family，sHSPs）是一类普遍存在的分子伴侣蛋白，其序列、大小和底物特异性均不均一，但具有保守的结合部分变性底物并以ATP非依赖性方式阻止其不可逆聚集的功能。本研究的长期目标是阐明sHSP分子伴侣的作用机制，并确定sHSP在氧化应激保护中的作用。我将解决的基本分子机制sHSP行动在氧化应激使用的模式生物，蓝藻，集胞藻PCC 6803。集胞藻只有一种sHSP，即Hsp16.6，它被证明是获得耐热性所必需的。重要的是，我最近的数据表明Hsp16.6也是氧化应激期间生存所必需的。我将在体外进行生化研究，使用纯化的重组sHSP，以揭示sHSP功能的机制细节，并在平行确定潜在的底物保护的sHSP在体内氧化应激。具体来说，我将1）确定是否有氧化应激诱导的热休克蛋白16.6的构象变化，2）确定是否sHSP可以保护酶不可逆的氧化和失活，和3）确定蛋白质与热休克蛋白16.6合作，在保护氧化应激。这些基础研究将更好地了解sHSPs在细胞氧化还原平衡中的作用，以及sHSPs如何影响广泛的人类疾病，包括神经退行性疾病，心血管疾病和癌症。

DESCRIPTION (provided by applicant): Molecular chaperones are proteins that are activated by cellular stress and assist in preventing the aggregation of misfolded proteins. The small heat shock protein chaperone family (sHSPs) is a ubiquitous class of molecular chaperones that are heterogeneous in sequence, size, and substrate specificity but have a conserved function to bind partially denatured substrates and prevent their irreversible aggregation in an ATP-independent manner. The long term goals of my research are to elucidate the mechanism of sHSP chaperone action and to define the role of sHSPs in protection from oxidative stress. I will address the basic molecular mechanism of sHSP action during oxidative stress using a model organism, the cyanobacterium, Synechocystis PCC 6803. Synechocystis has only one sHSP, Hsp16.6, which has been shown to be required for the acquisition of thermotolerance. Importantly, my recent data demonstrate Hsp16.6 is also necessary for survival during oxidative stress. I will conduct biochemical studies in vitro using purified recombinant sHSPs to uncover mechanistic details of sHSP function and in parallel identify potential substrates protected by an sHSP during oxidative stress in vivo. Specifically I will 1) determine if there are conformational changes in Hsp16.6 induced by oxidative stress, 2) Determine if sHSPs can protect enzymes from irreversible oxidation and inactivation, and 3) Identify proteins that cooperate with Hsp16.6 during protection from oxidative stress. These fundamental studies will provide a better understanding of the role of sHSPs in cellular redox equilibrium and how sHSPs impact a vast spectrum of human diseases including neurodegenerative disorders, cardiovascular disease, and cancer.