Biophysical and biological characterization of FIH-catalysed post-translational asparaginyl hydroxylation.

FIH 催化的翻译后天冬酰胺酰羟基化的生物物理和生物学表征。

• 批准号: BB/G014124/1
• 负责人: Peter Ratcliffe
• 金额: $ 134.81万
• 依托单位: University of Oxford
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2009
• 资助国家: 英国
• 起止时间: 2009 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FG014124%2F1
• 关键词: Biophysical; biological; characterization; FIH; catalysed

The work we are proposing to do is based on recent insights we have obtained into the way cells sense oxygen. Regulating the delivery of oxygen to tissues is a problem for all organisms - particularly large animals (such as humans) that are composed of many billions of cells. Many human diseases such as heart attacks, strokes, cancer, and anaemia involve compromise of cell function by low oxygen levels (hypoxia). In previous work (important components of which were supported by the BBSRC) we have identified a group of oxygenases (enzymes that catalyse the incorporation of atmospheric oxygen into their substrates) that act as cellular 'oxygen sensors', catalysing the hydroxylation (involving addition of an oxygen atom) of specific amino acid residues in a protein called HIF (hypoxia inducible factor). Hydroxylation destroys and inactivates HIF, but since it requires oxygen this reaction is suppressed in hypoxia, allowing HIF to become activate in hypoxic cells (hence its name). HIF is a transcription factor (a type of regulator of gene expression) and when it is switched on, it regulates a lot of genes that are involved in altering cell metabolism, growing new blood vessels, increasing blood production and other actions that help the body survive hypoxia. The work on HIF has raised a lot of questions as to whether this type of modification (hydroxylation) occurs for other types of protein within cells and what the effects might be. Recently we have found that one of the HIF-hydroxylases enzymes also hydroxylates a very common structural domain in proteins, the ankyrin repeat domain (ARD). ARDs occur in many types of protein with many different functions, such as in transcription, cell signalling, cell structure, ion channels, chromosome integrity and aging, inflammation and differentiation. These findings have opened up a new field of research on this type of protein modification, what it does, how it is regulated by hypoxia, and how it affects the cell's responses to hypoxia. It might also give fundamental clues about how the type of 'oxygen sensing' process that regulates HIF evolved. The work is goal directed and is seeking to address a problem in basic science that is potentially important for a number of different biological and biomedical fields. The two applicants are from different backgrounds (Biology and Chemistry) and are working in a partnership to combine their expertise across several biophysical, biochemical, and biological approaches to find out how common this type of protein modification is, how is affects the physical properties of the protein, and how this alters cell function.

我们提议要做的工作是基于我们最近对细胞感知氧气方式的了解。调节氧气向组织的输送是所有生物体面临的一个问题，尤其是由数十亿个细胞组成的大型动物（例如人类）。许多人类疾病，如心脏病、中风、癌症和贫血，都与低氧水平（缺氧）损害细胞功能有关。在之前的工作中（其重要组成部分得到了 BBSRC 的支持），我们已经鉴定了一组加氧酶（催化大气中的氧气掺入其底物的酶），它们充当细胞“氧传感器”，催化称为 HIF（缺氧诱导因子）的蛋白质中特定氨基酸残基的羟基化（涉及添加氧原子）。羟基化会破坏 HIF 并使 HIF 失活，但由于它需要氧气，因此这种反应在缺氧情况下会受到抑制，从而使 HIF 在缺氧细胞中变得活跃（因此得名）。 HIF 是一种转录因子（基因表达调节剂的一种），当它被打开时，它会调节许多基因，这些基因涉及改变细胞代谢、生长新血管、增加血液产量和其他帮助身体在缺氧中生存的行为。 HIF 的研究提出了很多问题，例如细胞内其他类型的蛋白质是否会发生这种类型的修饰（羟基化）以及可能产生什么影响。最近我们发现，一种 HIF 羟化酶还能羟基化蛋白质中非常常见的结构域，即锚蛋白重复结构域 (ARD)。 ARD 存在于多种具有不同功能的蛋白质中，例如转录、细胞信号传导、细胞结构、离子通道、染色体完整性和衰老、炎症和分化。这些发现开辟了一个新的研究领域，研究这种类型的蛋白质修饰、它的作用、它如何受缺氧调节，以及它如何影响细胞对缺氧的反应。它还可能提供有关调节 HIF 的“氧感应”过程类型如何进化的基本线索。这项工作以目标为导向，旨在解决基础科学中的一个问题，该问题对许多不同的生物和生物医学领域可能很重要。两位申请人来自不同的背景（生物学和化学），他们正在合作，结合他们在几种生物物理、生物化学和生物学方法方面的专业知识，以找出这种类型的蛋白质修饰有多常见，它如何影响蛋白质的物理性质，以及它如何改变细胞功能。

项目成果

Asparagine and aspartate hydroxylation of the cytoskeletal ankyrin family is catalyzed by factor-inhibiting hypoxia-inducible factor.

• DOI: 10.1074/jbc.m110.193540
• 发表时间: 2011-03-04
• 期刊: The Journal of biological chemistry
• 作者: Yang M;Ge W;Chowdhury R;Claridge TD;Kramer HB;Schmierer B;McDonough MA;Gong L;Kessler BM;Ratcliffe PJ;Coleman ML;Schofield CJ
• 通讯作者: Schofield CJ

The FIH hydroxylase is a cellular peroxide sensor that modulates HIF transcriptional activity.

• DOI: 10.1038/embor.2012.9
• 发表时间: 2012-03-01
• 期刊: EMBO REPORTS
• 影响因子: 7.7
• 作者: Masson, Norma;Singleton, Rachelle S.;Sekirnik, Rok;Trudgian, David C.;Ambrose, Lucy J.;Miranda, Melroy X.;Tian, Ya-Min;Kessler, Benedikt M.;Schofield, Christopher J.;Ratcliffe, Peter J.
• 通讯作者: Ratcliffe, Peter J.

Oxygenase-catalyzed ribosome hydroxylation occurs in prokaryotes and humans.

• DOI: 10.1038/nchembio.1093
• 发表时间: 2012-12
• 期刊: Nature chemical biology
• 影响因子: 14.8
• 作者: Ge W;Wolf A;Feng T;Ho CH;Sekirnik R;Zayer A;Granatino N;Cockman ME;Loenarz C;Loik ND;Hardy AP;Claridge TDW;Hamed RB;Chowdhury R;Gong L;Robinson CV;Trudgian DC;Jiang M;Mackeen MM;Mccullagh JS;Gordiyenko Y;Thalhammer A;Yamamoto A;Yang M;Liu-Yi P;Zhang Z;Schmidt-Zachmann M;Kessler BM;Ratcliffe PJ;Preston GM;Coleman ML;Schofield CJ
• 通讯作者: Schofield CJ