Function & Purification of the Fanconi Anemia Protein C*

功能

基本信息

批准号：
6623345
负责人：
GARY M KUPFER
金额：
$ 10.08万
依托单位：
UNIVERSITY OF VIRGINIA
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-07-01 至 2007-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6623345
关键词：
DNA damage Mammalia binding proteins blood proteins chemical conjugate chemical stability congenital aplastic anemia mitomycin C phosphorylation posttranslational modifications protein isoforms protein localization protein protein interaction protein purification protein structure function radiotracer tissue /cell culture

项目摘要

DESCRIPTION (provided by applicant): Fanconi anemia (FA) is a genetic disease with defects in development and hematopoiesis and propensity to cancer, indicating a vital and basic cell biology process at work. The hallmark of FA is genomic instability, evidenced bv gross chromosomal breakage and DNA alkylating agent hypersensitivity, which correlates with cancer susceptibility in general. Studies of FA are important in several ways. First, FA biology is involved across a spectrum of scientific disciplines. Including hematology oncology, and development. Second, since the known FA proteins are found only in mammalian cells and have no previously described protein domain, their study will yield the description of a novel pathway which promotes the maintenance of genomic stability. Third, work on other cancer susceptibility syndromes have proved to have wide applicability in science in general and cancer in particular, such as Li- Fraument syndrome (p53), ataxia telangiectasia (P13 kinase), and xeroderma pigmentosum (DNA nucleotide excision repair). Fourth, basic work on FA has already led to clinical use of reagents for diagnosis and genetic counseling, and gene therapy trials are currently underwav for treatment of FA. The focus of the grant is to determine the specific biochemical nature of the FA pathway. The emphasis is on the FA proteins, which are encoded by the FA genes, accounting for 5 of at least 7 complementation groups. The FA proteins bind together in a protein complex which forms and is nuclear in 6 of the 7 FA complementation groups, which indicates its importance in FA biology. Our hypothesis is that the FA nuclear protein complex functions as a multimeric complex which is regulated by postranslational phosphorylation. Specifically the purpose of this grant is to isolate the complex in order to purify additional binding proteins of the FA complex to lend an idea to the overall function of the FA complex. In my preliminary, studies I have isolated FA binding proteins whose interactions with FA proteins will be confirmed and whose hypothetical functions will be tested in the work proposed. In addition we will demonstrate how modifications of proteins and of the size of the FA protein complex regulates its function and localization. Our recent work has shown that the FancG protein becomes phosphorylated at mitosis and the entire complex exits condensed chromosomes. We will map the phosphorylation site, test for the kinase involved and assess the changes in complex size in response to multiple stimuli. Identification of new proteins and elucidation of FA pathway mechanisms promise to shed light on a novel area of cancer biology with the potential to provide direct clinical applicability.

描述（由申请人提供）：范可尼贫血（FA）是一种发育和发育缺陷的遗传性疾病。造血和癌症倾向，表明一个重要的基本细胞工作中的生物学过程。 FA 的标志是基因组不稳定性，已被证实由于染色体严重断裂和 DNA 烷化剂过敏，一般来说，与癌症易感性相关。 FA的研究很重要在几个方面。首先，FA 生物学涉及多个领域科学学科。包括血液肿瘤学和发育学。其次，由于已知的 FA 蛋白仅存在于哺乳动物细胞中并且具有之前没有描述过蛋白质结构域，他们的研究将产生描述促进维持基因组稳定性的新途径。第三，其他癌症易感性综合征的研究已被证明具有广泛的应用前景。在一般科学和特别是癌症中的适用性，例如 Li- Fraument 综合征 (p53)、毛细血管扩张性共济失调（P13 激酶）和干皮病色素沉着（DNA 核苷酸切除修复）。四、FA基础工作已经导致诊断和遗传咨询试剂的临床使用，目前正在进行治疗 FA 的基因治疗试验。焦点资助的目的是确定 FA 的具体生化性质途径。重点是 FA 蛋白，它是由 FA 编码的。基因，占至少 7 个互补组中的 5 个。 FA蛋白结合在一起形成蛋白质复合物，该复合物形成 7 种 FA 中的 6 种并成为核互补组，这表明了它在 FA 生物学中的重要性。我们的假设 FA 核蛋白复合物作为多聚体发挥作用复合物，受翻译后磷酸化调节。具体来说这笔赠款的目的是隔离该复合物以纯化 FA 复合物的其他结合蛋白为整体提供了思路 FA 复合体的功能。在我的初步研究中，我分离出了 FA 与 FA 蛋白相互作用的结合蛋白将被证实其假设的功能将在拟议的工作中得到测试。此外我们将演示如何修改蛋白质和 FA 的大小蛋白质复合物调节其功能和定位。我们最近的工作有表明 FancG 蛋白在有丝分裂时被磷酸化，整个细胞复合体退出浓缩染色体。我们将绘制磷酸化位点图，测试所涉及的激酶并评估复合物大小的变化对多种刺激的反应。新蛋白质的鉴定和 FA 途径机制的阐明有望揭示癌症生物学的一个新领域，并有可能提供直接的临床应用。