The biological activity of metal ion-gastrin complexes

金属离子-胃泌素复合物的生物活性

基本信息

批准号：
6508709
负责人：
GRAHAM S BALDWIN
金额：
$ 13.5万
依托单位：
UNIVERSITY OF MELBOURNE
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：
2002-08-01 至 2005-07-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): The long-term aim of this project is to understand the biological significance of the interactions between metal ions and peptides derived from the prohormone progastrin. Recent data from this laboratory has shown that progastrin-derived peptides (PDPs) selectively bind 2 ferric ions, and that recombinant human progastrin also binds a calcium ion with high affinity, at a site distinct from the ferric ion binding site. The specific aims of the project are: (1) to define the properties and structure of the complexes between metal ions and PDPs, (2) to define the role of metal ions in the biological activities of PDPs including receptor binding, cell proliferation and cell migration, and (3) to determine the role of the complexes in modulation of progastrin processing and in metal ion uptake by the gastrointestinal tract. The health significance of the project lies in the facts that PDPs act as growth factors for the normal gastric and colonic mucosa, accelerate the development of both gastric and colorectal cancer, and may be involved in disorders of iron homeostasis. The research design mirrors the specific aims. Firstly, the structures of the complexes between metal ions and PDPs will be determined by a combination of fluorescence, EPR and NMR spectroscopy. The structures will be used as the basis for the design of recombinant and synthetic PDPs with single amino acid substitutions which prevent the binding of either ferric or calcium ions. Secondly, the ability of the parent and mutant PDPs to bind to, stimulate proliferation in, and reduce adhesion of, a panel of gastrointestinal cell lines will be compared. Proliferation will be assessed by measurement of DNA synthesis in cell lines, isolated crypts, and the defunctioned colon in vivo. Adhesion will be assessed by immunochemical analysis of adhesion protein complexes, and migration and wound healing assays in tissue culture. Thirdly, the effect of changes in ferric ion concentration on progastrin processing will be measured in cell lines and in mice with altered iron status. The role of PDPs in the cellular uptake of ferric ions will also be determined. The demonstration that ferric and calcium ions are essential components of the biologically active forms of PDPs would alter completely our understanding of the role of metal ions in hormone function. The studies may also reveal an unexpected role for peptide hormones in ferric ion homeostasis. In the longer term definition of high affinity receptors for PDP-metal ion complexes may permit the development of novel therapies for treatment of disorders of iron metabolism and of gastrointestinal cancers.

描述（由申请人提供）：该项目的长期目的是了解金属离子和源自激素progastrin的肽之间相互作用的生物学意义。该实验室的最新数据表明，在与高亲和力的位点上，反相结合的人类前进蛋白有选择地结合2种铁离子，而重组人的progastrin也与高亲和力结合了钙离子，该钙离子在不同于基离子结合位点的位点。该项目的具体目的是：（1）定义金属离子和PDP之间复合物的特性和结构，（2）定义金属离子在PDP的生物学活性中的作用，包括受体结合，细胞增殖和细胞。迁移，（3）确定胃肠道摄取中配合物在progastrin加工和金属离子摄取中的作用。该项目的健康意义在于，PDP充当正常胃和结肠粘膜的生长因子，加速胃癌和结直肠癌的发展，并且可能参与铁稳态疾病。研究设计反映了具体目标。首先，金属离子和PDP之间的复合物的结构将由荧光，EPR和NMR光谱的组合确定。这些结构将用作设计重组和合成PDP的基础，并与单个氨基酸取代，从而防止铁离子或钙离子的结合。其次，将比较母体和突变体PDP与结合，刺激加剧并减少粘附的粘附力的能力。通过测量细胞系，分离的隐窝和体内缺失结肠的DNA合成来评估增殖。粘附将通过对粘附蛋白复合物的免疫化学分析以及组织培养中的迁移和伤口愈合分析来评估。第三，将在细胞系和铁状态改变的小鼠中测量铁离子浓度变化对进场蛋白加工的影响。 PDP在铁离子的细胞摄取中的作用也将得到确定。铁和钙离子是PDP的生物活性形式的重要组成部分的证明将完全改变我们对金属离子在激素功能中的作用的理解。这些研究还可能揭示了肽激素在铁离子稳态中的意外作用。在长期的PDP - 金属离子复合物的高亲和力受体的定义中，可以允许开发新的疗法来治疗铁代谢和胃肠道癌的疾病。