X-RAY STRUCTURAL STUDIES OF LACTOFERRIN

乳铁蛋白的 X 射线结构研究

• 批准号: 3319299
• 负责人: EDWARD N BAKER
• 金额: $ 7万
• 依托单位: MASSEY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-05-01 至 1990-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3319299
• 关键词: X ray crystallography; bicarbonates; chemical binding; divalent cations; human milk; iron; lactoferrin; oxaloacetates; point mutation; protein sequence; protein structure function; site directed mutagenesis; transferrin

Our aim is to gain a better understanding of the molecular basis for the biological roles of lactoferrin, the major iron-binding protein in milk, in many other bodily secretions, and in leukocytes. This will address its antibacterial properties, as well as mechanisms of uptake and release o metal ions. It will have important implications for understanding bodily defence mechanisms and in particular the biology of human milk and the bioavailability of trace elements. (i) Specifically, the three-dimensional structure of human lactoferrin will be determined at high resolution by X-ray crystallography. The structure will then be refined, so that the exact nature of the iron sites, their location and accessibility in the protein structure, and the role of functionally important anions (HCO3 ion) can be unambiguously determined. The high resolution analysis will build upon preliminary studies already carried out. Large crystals of lactoferrin, which are stable to X-rays, have been obtained; heavy atom derivatives have been prepared and characterised; and X-ray studies at 6A and 3.5A have given promising results. (ii) Once the high resolution structure has been determined, binding studies will be carried out with other metal ions (eg Zn2+, Cu2+, Ca2+, Cr3+, Mn2+) and anions (eg oxalate) which may bind either in the specific iron sites or in non-specific sites. This will help define the role of lactoferrin in binding trace element in milk, and in influencing their bioavailability. (iii) The structure will also be used to examine structural and functional relationships with other iron-binding proteins, notably serum transferrin, the iron transport protein in plasma, and ovotransferrin. (iv) Apolactoferrin, iron-binding fragments of lactoferrin, and lactoferrins of other species (e.g. bovine) will be examined in comparative studies. (v) As a long-term aim, the structure will be examined through site-directed mutagenesis studies presently being initiated by other researchers.

我们的目标是更好地了解 乳铁蛋白（牛奶中主要的铁结合蛋白）的生物学作用 许多其他身体分泌物和白细胞中。 这将解决其 抗菌性能，以及吸收和释放的机制， 金属离子 这将对理解身体的 防御机制，特别是人乳的生物学， 微量元素的生物利用度。 (i)具体而言，人乳铁蛋白的三维结构将 通过X射线晶体学以高分辨率测定。 结构 然后将被精炼，以便铁网站的确切性质， 蛋白质结构中的位置和可及性，以及 功能上重要的阴离子（HCO 3离子）可以明确地确定。 高分辨率分析将建立在初步研究的基础上， 贯彻 大晶体的乳铁蛋白，这是稳定的X射线， 已经获得;重原子衍生物已经制备， 特征;和X射线研究在6A和3.5A已经给出了有希望的 结果 (ii)一旦确定了高分辨率结构， 将用其它金属离子（例如Zn 2+，Cu 2+，Ca 2+， Cr 3+，Mn 2+）和阴离子（如草酸盐），它们可能结合在特定的 铁位点或非特异性位点。 这将有助于确定 乳铁蛋白与牛奶中微量元素结合及其影响 生物利用度 (iii)该结构还将用于检查结构和功能 与其他铁结合蛋白，特别是血清转铁蛋白， 血浆铁转运蛋白和卵转铁蛋白。 (iv)乳铁蛋白，乳铁蛋白的铁结合片段，和 其他物种（如牛）的乳铁蛋白将在比较中进行检查。 问题研究 (v)作为一个长期目标，该结构将通过以下方面进行审查： 定点诱变研究目前正在启动的其他 研究人员