ACCELERATING IRON CHELATION FROM SERUM TRANSFERRIN

说明(改编自应用程序) 血清转铁蛋白是一种通过血液运输铁的蛋白质。这个 蛋白质由两个相似的叶组成，只有一个铁结合部位 在每个裂片内的裂隙的底部。在铁超载的情况下， 转铁蛋白被铁饱和。然而，铁的兑换率 具有低分子量配体的速度往往非常慢，这严重 限制了治疗性铁络合剂针对这一点的能力 便利的铁质游泳池。 转铁蛋白释放铁的速度显示出对两者的复杂依赖关系。 配体浓度和无机盐浓度 缓冲区。对于许多配体来说，铁的最大释放速率似乎是 受限于缓慢的蛋白质构象变化，但这一过程并不顺利 明白了。一些配体似乎能够通过以下两种方式避免这种限制 利用一条单独的一阶路径，以某种方式避免了 构象门控，或通过结合在特征不佳的变构 加速构象变化的阴离子结合部位。这个 这项研究的主要目标是确定这一机制的细节 转铁蛋白与低分子量配体之间的铁交换 利用这些信息设计新的助剂以加速除铁 生理条件下的转铁蛋白。 拟议的工作将侧重于重组转铁蛋白的研究。 半分子、Tf/2N和Tf/2C以及一系列定点突变体 这些分子中。铁的释放动力学将遵循可见光 和荧光光谱仪。天然和突变体的动力学研究 蛋白质将表征可用于铁释放的途径。两者都有 计算研究和定点突变将被用来定位和 表征变构阴离子结合部位并确定阴离子如何 这个部位的结合能够加速铁的释放。新的化合物将是 人工合成的，旨在避免蛋白质的限制 构象变化要么是通过单独的， 一级途径或通过靶向变构阴离子结合部位 加快门控构象变化的速度。这是一个 有分子生物学专业知识的教员参与的合作项目， 溶液动力学、有机合成和计算化学。

DESCRIPTION (adapted from the application) Serum transferrin is the protein that transports iron through blood. The protein consists of two similar lobes, with a single iron binding site located at the base of a cleft within each lobe. Under conditions of iron overload, transferrin becomes saturated with iron. However, the rate of iron exchange with low molecular weight ligands tends to be very slow, which severely restricts the ability of therapeutic iron chelating agents to target this readily accessible iron pool. The rate of iron release from transferrin shows a complex dependence on both the ligand concentration and the concentration of inorganic salts in the buffer. For many ligands it appears that the maximum rate of iron release is limited by a slow protein conformational change, but this process is not well understood. Some ligands appear to be able to avoid this limitation either by taking advantage of a separate, first-order pathway that somehow avoids the conformational gating, or by binding at a poorly characterized allosteric anion-binding site that accelerates the rate of the conformational change. The primary objectives of this study are to determine the details of the mechanism of iron exchange between transferrin and low molecular weight ligands and to use this information to design new agents for accelerating iron removal from transferrin under physiological conditions. The proposed work will emphasize studies on recombinant transferrin half-molecules, Tf/2N and Tf/2C, as well as a series of site directed mutants of these molecules. The kinetics of iron release will be followed by visible and fluorescence spectroscopies. Kinetic studies on the native and mutant proteins will characterize the pathways available for iron release. Both computation studies and site directed mutagenesis will be used to locate and characterize the allosteric anion binding site and to determine how anion binding at this site is able to accelerate iron release. New compounds will be synthesized that will be designed to avoid the limitations of the protein conformational change either by accelerate iron release through a separate, first-order pathway or by targeting the allosteric anion binding site to accelerate the rate of the gating conformational change. This is a collaborative project involving faculty with expertise in molecular biology, solution kinetics, organic synthesis, and computational chemistry.