PEPTIDES BASED ON ACTIVE SITES OF ACTIN-BINDING PROTEINS

拟议合作的总体目标是提供信息， 关于运动细胞如巨噬细胞的结构变化 血小板和淋巴细胞是在这些细胞响应于 细胞外信号 细胞运动对于这种生理过程是必要的。 如吞噬作用，凝血过程中的血小板反应， 伤口愈合和神经发育。 了解链接 细胞信号和运动之间的联系对于理解 导致运动受损或失控的缺陷， 增殖 这项工作的重点是actin顺从性切断 蛋白质凝溶胶蛋白、绒毛蛋白和其他肌动蛋白结合蛋白， 由聚磷酸肌醇（PPI）调节的profilin。 聚磷酸肌醇是细胞中普遍存在的磷脂成分 膜的磷酸化或水解，由几个不同的 酶参与许多信号转导途径。 母 这项合作所基于的项目涉及生物化学和 肌动蛋白和其他蛋白质网络的生物物理研究，旨在 确定这些网络是如何重塑的，以及它们的重塑如何 参与到运动的产生中。 此扩展将允许详细的 分析聚磷酸肌醇结合的蛋白质上的位点， 由这种结合引起的蛋白质结构变化。 本提案的具体目的是审查 具有基于以下序列的合成肽的聚磷酸肌醇 在蛋白质内，通过研究， 蛋白水解衍生的蛋白片段或突变体。 的优点 研究小的合成肽是，它们的结构可以 比完整的蛋白质更严格地研究，在某些情况下， 溶液结构可以完全通过NMR和其它方法确定 方法如在父母补助金中所讨论的。 肽合成需要 专业的技能和设备， 研究细胞生物学或肽的影响所需的，因为 例如，凝溶胶蛋白/PPI相互作用，因此， 不提供肽合成。 有机会与 一个专门研究多肽的有机合成实验室 可以应用于研究调节的实验范围 肌动蛋白结合蛋白，并将增加精确度， 可以确定PPI调节蛋白质所必需的位点。 的 后一种可能性可能与设计有关， 调节PPI代谢或细胞骨架的药物 体内重塑。

The overall goal of the proposed collaboration is to provide information about how structural changes within motile cells such as macrophages, platelets and lymphocytes are achieved when these cells respond to extracellular signals. Cell motility is necessary for such physiologic processes as phagocytosis, platelet response during blood coagulation, wound healing, and neurologic development. Understanding the link between cell signalling and motility is important for understanding the defects that causes impaired or uncontrolled movements and proliferation. This work focuses on the actin filament-severing proteins gelsolin, villin, and other actin-binding proteins such as profilin that are regulated by polyphosphoinositides (PPI's). Polyphosphoinositides are ubiquitous phospholipid components of the cell membrane whose phosphorylation or hydrolysis by several different enzymes is involved in many signal transduction pathways. The parent project on which this collaboration is based involves biochemical and biophysical studies of actin and other protein networks, designed to determine how such networks are remodeled and how their remodeling may be involved in producing motion. This extension will allow detailed analysis of the site on proteins to which polyphosphoinositides bind and of the structural changes within the protein caused by this binding. The specific aim of this proposal is to examine interactions of polyphosphoinositides with synthetic peptides based on the sequences within proteins implicated as potential PPI-binding sites by studies of proteolytically-derived protein fragments or mutants. The advantage of studying small synthetic peptides is that their structures can be studied more rigorously than intact proteins, and in some cases their solution structures can be completely determined by nmr and other methods as discussed in the parent grant. Peptide synthesis requires specialized skills and equipment that are very different from those required to study cell biology or the effects of the peptides on, for example, gelsolin/PPI interactions, and therefore the parent grant made no provision for peptide synthesis. The opportunity to collaborate with a laboratory of organic synthesis that specializes in peptides expands the range of experiments that can be applied to study the regulation of actin-binding proteins and will increase the precision with which the sites necessary for protein regulation by PPI's can be defined. The latter possibility could potentially be relevant to the design of pharmacologic agents that modulate PPI-metabolism or cytoskeletal remodeling in vivo.