Computational analysis of phosphoinositide signaling

描述（由申请人提供）：我们的长期目标是继续发展计算方法，在分子水平上定量描述导致膜表面大分子蛋白质/脂质复合物形成的物理力量。这些复合物在多种细胞过程中起作用，如界面信号转导。我们对当前提案的总体假设是，静电在磷酸肌醇信号传导中起着普遍作用，并通过磷酸肌醇介导的膜结合和蛋白质和脂质的横向组织表现出来。第一个特定目标是预测磷酸肌苷结合区域的膜相关取向，并定量描述结合配体对这些区域静电特性的影响。描述磷酸肌苷如何改变这些结构域的静电特性对于理解膜结合是如何调节的至关重要：我们假设高负电荷磷酸肌苷的结合中和了配体结合口袋中的碱性基团，并减少了疏水基序插入膜界面的能量屏障。第二个特定目的是预测磷酸肌苷如何影响C2结构域的膜相关取向，从而促进钙介导的细胞过程，如突触囊泡胞吐。我们假设这些相互作用主要是由非特定静电力驱动的。第三个特定目标是模拟膜表面蛋白质和磷酸肌苷之间的横向相互作用，以表征在其组织上形成的多聚体结构。这些复合物在细胞内钙释放和细胞趋化中起关键作用。我们假设，膜表面的负电位和正电位局部区域充当了具有相反静电特性的分子的吸引力盆地，并且静电是形成这些复合物的主要驱动力。我们的计算将为磷酸肌肽结合域的膜结合、膜介导的相互作用以及两者的调节提供实验上可测试的假设。对静电相互作用的全面描述将为今后研究其他重要的相互作用铺平道路。影响膜靶向的突变与癌症和微生物敏感性有关。因此，这项研究将提供对正常和异常功能的深入了解。

DESCRIPTION (provided by applicant): Our long-term goal is to continue the development of computational methods to describe quantitatively, at the molecular level, the physical forces responsible for the formation of macromolecular protein/lipid complexes at membrane surfaces. These complexes function in a variety of cellular processes such as interfacial signal transduction. Our overall hypothesis for the current proposal is that electrostatics plays a general role in phosphoinositide signaling and is manifested through phosphoinositide-mediated membrane association and the lateral organization of proteins and lipids. The first Specific Aim is to predict the membrane-associated orientations of phosphoinositide-binding domains and to describe quantitatively the effect of bound ligand on the electrostatic properties of these domains. Describing how phosphoinositides change the electrostatic character of these domains is crucial to understanding how membrane association is regulated: We hypothesize that binding of the highly negatively charged phosphoinositide neutralizes basic groups in the ligand-binding pocket and reduces the energetic barrier to insertion of hydrophobic motifs into the membrane interface. The second Specific Aim is to predict how phosphoinositides affect the membrane- associated orientations of C2 domains and, thus, contribute to calcium-mediated cellular processes such as synaptic vesicle exocytosis. We hypothesize that these interactions are driven mainly by non-specific electrostatic forces. The third Specific Aim is to simulate the lateral interactions among proteins and phosphoinositides at membrane surfaces in order to characterize the multimeric structures that form upon their organization. Such complexes play key roles in intracellular calcium release and cellular chemotaxis. We hypothesize that localized regions of negative and positive potential at the membrane surface serve as basins of attraction for molecules with opposite electrostatic characteristics and that electrostatics is a major driving force in forming these complexes. Our calculations will provide experimentally testable hypotheses for the membrane association of phosphoinositide binding domains, for membrane-mediated interactions, and for the regulation of both. A full description of electrostatic interactions will pave the way for the future study of other important interactions. Mutations that affect membrane targeting are implicated in cancer and microbial susceptibility. Thus, this research will provide insight into both normal and aberrant functionality.