MOLECULAR INTERACTIONS AT THE CELL SURFACE

细胞表面的分子相互作用

• 批准号: 3271101
• 负责人: PAUL B SIGLER
• 金额: $ 20.3万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-01-01 至 1990-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3271101
• 关键词: atomic absorption spectrometry; calcium binding protein; computer graphics /printing; enzyme complex; enzyme inhibitors; enzyme substrate analog; hormone binding protein; hormone receptor; membrane activity; membrane lipids; membrane proteins; phospholipase A2; phospholipids; stereochemistry; steroid delta isomerase; steroid hormone metabolism

Structure/function analysis will continue on the interaction of proteins with the surface of membranes and with steroids. Both processes figure heavily in the control of intracellular metabolism; the former through the enzymatic release of second mssengers from the cytosolic leaflet of the plasma membrane; the later through steroid activation of genetic control elements. Our principal tool will be high resolution crystallography of the appropriate proteins and of their physiologically important complexes. Phosphlipase A2 (PLA2) offers an ideal model for studying the interaction of proteins with membrane surfaces since it prefers to attack phospholipid head groups if they are in membranes or membrane-like aggregates. Despite several high resolution crystal structures of PLA2 we do not yet understand its action. To resolve this issue we will continue to systematically build a stereochemical picture of the enzyme's functional interactions. The steps are: (1) To complete the 1.7 A refinement of the ligand free form of C. atrox PLA2; (2) To solve the high resolution crystal structure of the calcium complex of C. atrox of PLA2; (3) To solve the high resolution crystal structure of crotoxin, a phosphlipiasic neurotoxin whose action is targeted to the presynaptic membrane; (4) To prepare crystals of PLA2 in complexes with phosphlipid analogues that emulate the interaction between PLA2 and phosphlipids in membrane-like aggregates. This requires the synthesis of stable analogues in which the head groups are constrained to resemble those in an aggregate, but that are small enough to cocrystallize with the enzyme; (5) To explore the preparation of crystalline intracellular phospholipases that mediate internal cell responses. As a first model for protein steroid interaction, we will interpret the 2.5 A map of delta 5-KSI and contrast the structure with that of its isomorphous steroid inhibitor complex. Once refined, these structures should reveal the enzyme's mechanism of specific steroid binding and catalysis. We hope to prepare crystals of the rather plentiful human sex steroid binding globulin and ultimately to crystallize estrogen and progesterone receptors.

结构/功能分析将继续进行蛋白质的相互作用 膜表面和类固醇。 两个过程都数字 严重控制细胞内代谢；前者通过 酶促释放第二名MSSENGERS从胞质小叶 质膜；后来通过遗传控制的类固醇激活 元素。 我们的主要工具将是高分辨率的晶体学 适当的蛋白质及其生理上重要的复合物。 磷脂酶A2（PLA2）提供了研究相互作用的理想模型 具有膜表面的蛋白质，因为它更喜欢攻击磷脂 头部组在膜或膜状聚集体中。 尽管 我们尚不了解PLA2的几种高分辨率晶体结构 它的行动。 为了解决这个问题，我们将继续系统地构建 酶功能相互作用的立体化学图片。 这 步骤是：（1）完成1.7的不含配体形式的改进 C. Atrox Pla2; （2）解决高分辨率的晶体结构 Pla2的C. aTrox的钙复合物； （3）解决高分辨率 Crotoxin的晶体结构，一种磷脂神经毒素的作用是 针对突触前膜； （4）准备pla2晶体 与磷脂类似物的复合物，模仿 膜状聚集体中的PLA2和磷脂。 这需要 稳定类似物的合成，其中头部组约束至 类似于聚集的人，但足够小，可以共结合 用酶； （5）探索结晶的制备 介导内部细胞反应的细胞内磷脂酶。 作为蛋白质类固醇相互作用的第一个模型，我们将解释2.5 Delta 5-Ki的地图，并将结构与其结构与之对比 异态类固醇抑制剂复合物。 一旦完善，这些结构 应揭示酶的特定类固醇结合机制和 催化。 我们希望准备相当丰富的人性的晶体 类固醇结合球蛋白，最终使雌激素结晶 孕酮受体。