STRUCTURAL DETERMINATION OF SHP 1 & PEPTIDE COMPLEXES: IMMUNOLOGY

SHP 1 的结构测定

• 批准号: 6220532
• 负责人: GUANGWEN WAYNE ZHOU
• 金额: $ 1.77万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-08-15 至 2000-08-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6220532
• 关键词: bioimaging /biomedical imaging; biological products; biomedical resource; proteins; radiography; structural biology

Over the past year, we have carried out crystallographic studies of catalytic intermediates supported by the catalytic site of Gia1. The structure of the GDP-bound complex has been determied at a series of Mg2+ concentrations in the presence of sulfate, an analog of inorganic phospate, a product of GTP hydrolysis. These crystal structures, determined with data obtained from the CHESS A1 beam line, demonstrate that restructuring of the switch II helix are concomitant with cleavage of the b-g phosphate bond of GTPgS, GDP and GDPPi complexes using the A1 and F1 beamlines, reveaing the conformation of the protein in all three stable catalytic intermediate states. The complex between RGS4 (Regulator of G protein Signalling) and the GDP-Aluminum fluroride-Mg2+ bound form of Gia1, was also determined at 2.8
resolution using radiation from the F2 line. This represents the first structure of a G protein-G protein GAP (GTPase Activating Protein) complex to be determined, and demonstrates, in part, how RGS4 stabilizes the transition state for GTP hydrolysis. Most recently, the structures of the stimulatory G protein alpha subunit Gsa and its complex with the soluble catalytic domains of its effector, adenylyl cyclase, have been determined. These structures, determined at 2.8 - 2.3
resolutions (for the series of complexes studied) have revealed the mode of interacion between a heterotrimeric G protein and its effector, as well as the binding sites of ATP in the catalytic site of adenylyl cyclase and that of the diterpine activator, forskolin. These studies, enabled with data measured at the CHESS A1 beamlines, represent a milestone in structural studies of G proteins, and suggest for the first time, how heterotrimeric G proteins activate their effectors. Synchrotron radiation was essential to the success of all of the projects described above, due to the weak diffracing power and limiting size of the crystals utilized in each of the experiments

在过去的一年里，我们进行了晶体学研究 由Gia 1的催化位点支持的催化中间体。 GDP结合复合物的结构已被一系列确定 的Mg 2+浓度在硫酸盐的存在下， 无机磷酸盐，GTP水解产物。 这些水晶 结构，用从CHESS A1光束线获得的数据确定， 证明开关II螺旋的重组是伴随的， GTPgS、GDP和GDP的b-g磷酸键断裂Pi 复合物使用A1和F1光束线，揭示了构象 蛋白质处于所有三种稳定的催化中间状态。 的 RGS4（Regulator of G protein Signalling）与 Gia 1的GDP-氟化铝-Mg 2+结合形式也在 2.8
分辨率使用来自F2线的辐射。 这代表 G蛋白的第一结构-G蛋白GAP（GTTT活化 蛋白质）复合物，并证明，部分，如何RGS 4 稳定GTP水解的过渡态。 最近， 刺激性G蛋白α亚基Gsa及其 与其效应物腺苷酸的可溶性催化结构域复合 环化酶，已确定。 这些结构，在2.8 - 2.3
决议（为一系列复杂的研究）已经揭示 异源三聚体G蛋白与其 效应器，以及ATP的结合位点的催化位点， 腺苷酸环化酶和地萜品激活剂，毛喉素。 这些研究利用在CHESS A1光束线上测量的数据进行， 代表了G蛋白结构研究的里程碑，并建议 第一次研究了异三聚体G蛋白如何激活它们的 效应器 同步加速器辐射对所有的成功都至关重要。 在上述项目中，由于分散能力弱， 每个实验中使用的晶体的限制尺寸