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STRUCTURE AND FUNCTION OF IMMUNOPHILIN

亲免素的结构和功能

基本信息

批准号：
2633522
负责人：
Hengming Ke
金额：
$ 18.53万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
1994
资助国家：
美国
起止时间：
1994-01-01 至 1998-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2633522
关键词：
X ray crystallography cis trans isomerization computer program /software computer simulation crystallization cyclosporines drug receptors enzyme complex enzyme mechanism enzyme structure enzyme substrate enzyme substrate complex immunosuppressive isozymes peptidylprolyl isomerase physical model proline structural biology

项目摘要

Cytoplasmic signal transduction during T cell activation is mediated by a complex of calcineurin and cyclophilin (CyP). CyP is a binding protein for the immunosuppressive drug cyclosporine A (CsA) and also an enzyme catalyzing the cis yields trans isomerization of peptidyl-prolyl bonds. Calcineurin, a seine/threonine phosphatase has been recently identified as a receptor of the CyP-CsA complex in vitro. This proposal is aimed at structural studies of mammalian CyPs and their complexes with substrates and CsA by X-ray protein crystallography, including: (I) determination of three-dimensional structures of human CyP A complexed with CsA and the CsA derivatives of dimethyl-Bmt1-CsA and MeAla6-CsA (II) determination of three-dimensional structures of human CyP A complexed with three proline substrates of Ser-Pro, His-Pro, and Ala-Ala-Pro-Phe, (III) structural comparison between CyP-CsA and CyP-substrate, (IV) determination of the three-dimensional structure of human CyP B, and (V) determination of the three-dimensional structure of murine CyP C. These studies will present a structural basis for cytoplasmic signal transduction in T cell activation, provide insight into the mechanism of peptidyl-prolyl isomerization and its relationship to immunosuppression, and serve as a guideline for the design of new immunosuppressive drugs. Routine methods will be used for crystallization (dialysis or vapor diffusion), preliminary characterization of crystals (precession and still photos), and heavy atom derivative preparation. Diffraction data will be collected on either a Hamlin multiwire detector or a phosphate image plate in our laboratory. Crystal structures will be determined by the molecular replacement or multiple isomorphous replacement method. Models will be built in an ESV10 graphic system and refined by the PROLSQ or XPLOR program.

T细胞活化过程中的胞质信号转导由钙调神经磷酸酶和亲环蛋白（CyP）的复合物。CyP是一种结合蛋白免疫抑制药物环孢素A（CsA）和一种酶催化顺式产生肽基-脯氨酰基键的反式异构化。钙调神经磷酸酶是新近发现的一种丝氨酸/苏氨酸磷酸酶作为CyP-CsA复合物的受体。该提案旨在在哺乳动物CyPs及其与底物和CsA的X射线蛋白质晶体学，包括：（I）人CyP A复合物三维结构的测定与CsA和二甲基-Bmt 1-CsA和MeAla 6-CsA的CsA衍生物（II）人CyP A复合物三维结构的测定用三种脯氨酸底物Ser-Pro、His-Pro和Ala-Ala-Pro-Phe， (III)CyP-CsA和CyP-底物之间的结构比较，（IV）确定人CyP B的三维结构，和（V）小鼠CyP C的三维结构的测定。这些研究将为胞质信号提供结构基础转导的T细胞活化，提供深入了解的机制，肽基脯氨酰异构化及其与免疫抑制的关系，并作为设计新的免疫抑制药物的指南。常规方法将用于结晶（透析或蒸汽扩散），晶体的初步表征（旋进和静态照片）和重原子衍生物制备。衍射数据将在哈姆林多线探测器或磷酸盐探测器上收集成像板。晶体结构将由以下决定：分子置换或多个同晶置换法。模型将在ESV 10图形系统中构建，并由PROLSQ进行细化或XPLOR程序。