SULFATION OF HUMAN PLASMA PROTEIN

酪氨酸残基的硫化是一种生物合成修饰 包括第四组分在内的具有重要生理意义的血浆蛋白 补体、凝血因子V和凝血因子VIII、纤维连接蛋白和α2- 抗纤溶酶。在其中的几个例子中，人们认识到他们的 硫酸盐化对活性有很大影响。我们假设这些地点 硫酸盐化通常作为识别元素和接触点 蛋白质之间的相互作用。目前的提案对此进行了审查 假说，并扩展了我们以前在生物合成和 血浆蛋白中酪氨酸硫酸盐残基的功能。目标 1)确定酪氨酸硫酸盐残基在相互作用中的作用 凝血、纤溶和补体成分，2)检查 酪氨酸硫酸盐化的化学计量和生理变化，以及 3)识别指导特定硫化的识别信号 网站。 硫酸盐基团在血浆蛋白与蛋白质相互作用中的作用 凝血酶、纤溶酶和C1S亚组分 补体的研究将通过比较硫酸盐化的活性和 非硫化形式的蛋白质和与之相对应的合成肽 硫酸盐化的地点。将对活动进行功能评估 补体和凝血试验以及纯化的酶。这个 多肽与蛋白质结合的亲和力和热力学 将通过滴定量热法进行测量。目前还不清楚是否 在硫酸盐化过程中存在生理或病理变化 蛋白质。这个问题将通过测量酪氨酸硫酸盐来研究 不同性别人群几种血浆蛋白和尿液含量的变化 个人。将通过免疫亲和来纯化特定的蛋白质 氨基酸法测定硫酸酪氨酸含量及其方法 碱性水解物的分析或所产生的多肽的高效液相分析 通过蛋白质分解或化学裂解。一系列合成肽 将作为酪氨酸蛋白磺基转移酶的底物进行研究 确定如何确定底物位置，以提供最佳 用于酶的测定和亲和纯化的底物，以及 作为设计抑制剂的基础。长期目标是 理解硫酸盐化的生理意义和变化 蛋白质中的酪氨酸残留量。此外，这项工作将提供 关于凝血和补体相互作用的基本信息 组件。

Sulfation of tyrosine residues is a biosynthetic modification of physiologically important plasma proteins including the fourth component of complement, coagulation factors V and VIII, fibronectin, and alpha2- antiplasmin. In several of these examples, it is recognized that their sulfation has major effects on activity. We hypothesize that sites of sulfation often serve as recognition elements and points of contact in protein-protein interactions. The current proposal examines this hypothesis and extends our previous work on the biosynthesis and function of tyrosine sulfate residues in plasma proteins. Objectives are: 1) To define the roles of tyrosine sulfate residues in interactions of coagulation, fibrinolytic, and complement components, 2) To examine the stoichiometry and physiological variation of tyrosine sulfation, and 3) To identify recognition signals that direct the sulfation of specific sites. Contributions of sulfate groups to interactions of plasma proteins with the proteinases thrombin, plasmin, and the C1s subcomponent of complement will be studied by comparing activities of sulfated and nonsulfated forms of proteins and of synthetic peptides corresponding to sites of sulfation. Activities will be assessed with functional complement and coagulation assays and with purified enzymes. The affinities and thermodynamics of the binding of peptides to proteins will be measured by titration calorimetry. It is not known whether there is physiological or pathological variation in the sulfation of proteins. This issue will be studied by measuring the tyrosine sulfate content of several plasma proteins and of urine from different individuals. Specific proteins will be purified by immunoaffinity methods and their tyrosine sulfate content assessed by amino acid analysis of base hydrolysates or by HPLC analysis of peptides produced by proteolytic or chemical cleavage. A series of synthetic peptides will be studied as substrates of tyrosylprotein sulfotransferase to determine how substrate sites are identified, to provide optimal substrates for assay and affinity purification of the enzyme, and to serve as a basis for designing inhibitors. Long term goals are to understand the physiological significance and variation of the sulfation of tyrosine residues in proteins. In addition, this work will provide basic information about interactions between coagulation and complement components.