ROLES OF PHOSPHOLIPID AND CA2+ IN BRAIN

此续订申请的总体目标主要包括 在分子、免疫学、药理学和调控方面 新近发现的磷脂敏感的钙依赖蛋白激酶 脑内(Pl-Ca-PK)系统。具体目标如下：(1) PL-Ca-PK，这种酶。将努力改进对水的提纯 从猪或牛脑中提取的酶。同样，单克隆化的生产 这种酶的抗体将会得到改善，而且这种酶的分布 大脑中的酶将通过免疫细胞化学进行研究。(2)内生性 底物。髓鞘碱性蛋白(MBP)的磷酸化位点 将测定Pl-Ca-Pk。一旦这样做了，最小的结构 将探索PL-Ca-PK底物专一性的决定因素 使用合成的寡肽。MBP和MBP的磷酸化作用 其片段或多肽对其诱导能力或能力的影响 将探索实验性变态反应性脑脊髓炎。除了……之外 MBP，大脑中PL-Ca-PK的其他一些内源性底物将是 进一步调查。(3)PL-Ca-PK系统的调控。由以下人士订立的规例 各种试剂对酶的活性、磷酸化状态的影响 内源性底物与PL-Ca-PK在脑内可能的“移位” 将会被调查。(4)一种新的S-100调节的蛋白质磷酸化 系统。S-100调节酶(蛋白激酶X)及其19K磁共振 大鼠脑中的底物蛋白将进一步纯化和鉴定。 希望这项提案将产生关于以下方面的新知识 磷脂、钙离子和S-100对脑生物学和病理学的调节 蛋白质通过蛋白质磷酸化反应。

The overall goals of this renewal application cover primarily the molecular, immunological, pharmacological and regulatory aspects of the recently-identified phospholipid-sensitive Ca2+-dependent protein kinase (PL-Ca-PK) system in brain. The specific aims are as follows: (1) PL-Ca-PK, the enzyme. Attempts will be made to improve the purification of the enzyme from pig or bovine brain. Similarly, production of monoclonal antibodies to the enzyme will be improved, and the distribution of the enzyme in brain will be studied immunocytochemically. (2) Endogenous substrates. The sites of phosphorylation in myelin basic protein (MBP) by PL-Ca-PK will be determined. Once this is done, the minimal structural determinants of the substrate specificity for PL-Ca-PK will be explored using synthetic oligopeptides. The effects of phosphorylation of MBP and its fragments or peptides on their ability or inability to induce experimental allergic encephalomyelitis will be explored. In addition to MBP, a number of other endogenous substrates for PL-Ca-PK in brain will be further investigated. (3) Regulation of PL-Ca-PK system. Regulations by various agents of the enzyme activity, the phosphorylation state of endogenous substrates and possible "translocation" of PL-Ca-PK in brain will be investigated. (4) A new S-100-modulated protein phosphorylation system. The S-100-modulated enzyme (protein kinase X) and its l9K Mr substrate protein in rat brain will be further purified and characterized. It is hoped that the proposal will produce new knowledge regarding regulation of brain biology and pathology by phospholipid, Ca2+ and S-100 protein via protein phosphorylation reaction.