MUTAGENESIS STUDIES OF MAP REGULATORY PROTEIN DOMAINS

ARF、ras(P21)、src、cAMP依赖的蛋白激酶和 G蛋白家族，都是调节蛋白，其细胞内 定位和/或活性受共价连接的影响 在脂肪酸中，肉豆蔻酸酯或棕榈酸酯。岗位的作用-- 调节蛋白的翻译共价酰化将是 通过使用Site-Site生成一系列突变基因进行检查 定向诱变。然后，正常和突变的cDNA将被 用于内源性酶检测的原核表达和纯化蛋白 活动以及参与监管、 蛋白质之间的相互作用。将使用免疫细胞化学 确定突变蛋白的细胞定位 在适当的细胞系统中表达。另外，变种人 蛋白质将作为潜在的底物或抑制物进行测试 肉豆蔻醇和棕榈基转移酶。

ARF, ras (p21), src, cAMP-dependent protein kinase, and the family of G-proteins, are all regulatory proteins whose cellular localization and/or activities are affected by covalent attachment of the fatty acids, myristate or palmitate. The role of post- translational covalent acylation of regulatory proteins will be examined by generating a series of mutant genes using site- directed mutagenesis. The normal and mutant cDNAs will then be expressed and proteins purified for tests of intrinsic enzymatic activities as well as the ability to participate in regulatory, protein-protein interactions. Immunocytochemistry will be used to determine the cellular localization of mutant proteins expressed in appropriate cellular systems. In addition, the mutant proteins will be tested as potential substrates or inhibitors of myristyl and palmityl transferases.