STRUCTURE AND MECHANISM OF PROTEIN PRENYLTRANSFERASES

这项建议涉及哺乳动物蛋白质戊烯基转移酶蛋白质：法尼基转移酶(FTase)和香叶基香叶基转移酶(GGTase-1)的结构和机制。这些蛋白质催化将类异戊二烯类脂加成到一些小的调节G蛋白上。这种修饰(戊烯基化)对于G蛋白的活性和靶向细胞膜是必不可少的。FTase介导的RAS癌基因蛋白的法尼化是其致癌和细胞转化活性所必需的。用FTase抑制剂处理RAS转化的细胞，在组织培养细胞和动物模型中可以逆转转化的表型，而没有明显的毒性。抑制RAS法尼化目前被认为是最有希望的抗癌靶点之一。超过25%的人类癌症与致癌形式的RAS有关。因此，阐明FTase及其底物复合体的三维结构对于理解信号转导的基本过程以及开发具有药理活性的抗癌药物或现有药物的改进衍生物都具有重要意义。GGTase-1作为药物开发的潜在靶点，最近也受到了关注。细胞中的大多数前烯基化蛋白质都被香叶基香叶基异戊二烯类化合物修饰。GGTase-1催化参与细胞增殖的几个RAS超家族成员的前烯基化。抑制GGTase-1可能不仅在癌症化疗中很重要，而且在心血管疾病的治疗和抗真菌化合物的开发中也是重要的。此外，GGTase-1的结构，特别是与相关FTase的结构相比较，可能会加深我们对多肽和类异戊二烯底物专一性的机制的理解。此外，了解FTase和GGTase底物特异性差异的分子基础可能有助于开发更具选择性的FTase(和GGTase-1)抑制剂用于抗癌治疗。这项建议的总体目标是利用X射线结晶学研究哺乳动物蛋白质戊烯基转移酶蛋白质的机制和底物特异性的分子基础：1)确定哺乳动物FTase底物、产物和抑制物复合体的高分辨率三维结构；2)确定哺乳动物GGTase-1的三维结构；3)确定GGTase-1及其底物、产物和抑制剂的结构。

This proposal is concerned with the structure and mechanism of mammalian protein prenyltransferase proteins: farnesyltransferase (FTase) and geranylgeranyltransferase (GGTase-1). These proteins catalyze the addition of an isoprenoid lipid to a number of small regulatory G-proteins. This modification (prenylation) is essential for the activity and targeting of G-proteins to cell membranes. The FTase-mediated farnesylation of Ras oncogene proteins is absolutely required for their oncogenic, cell transforming activity. Treatment of Ras transformed cells with inhibitors of FTase has been shown to result in reversion of the transformed phenotype in tissue culture cells and animal models without apparent toxicity. Inhibition of Ras farnesylation is currently considered one of the most promising anti-cancer targets. Over 25 percent of human carcinomas are associated with oncogenic forms of Ras. Elucidation of the three-dimensional structure of FTase and its substrate complexes is therefore important both for understanding fundamental processes in signal transduction and for the development of pharmocologically active anti-cancer drugs or improved derivatives of existing drugs. GGTase-1 has also received attention recently as a potential target of the development of pharmaceuticals. The majority of prenylated proteins in the cell are modified by the geranylgeranyl isoprenoid. GGTase-1 catalyzes the prenylation of several members of the Ras superfamily involved in cell proliferation. Inhibition of GGTase-1 may be important not only for cancer chemotherapy, but also in the treatment of cardiovascular disease and the development of anti-fungal compounds. Furthermore, the structure of GGTase-1, particularly when compared to the structure of the related FTase, may further our understanding of the mechanisms of both peptide and isoprenoid substrate specificity. In addition, understanding the molecular basis for the differences between FTase and GGTase substrate specificities may facilitate the development of more selective FTase (and GGTase-1) inhibitors for anti-cancer therapy. The overall goal of this proposal is to study the molecular basis for the mechanism and substrate specificity of mammalian protein prenyltransferase proteins using X-ray crystallography to 1) determine the high resolution, three-dimensional structures of mammalian FTase substrate, product, and inhibitor complexes 2) determine the three-dimensional structure of mammalian GGTase-1 3) determine the structures of GGTase-1 together with substrates, products, and inhibitors.