STRUCTURE AND MECHANISM OF PROTEIN PRENYLTRANSFERASES

本论文主要研究哺乳动物异戊二烯基转移酶的结构和作用机制：法尼基转移酶（FTase）和香叶基香叶基转移酶（GGTase-1）。 这些蛋白质催化类异戊二烯脂质与许多小的调节性G蛋白的加成。 这种修饰（异戊二烯化）对于G蛋白的活性和靶向细胞膜是必不可少的。 Ras癌基因蛋白的FT酶介导的法尼基化是其致癌细胞转化活性所绝对需要的。 用FTase抑制剂处理Ras转化细胞已显示导致组织培养细胞和动物模型中转化表型的逆转，而没有明显的毒性。 Ras法尼基化的抑制目前被认为是最有前途的抗癌靶点之一。 超过25%的人类癌症与Ras的致癌形式有关。 因此，阐明FTase及其底物复合物的三维结构对于理解信号转导中的基本过程和开发药理活性抗癌药物或现有药物的改进衍生物都是重要的。GGTase-1作为药物开发的潜在靶标最近也受到关注。 细胞中的大多数异戊二烯化蛋白质被香叶基香叶基类异戊二烯修饰。 GGT酶-1催化参与细胞增殖的Ras超家族的几个成员的异戊烯化。 抑制GGT酶-1可能不仅对癌症化疗很重要，而且对心血管疾病的治疗和抗真菌化合物的开发也很重要。 此外，GGT酶-1的结构，特别是当与相关的FTase的结构相比时，可以进一步理解肽和类异戊二烯底物特异性的机制。 此外，了解FTase和GGT酶底物特异性之间差异的分子基础可能有助于开发更具选择性的FTase（和GGT酶-1）抑制剂用于抗癌治疗。 本提案的总体目标是使用X射线晶体学研究哺乳动物蛋白异戊二烯基转移酶蛋白的机制和底物特异性的分子基础，以1）确定哺乳动物FT酶底物、产物和抑制剂复合物的高分辨率三维结构2）确定哺乳动物GGT酶-1的三维结构3）确定GGT酶-1与底物的结构，产品和抑制剂。

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.