Studies on the cellular functions of lipid-modified proteins

脂质修饰蛋白的细胞功能研究

• 批准号: 07557162
• 负责人: OHMURA Satoshi
• 金额: $ 5.12万
• 依托单位: The Kitasato Institute
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (A)
• 财政年份: 1995
• 资助国家: 日本
• 起止时间: 1995 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07557162/
• 关键词: Myristoylation; Farnesyltransferase; HIV Gag protein; Triacsin; Acly-Coa synthetase; Inhibitor; Ras protein; Andrastin; 抗エイズ薬; 抗癌剤; 脂質修飾蛋白質; アシル化; 抗エイズ剤

Many cellular proteins are modified with lipids such as acyl(palmitoyl or myristoly)and/or prenly(farnesly or geranylgeranyl)residues. In this researach project, the biochemical functions of lipid-modified proteins were studied and searaach for microbial inhibitors of their formation wes carried out as a new approach for anticancer or antiviral agents.HIV Gag protein is myrisstoylataed at the N-terminal glycine residue, which plays an important role in virus particlebdding. The myristoylation levels were controlled to investigate the effect on the particle budding by using the specifiv acyl-CoA synthetase inhibitor triacsin previousy discovered by our group. We showed that the inhibition of Gag myristoylation by triacsin follows dose-dependent kinetics but that the particle budding exhibits sudden shutoff kinetics, suggesting that only a relatively small proportion of total Gag molecules need to be myristoylated for efficient budding and indicating that total inhibition of myristoylation will be required for effective anti-HIV therapy.Oncogene product Ras proteins are posttranslationally farnesylated at the cysteine residue near the C-terminus, in which protein farnesyltransferase(PFTase)is involved. PFTase is expected as a novel target of inhibition since the inhibition causes altering membrane localization and blocking activation of Ras proteins. An efficient screening system was conducted by utilizing Saccharomyces cerevisiae, resulting in discovery of two series of new PTFase inhibitors, andrastins produced by Penicillium sp.FO-3929 and kurasoins by Paecilomyces sp.FO-3684. The structure elucidation including stereochemistries, biosynthesis of andrastin and total sylnthesis of kurasoin were studied. Their inhibitory activity against PFTase (IC_<50>) was 10-60muM.It still remains to be investigated whether or not these compounds inhibit PFTase in cells and show in fvivo anticancer activity.

许多细胞蛋白质用脂质如酰基（棕榈酰基或肉豆蔻酰基）和/或prenly（法尼基或香叶基香叶基）残基修饰。HIV Gag蛋白的N端甘氨酸残基被肉豆蔻酰化，在病毒颗粒形成过程中起重要作用。通过使用本小组先前发现的特异性酰基辅酶A合成酶抑制剂三酸甘油酯来控制肉豆蔻酰化水平以研究对颗粒出芽的影响。我们表明，三酸甘油酯对Gag豆蔻酰化的抑制遵循剂量依赖性动力学，但颗粒出芽表现出突然关闭动力学，这表明只有相对小比例的总Gag分子需要被豆蔻酰化以有效出芽，并表明有效的抗增殖需要完全抑制豆蔻酰化。HIV治疗：癌基因产物Ras蛋白在C-末端附近的半胱氨酸残基处进行法尼基化，其中涉及蛋白法尼基转移酶（PFT酶）。PFT酶被认为是一种新的抑制靶点，因为它的抑制作用改变了Ras蛋白的膜定位并阻断了Ras蛋白的激活。利用酿酒酵母（Saccharomycescerevisiae）进行了一个高效的筛选系统，发现了两个新的PTFase抑制剂系列，即青霉（Penicilliumsp.FO-3929）产生的andrastins和拟青霉（Paecilomycessp.FO-3684）产生的kurasoins。本论文对苦皮素的立体化学、生物合成和全合成进行了研究。这些化合物对PFT酶的抑制活性（IC_2<50>）为10- 60 μ M，它们是否能抑制细胞内PFT酶并显示出体内抗癌活性还有待进一步研究。

项目成果

Sunazuka T.: "Synthesis and absolute structures of novel protein farnesyltransferase inhibitors,kurasoins A and B." J.Antibiot.50. 453-455 (1997)

Sunazuka T.：“新型蛋白质法呢基转移酶抑制剂 kurasoins A 和 B 的合成和绝对结构。”

Jones I.: "The molecular basis of HIV capsid assembly." Rev.Med.Viol. (1998)

Jones I.：“HIV 衣壳组装的分子基础。”

Matsuzawa A.et al.: "Prorection against stress-induced cell death by intracellular PAF acetyihydrolaseII" J.Biol.Chem.272. 32315-32320 (1997)

Matsuzawa A.et al.：“通过细胞内 PAF 乙酰水解酶 II 预防应激诱导的细胞死亡”J.Biol.Chem.272。

Uchida R: "Kurasoins A and B,new protein farnesytransferase inhibitorsproduced by Penicillium sp.FO-3684.I.Producing strain,fermentation,isolation and biological activities." J.Antibiot.49. 932-934 (1996)

Uchida R：“Kurasoins A和B，青霉属sp.FO-3684产生的新型蛋白质法尼基转移酶抑制剂。I.生产菌株、发酵、分离和生物活性。”

Jones I.M.& Morikawa Y.: "The molecular bisis of HIV capsid assembly" Rev.Med.Virol.(印刷中). (1998)

Jones I.M. 和 Morikawa Y.：“HIV 衣壳组装的分子二元”Rev.Med.Virol.（出版中）。