新型SIRT5抑制剂及抑制策略的探索研究

发布时间：2018-03-26 01:21

本文选题：sirtuin　切入点：SIRT5　出处：《江苏大学》2017年硕士论文

【摘要】：蛋白翻译后修饰(PTMs)越来越多的表明在调节蛋白功能以及在许多生理和病理过程中起了很大作用。赖氨酸乙酰化是被研究最多的修饰和调节之一,参与众多至关重要的细胞内过程包括蛋白降解,转录激活以及新陈代谢。沉默信息调节因子2(Sir2)或者说sirtuin是一类高度保守的依赖β-烟酰胺腺嘌呤二核苷酸(β-NAD+)的酶,它调节了许多代谢通路。自从2000年以来,哺乳动物中已经发现了七个sirtuin家族成员,即SIRT1-7。Sirtuin被视为是当代癌症、代谢类疾病以及神经退行性疾病的治疗靶点。因此,其抑制剂在过去这些年成为了研究重点。其中四个成员(SIRT4-7)没有检测到或者只检测到很微弱的去乙酰活性,倾向于去除长链脂肪酰基、丁二酰基等。在这份论文工作中,我们选用存在于线粒体和细胞质中的SIRT5作为研究目标。SIRT5是体内高效的赖氨酸去丙二酰、去丁二酰、去戊二酰的去酰化酶。对于这种去除负电羧基的偏好可以解释为由于105位的精氨酸残基以及102位的酪氨酸残基组成的人类SIRT5去酰化口袋。针对目前SIRT5抑制剂研究不够充分的情况,更多高效且有选择性的抑制剂需要被发现。首先,我们选用高效的催化机制导向的抑制弹头---Nε-羧乙基-硫代氨甲酰-赖氨酸SIRT5抑制弹头,将其运用到我们设计的环肽骨架中去。设计合成了一系列带有该弹头的环肽化合物,用于抑制活性测试和构效关系研究(SAR)。根据这些化合物SIRT5活性抑制测试的结果,这个环肽的抑制策略被证明是一个有效增强sirtuin抑制剂活性的方法。化合物3发现是一个强效的SIRT5抑制剂(IC50=2.2±0.89μM),是相应的直链肽化合物7的SIRT5抑制强度的42.3倍左右。此外,在蛋白酶水解测试中发现,化合物3相较于直链肽化合物7其蛋白酶水解的耐受性很强。这也证明了环肽骨架的优越性。再有,我们不满足于现有的SIRT5抑制弹头,尝试性地设计了新型弹头。将之前弹头末端丙酸替换成了苯甲酸,向其中引入芳香环,期待能更紧密占据SIRT5酰基化口袋。我们也将弹头中硫脲结构替换成硒脲,化合物9(IC50=1.7±0.1μM)相较于之前使用相同肽骨架的硫脲化合物(IC50=5±1.9μM)的SIRT5抑制活性提高了2.9倍左右。预期这一变化会成为整个sirtuin抑制剂研究的有力武器。
[Abstract]:Protein post-translational modification (PTMs) has been increasingly shown to play a significant role in regulating protein function and in many physiological and pathological processes. Lysine acetylation is one of the most widely studied modifications and modulations. Involved in a number of critical intracellular processes including protein degradation, transcriptional activation, and metabolism. Silencing signaling factor 2 / Sir2) or sirtuin is a highly conserved enzyme that relies on 尾 -nicotinamide adenine dinucleotides (尾 -NAD). It regulates many metabolic pathways. Since 2000, seven members of the sirtuin family, known as SIRT1-7.Sirtuin, have been identified as targets for modern cancer, metabolic diseases and neurodegenerative diseases. Its inhibitors have been the focus of research in the past few years. Four of its members, SIRT4-7, have not detected or only detected very weak deacetylation activities, and tend to remove long-chain fatty acyl, succinyl, and so on. We selected SIRT5 in mitochondria and cytoplasm as our research objective. SIRT5 is highly efficient lysine demalonyl and succinyl in vivo. De-glutaryl deacylase. The preference for the removal of negatively charged carboxyl groups can be explained by a human SIRT5 deacylation pocket consisting of 105th arginine residues and 102nd tyrosine residues. Insufficient circumstances, More efficient and selective inhibitors need to be found. First, we use a highly efficient catalytic mechanism directed inhibitory warhead-N 蔚 -carboxyethyl-thiocarbamate-lysine SIRT5 inhibitory warhead. A series of cyclic peptide compounds with this warhead were designed and synthesized for inhibition activity testing and structure-activity relationship study. Based on the results of SIRT5 activity inhibition tests of these compounds, The inhibition strategy of this cyclic peptide has been proved to be an effective method to enhance the activity of sirtuin inhibitor. Compound 3 was found to be a potent SIRT5 inhibitor, IC502.2 卤0.89 渭 M-1, which is about 42.3 times of the SIRT5 inhibitory intensity of the corresponding straight chain peptide compound 7. In the proteolytic test, compound 3 was found to be more tolerant of proteolysis than that of straight chain peptide compound 7. This also proves the superiority of cyclic peptide skeleton. Moreover, we are not satisfied with the existing SIRT5 inhibitory warhead. A new warhead was tentatively designed. Propionic acid was replaced with benzoic acid at the end of the warhead, and aromatic rings were introduced into it, hoping to occupy more closely the SIRT5 acylation pocket. We also replaced the thiourea structure in the warhead with selenourea. The SIRT5 inhibitory activity of compound 9(IC50=1.7 卤0.1 渭 M was about 2.9-fold higher than that of thiourea compound IC5050 卤1.9 渭 M using the same peptide skeleton. It is expected that this change will become a powerful weapon in the study of sirtuin inhibitors.
【学位授予单位】：江苏大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R91

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