增加的eIF2A水平通过隔离40S核糖体子单元来抑制翻译
Daisy J Grove1,2, Daniel J Levine2, Michael G Kearse1,2
1The Ohio State Biochemistry Program, The Ohio State University, Columbus, OH 43210, USA.
Nucleic acids research
|August 21, 2023
概括
超量的真核转化因子eIF2A通过隔离40S核糖体子单元来抑制蛋白质合成. 这一发现澄清了eIF2A在翻译启动中的神秘作用.
科学领域:
- 分子生物学分子生物学
- 细胞生物学 细胞生物学
- 生物化学 生化学
背景情况:
- 欧核生物启动因子2亚单元α (eIF2A) 的确切功能仍然不清楚,这是第一个确定的欧核生物启动因子tRNA载体.
- eIF2A在人类癌细胞中表现出不寻常的非细胞质局部化,与典型的翻译启动因子不同.
- 之前的研究面临的挑战是生产足够的可溶性重组eIF2A用于机械研究.
研究的目的:
- 为复合人类eIF2A开发一种有效的净化方法.
- 在翻译中阐明eIF2A的抑制机制.
- 为了研究eIF2A与核糖体子单元的相互作用.
主要方法:
- 开发了从大肠杆菌和昆虫细胞中复合人类eIF2A的新型净化策略,实现了显著更高的产量.
- 使用哺乳动物体外翻译系统来评估eIF2A对记者mRNA翻译的影响.
- 进行了拉下测试,以确定eIF2A和40S核糖体子单元之间的结合相互作用.
主要成果:
- 实现了重组人类eIF2A产量的大幅增加 (大肠杆菌的360倍,昆虫细胞的6000倍).
- 证明高的eIF2A水平在开始编码子识别之前抑制了各种mRNA的翻译,不论是开始编码子类型.
- 表明eIF2A抑制了病毒内部核糖体进入部位 (IRESs) 介导的翻译,包括那些独立于启动因子的部位.
- 证实过多的eIF2A会隔离40S核糖体子单元,这种效应通过补充额外的40S子单元来逆转.
- 通过拉下测试,确定了重组eIF2A和40S子单元之间的直接结合.
结论:
- 过多的eIF2A通过隔离40S核糖体子单元来抑制翻译.
- eIF2A与40S子单元的相互作用是它对蛋白质合成的抑制作用的关键机制.
- 保持适当的eIF2A水平至关重要,以防止其干扰翻译机器.
相关概念视频
02:07Improving Translational Accuracy
11.6K
Base complementarity between the three base pairs of mRNA codon and the tRNA anticodon is not a failsafe mechanism. Inaccuracies can range from a single mismatch to no correct base pairing at all. The free energy difference between the correct and nearly correct base pairs can be as small as 3 kcal/ mol. With complementarity being the only proofreading step, the estimated error frequency would be one wrong amino acid in every 100 amino acids incorporated. However, error frequencies observed in...
11.6K
02:33Initiation of Translation
34.1K
Initiating translation is complex because it involves multiple molecules. Initiator tRNA, ribosomal subunits, and eukaryotic initiation factors (eIFs) are all required to assemble on the initiation codon of mRNA. This process consists of several steps that are mediated by different eIFs.
First, the initiator tRNA must be selected from the pool of elongator tRNAs by eukaryotic initiation factor 2 (eIF2). The initiator tRNA (Met-tRNAi) has conserved sequence elements including modified bases at...
First, the initiator tRNA must be selected from the pool of elongator tRNAs by eukaryotic initiation factor 2 (eIF2). The initiator tRNA (Met-tRNAi) has conserved sequence elements including modified bases at...
34.1K
01:29Translational Regulation
44
Translational regulation in prokaryotes ensures efficient protein synthesis by controlling ribosome access to mRNA. This regulation is mediated by secondary RNA structures, including translational riboswitches, RNA thermometers, and small RNAs (sRNAs), which respond to intracellular and environmental signals to modulate gene expression.Translational RiboswitchesRiboswitches in the leader region of mRNAs can regulate translation by altering the accessibility of the Shine-Dalgarno (SD) sequence,...
44
01:23Stringent Response in E. coli
31
Bacterial growth is closely tied to nutrient availability, with cells proliferating exponentially under favorable conditions and entering a stationary phase when resources become scarce. This transition is mediated by a regulatory mechanism known as the stringent response, which allows bacteria to adapt to nutrient deprivation by modulating gene expression and metabolic activity.During nutrient scarcity, intracellular amino acid levels decline. It results in the accumulation of uncharged tRNAs...
31
01:44Termination of Translation
25.5K
The large ribosomal subunit has several important structures essential to translation. These include the peptidyl transferase center (PTC) - which is the site where the peptide bond is formed - and a large, internal, water-filled tube through which the nascent polypeptide moves. This latter structure is called the Peptide Exit Tunnel, and it begins at the PTC and spans the body of the large ribosomal subunit. During translation, as the nascent polypeptide chain is synthesized, it passes through...
25.5K
02:28Leaky Scanning
5.2K
During most eukaryotic translation processes, the small 40S ribosome subunit scans an mRNA from its 5' end until it encounters the first start AUG codon. The large 60S ribosomal subunit then joins the smaller one to initiate protein synthesis. The location of the translation initiation is largely determined by the nucleotides near the start codon as there may be multiple translation initiation sites present on the mRNA. Marilyn Kozak discovered that the sequence RCCAUGG (where R...
5.2K