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通过测量二肽和三肽形成来研究真核生物mRNA翻译的简化体外系统。

Simplified in vitro system for study of eukaryotic mRNA translation by measuring di- and tripeptide formation.

作者信息

Cenatiempo Y, Twardowski T, Redfield B, Reid B R, Dauerman H, Weissbach H, Brot N

出版信息

Proc Natl Acad Sci U S A. 1983 Jun;80(11):3223-6. doi: 10.1073/pnas.80.11.3223.

DOI:10.1073/pnas.80.11.3223
PMID:6574481
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC394012/
Abstract

An in vitro system for measurement of rabbit globin mRNA translation has been developed based on the formation of the NH2-terminal dipeptide, fMet-Val. The basic components include a partially purified initiation factor preparation from rabbit reticulocytes supplemented with eukaryotic initiation factor 4A, purified and formylated yeast Met-tRNAi, and rabbit liver or Escherichia coli Val-tRNA1Val. Picomole quantities of fMet-Val are synthesized, dependent on mRNA, and the dipeptide is readily assayed by a simple extraction procedure. In the presence of Leu-tRNA or His-tRNA, the tripeptides fMet-Val-Leu and fMet-Val-His are synthesized, corresponding to the NH2-terminal sequence of alpha- and beta-globin, respectively. Therefore, tripeptide synthesis provides a simple means to distinguish between the expression of the alpha- and beta-globin mRNA species.

摘要

基于氨基末端二肽fMet-Val的形成，已开发出一种用于测量兔珠蛋白mRNA翻译的体外系统。基本成分包括来自兔网织红细胞的部分纯化起始因子制剂，并补充真核起始因子4A、纯化的甲酰化酵母Met-tRNAi以及兔肝或大肠杆菌Val-tRNA1Val。皮摩尔量的fMet-Val依赖于mRNA进行合成，并且该二肽可通过简单的提取程序轻松检测。在亮氨酸tRNA或组氨酸tRNA存在的情况下，分别合成对应于α-和β-珠蛋白氨基末端序列的三肽fMet-Val-Leu和fMet-Val-His。因此，三肽合成提供了一种区分α-和β-珠蛋白mRNA种类表达的简单方法。

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本文引用的文献

1
Translational control of ribosomal protein L10 synthesis occurs prior to formation of first peptide bond.核糖体蛋白L10合成的翻译控制发生在第一个肽键形成之前。
Proc Natl Acad Sci U S A. 1981 Jul;78(7):4261-4. doi: 10.1073/pnas.78.7.4261.
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Use of different tRNASer isoacceptor species in vitro to discriminate between the expression of plasmid genes.在体外使用不同的丝氨酸tRNA同工受体种类来区分质粒基因的表达。
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In vitro expression of Escherichia coli ribosomal protein L 10 gene: tripeptide synthesis as a measure of functional mRNA.大肠杆菌核糖体蛋白L10基因的体外表达：以三肽合成作为功能性信使核糖核酸的衡量指标。
Arch Biochem Biophys. 1982 Oct 15;218(2):572-8. doi: 10.1016/0003-9861(82)90381-2.
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In vitro synthesis of the first dipeptide of the beta subunit of Escherichia coli RNA polymerase.大肠杆菌RNA聚合酶β亚基首个二肽的体外合成
Proc Natl Acad Sci U S A. 1982 Aug;79(15):4609-12. doi: 10.1073/pnas.79.15.4609.
5
Methionyl soluble ribonucleic acid transformylase. I. Purification and partial characterization.甲硫氨酰可溶性核糖核酸转甲酰酶。I. 纯化及部分特性鉴定
J Biol Chem. 1967 Apr 10;242(7):1522-5.
6
Initial dipeptide formation in hemoglobin biosynthesis.血红蛋白生物合成中的初始二肽形成。
Proc Natl Acad Sci U S A. 1971 Aug;68(8):1810-4. doi: 10.1073/pnas.68.8.1810.
7
Alpha and beta globin messenger ribonucleic acid. Different amounts and rates of initiation of translation.α和β珠蛋白信使核糖核酸。不同的翻译起始量和速率。
J Biol Chem. 1971 Dec 10;246(23):7131-8.
8
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A supernatant factor involved in initiation complex formation with eukaryotic ribosomes.一种参与与真核核糖体形成起始复合物的上清因子。
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