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

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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