Department of Microbiology and Cell Science, University of Florida, Gainesville, FL, USA.
Biol Direct. 2010 Jan 13;5:3. doi: 10.1186/1745-6150-5-3.
The bacterial elongation factor P (EF-P) is strictly conserved in bacteria and essential for protein synthesis. It is homologous to the eukaryotic translation initiation factor 5A (eIF5A). A highly conserved eIF5A lysine is modified into an unusual amino acid derived from spermidine, hypusine. Hypusine is absolutely required for eIF5A's role in translation in Saccharomyces cerevisiae. The homologous lysine of EF-P is also modified to a spermidine derivative in Escherichia coli. However, the biosynthesis pathway of this modification in the bacterial EF-P is yet to be elucidated.
Here we propose a potential mechanism for the post-translational modification of EF-P. By using comparative genomic methods based on physical clustering and phylogenetic pattern analysis, we identified two protein families of unknown function, encoded by yjeA and yjeK genes in E. coli, as candidates for this missing pathway. Based on the analysis of the structural and biochemical properties of both protein families, we propose two potential mechanisms for the modification of EF-P.
This hypothesis could be tested genetically by constructing a bacterial strain with a tagged efp gene. The tag would allow the purification of EF-P by affinity chromatography and the analysis of the purified protein by mass spectrometry. yjeA or yjeK could then be deleted in the efp tagged strain and the EF-P protein purified from each mutant analyzed by mass spectrometry for the presence or the absence of the modification. This hypothesis can also be tested by purifying the different components (YjeK, YjeA and EF-P) and reconstituting the pathway in vitro.
The requirement for a fully modified EF-P for protein synthesis in certain bacteria implies the presence of specific post-translational modification mechanism in these organisms. All of the 725 bacterial genomes analyzed, possess an efp gene but only 200 (28%) possess both yjeA and yjeK genes. In the other organisms, EF-P may be modified by another pathway or the translation machinery must have adapted to the lack of EF-P modification. Our hypotheses, if confirmed, will lead to the discovery of a new post-translational modification pathway.
This article was reviewed by Céline Brochier-Armanet, Igor B. Zhulin and Mikhail Gelfand. For the full reviews, please go to the Reviewers' reports section.
细菌延伸因子 P(EF-P)在细菌中严格保守,对蛋白质合成至关重要。它与真核翻译起始因子 5A(eIF5A)同源。eIF5A 的一个高度保守赖氨酸被修饰为来自亚精胺的不寻常氨基酸,即 hypusine。在酿酒酵母中,hypusine 绝对是 eIF5A 翻译功能所必需的。大肠杆菌中的 EF-P 的同源赖氨酸也被修饰为亚精胺衍生物。然而,细菌 EF-P 中这种修饰的生物合成途径尚未阐明。
在这里,我们提出了 EF-P 翻译后修饰的潜在机制。通过使用基于物理聚类和系统发育模式分析的比较基因组方法,我们确定了大肠杆菌 yjeA 和 yjeK 基因编码的两个未知功能的蛋白质家族是该缺失途径的候选者。基于对这两个蛋白质家族的结构和生化特性的分析,我们提出了 EF-P 修饰的两种潜在机制。
这个假说可以通过构建带有标记 efp 基因的细菌菌株来进行遗传检验。该标签将允许通过亲和层析纯化 EF-P,并通过质谱分析对纯化的蛋白质进行分析。然后可以在 efp 标记的菌株中删除 yjeA 或 yjeK,并通过质谱分析从每个突变体中纯化的 EF-P 蛋白是否存在修饰。该假说也可以通过纯化不同的成分(YjeK、YjeA 和 EF-P)并在体外重新构建途径来进行测试。
某些细菌中蛋白质合成需要完全修饰的 EF-P 意味着这些生物体中存在特定的翻译后修饰机制。在分析的 725 个细菌基因组中,都有 efp 基因,但只有 200 个(28%)同时具有 yjeA 和 yjeK 基因。在其他生物体中,EF-P 可能通过另一种途径进行修饰,或者翻译机制必须适应缺乏 EF-P 修饰的情况。如果我们的假说得到证实,将发现一种新的翻译后修饰途径。
这篇文章由 Céline Brochier-Armanet、Igor B. Zhulin 和 Mikhail Gelfand 进行了评审。有关完整的评审,请前往评审报告部分。