Kaberdin Vladimir R
Max F. Perutz Laboratories, Department of Microbiology and Genetics, University Department at the Vienna Biocenter, Dr. Bohrgasse 9/4, A-1030 Vienna, Austria.
Nucleic Acids Res. 2003 Aug 15;31(16):4710-6. doi: 10.1093/nar/gkg690.
Endoribonuclease RNase E has a central role in both processing and decay of RNA in Escherichia coli, and apparently in many other organisms, where RNase E homologs were identified or their existence has been predicted from genomic data. Although the biochemical properties of this enzyme have been already studied for many years, the substrate specificity of RNase E is still poorly characterized. Here, I have described a novel oligonucleotide-based assay to identify specific sequence determinants that either facilitate or impede the recognition and cleavage of RNA by the catalytic domain of the enzyme. The knowledge of these determinants is crucial for understanding the nature of RNA-protein interactions that control the specificity and efficiency of RNase E cleavage and opens new perspectives for further studies of this multi-domain protein. Moreover, the simplicity and efficiency of the proposed assay suggest that it can be a valuable tool not only for the characterization of RNase E homologs but also for the analysis of other site-specific nucleases.
核糖核酸酶E（Endoribonuclease RNase E）在大肠杆菌的RNA加工和降解过程中发挥着核心作用，显然在许多其他生物中也是如此，在这些生物中已鉴定出RNase E的同源物，或者根据基因组数据预测了它们的存在。尽管对这种酶的生化特性已经研究了多年，但RNase E的底物特异性仍未得到很好的表征。在这里，我描述了一种基于寡核苷酸的新检测方法，以鉴定促进或阻碍该酶催化结构域对RNA识别和切割的特定序列决定因素。了解这些决定因素对于理解控制RNase E切割特异性和效率的RNA-蛋白质相互作用的本质至关重要，并为进一步研究这种多结构域蛋白开辟了新的视角。此外，所提出的检测方法的简单性和高效性表明，它不仅可以作为表征RNase E同源物的有价值工具，还可以用于分析其他位点特异性核酸酶。
