Institute of Molecular and Cell Biology, University of Tartu, Riia 23, 51010 Tartu, Estonia.
Biochimie. 2012 May;94(5):1080-9. doi: 10.1016/j.biochi.2012.02.023.
Nucleoside modifications are introduced into the ribosomal RNA during the assembly of the ribosome. The number and the localization of the modified nucleosides in rRNAs are known for several organisms. In bacteria, rRNA modified nucleosides are synthesized by a set of specific enzymes, the majority of which have been identified in Escherichia coli. Each rRNA modification enzyme recognizes its substrate nucleoside(s) at a specific stage of ribosome assembly. Not much is known about the specificity determinants involved in the substrate recognition of the modification enzymes. In order to shed light on the substrate specificity of RluD and RlmH, the enzymes responsible for the introduction of modifications into the stem-loop 69 (H69), we monitored the formation of H69 pseudouridines (Ψ) and methylated pseudouridine (m3Ψ) in vitro on ribosomes with alterations in 23S rRNA. While the synthesis of Ψs in H69 by RluD is relatively insensitive to the point mutations at neighboring positions, methylation of one of the Ψs by RlmH exhibited a much stronger sensitivity. Apparently, in spite of synthesizing modifications in the same region or even at the same position of rRNA, the two enzymes employ different substrate recognition mechanisms.
核苷修饰是在核糖体组装过程中引入核糖体 RNA 中的。几种生物体中已知 rRNA 修饰核苷的数量和位置。在细菌中，rRNA 修饰核苷由一组特定的酶合成，其中大多数在大肠杆菌中已被鉴定。每个 rRNA 修饰酶都在核糖体组装的特定阶段识别其底物核苷。关于修饰酶底物识别中涉及的特异性决定因素，我们知之甚少。为了阐明负责引入茎环 69（H69）修饰的酶 RluD 和 RlmH 的底物特异性，我们在核糖体上监测了 23S rRNA 突变对 H69 假尿嘧啶核苷（Ψ）和甲基假尿嘧啶核苷（m3Ψ）形成的影响。虽然 RluD 合成 H69 中的 Ψ 相对不敏感于相邻位置的点突变，但 RlmH 对一个 Ψ 的甲基化表现出更强的敏感性。显然，尽管在 rRNA 的相同区域甚至相同位置合成修饰，但这两种酶采用不同的底物识别机制。
