Ivanesthi Indira Rizqita, Rida Gita Riswana Nawung, Setiawibawa Aditya Aryandi, Tseng Yi-Kuan, Muammar Arief, Wang Chien-Chia
Department of Life Sciences, National Central University, Jungli District, Taoyuan, Taiwan.
Graduate Institute of Statistics, National Central University, Jungli District, Taoyuan, Taiwan.
Microbiol Spectr. 2023 Feb 22;11(2):e0462122. doi: 10.1128/spectrum.04621-22.
The 5' extra guanosine with 5'-monophosphate at position -1 (G-1) of tRNA (p-tRNA) is a nearly universal feature that establishes tRNA identity. G-1 is either genome encoded and retained after processing by RNase P (RNase P) or posttranscriptionally incorporated by tRNA guanylyltransferase (Thg1) after RNase P cleavage. However, RNase P is not found in the hyperthermophilic archaeum Nanoarchaeum equitans; instead, all of its tRNAs, including tRNA, are transcribed as leaderless tRNAs with 5'-triphosphate (ppp-tRNAs). How N. equitans histidyl-tRNA synthetase (NeHisRS) recognizes its cognate tRNA (NetRNA) is of particular interest. In this paper, we show that G-1 serves as the major identity element of NetRNA, with its anticodon performing a similar role, though to a lesser extent. Moreover, NeHisRS distinctly preferred p-tRNA over ppp-tRNA (~5-fold difference). Unlike other prokaryotic HisRSs, which strongly prefer tRNA with C73, this enzyme could charge tRNAs with A73 and C73 with nearly equal efficiency. As a result, mutation at the C73-recognition amino acid residue Q112 had only a minor effect (<2-fold reduction). This study suggests that NeHisRS has evolved to disregard C73, but it still maintains its evolutionarily preserved preference toward tRNA with 5'-monophosphate. Mature tRNA has, at its 5'-terminus, an extra guanosine with 5'-monophosphate, designated G-1. G-1 is the major recognition element for histidyl-tRNA synthetase (HisRS), regardless of whether it is of eukaryotic or prokaryotic origin. However, in the hyperthermophilic archaeum Nanoarchaeum equitans, all its tRNAs, including tRNA, are transcribed as leaderless tRNAs with 5'-triphosphate. This piqued our curiosity about whether N. equitans histidyl-tRNA synthetase (NeHisRS) prefers tRNA with 5'-triphosphate. We show herein that G-1 is still the major recognition element for NeHisRS. However, unlike other prokaryotic HisRSs, which strongly prefer tRNA with C73, this enzyme shows almost the same preference for C73 and A73. Most intriguingly, NeHisRS still prefers 5'-monophosphate over 5'-triphosphate. It thus appears that the preference of HisRS for tRNA with 5'-monophosphate emerged very early in evolution.
转运RNA（p-tRNA）在-1位带有5'-单磷酸的5'端额外鸟苷（G-1）是确定tRNA身份的一个几乎普遍存在的特征。G-1要么是基因组编码的，并在经过核糖核酸酶P（RNase P）加工后保留下来，要么在RNase P切割后由tRNA鸟苷酰转移酶（Thg1）进行转录后掺入。然而，在超嗜热古菌嗜纳米古菌中未发现RNase P；相反，其所有的tRNA，包括tRNA，转录时都是无前导序列的带有5'-三磷酸的tRNA（ppp-tRNAs）。嗜纳米古菌组氨酰-tRNA合成酶（NeHisRS）如何识别其同源tRNA（NetRNA）特别令人感兴趣。在本文中，我们表明G-1是NetRNA的主要身份识别元件，其反密码子也发挥类似作用，尽管程度较小。此外，NeHisRS明显更倾向于p-tRNA而非ppp-tRNA（相差约5倍）。与其他强烈偏好带有C73的tRNA的原核组氨酰-tRNA合成酶不同，这种酶对带有A73和C73的tRNA的充电效率几乎相同。因此，C73识别氨基酸残基Q112处的突变只有轻微影响（降低不到2倍）。这项研究表明NeHisRS在进化过程中已不再依赖C73，但它仍然保持着对带有5'-单磷酸的tRNA在进化上保留下来的偏好。成熟的tRNA在其5'端有一个带有5'-单磷酸的额外鸟苷，称为G-1。G-1是组氨酰-tRNA合成酶（HisRS）的主要识别元件，无论其起源是真核还是原核。然而，在超嗜热古菌嗜纳米古菌中，其所有的tRNA，包括tRNA，转录时都是无前导序列的带有5'-三磷酸的tRNA。这引发了我们对于嗜纳米古菌组氨酰-tRNA合成酶（NeHisRS）是否更偏好带有5'-三磷酸的tRNA的好奇。我们在此表明G-1仍然是NeHisRS的主要识别元件。然而，与其他强烈偏好带有C73的tRNA的原核HisRS不同，这种酶对C73和A73的偏好几乎相同。最有趣的是，NeHisRS仍然更偏好5'-单磷酸而非5'-三磷酸。因此，HisRS对带有5'-单磷酸的tRNA的偏好似乎在进化过程中很早就出现了。
