Recognition of tRNAIle with a UAU anticodon by isoleucyl‐tRNA synthetase in lactic acid bacteria
Recognition of tRNAIle with a UAU anticodon by isoleucyl‐tRNA synthetase in lactic acid bacteria
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乳酸菌中异亮氨酰-tRNA合成酶对带有UAU反密码子的tRNAIle的识别
DOI：
10.1111/febs.16389
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发表时间：
2022
期刊：
影响因子：
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通讯作者：
Takai Kazuyuki
中科院分区：
文献类型：
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作者：
Uesugi Gakuto;Fukuba Yuho;Yamamoto Takayuki;Inaba Nozomi;Furukawa Haruyuki;Yoshizawa Satoko;Tomikawa Chie;Takai Kazuyuki
在几乎所有的真细菌中，AUA密码子由在摆动位置上携带tRNAIle 2的赖氨酸翻译。赖氨酸是由tRNAIle赖氨酸合成酶（TilS）通过转录后修饰tRNAIle 2（CAU）的胞苷而引入的。干酪乳杆菌和植物乳杆菌具有TilShomologues和tRNAIle 2（CAU）基因。此外，干酪乳杆菌还具有另一个tRNAIle 2基因，该基因具有一个UAU反密码子。在这里，我们证明了L.caseitRNAIle2（UAU）通过L. caseiisoleucyl-tRNA合成酶（IleRS）而不是通过L. plantarumIleRS装载异亮氨酸，尽管这两种酶的氨基酸同一性超过60%。据报道，在移动的支原体中，它有它的tRNAIle 2（UAU），但没有IleRS的865位的Arg残基是识别UAU反密码子所必需的。这一位置也被两种乳酸杆菌IleRS中的Arg占据。因此，干酪乳杆菌IleRS中的其他残基也有助于识别tRNAIle 2（UAU）。我们发现，其中N-末端结构域被植物乳杆菌IleRS的相应区域取代的嵌合体干酪乳杆菌IleRS对tRNAIle 2（UAU）和tRNAIle 1（GAU）具有非常低的氨酰化活性。A18 G突变体对任一tRNAIle几乎没有可检测的氨酰化活性。然而，A18 G和G19 N的双点突变体使tRNAIle 1（GAU）氨酰化，而不使tRNAIle 2（UAU）氨酰化。我们的研究结果表明，对于L. caseiIleRS，Ala 18和Gly 19在识别tRNAIle 2（UAU）中也起着关键作用。
In almost all eubacteria, the AUA codon is translated by tRNAIle2bearing lysidine at the wobble position. Lysidine is introduced by tRNAIlelysidine synthetase (TilS) via post‐transcriptional modification of the cytidine of tRNAIle2(CAU).Lactobacillus caseiandLactobacillus plantarumhavetilShomologues and tRNAIle2(CAU) genes. In addition,L. caseialso has another tRNAIle2gene with an UAU anticodon.L. plantarumhas a tRNAIle(UAU)‐like RNA. Here, we demonstrate thatL. caseitRNAIle2(UAU) is charged with isoleucine byL. caseiisoleucyl‐tRNA synthetase (IleRS) but not byL. plantarumIleRS, even though the amino acid identity of these two enzymes is over 60%. It has been reported that, inMycoplasma mobile, which has its tRNAIle2(UAU) but notilShomologue, an Arg residue at position 865 of the IleRS is required for recognition of the UAU anticodon. This position is occupied by an Arg also in the IleRSs from both of theLactobacillusspecies. Thus, other residues inL. casei, IleRS should also contribute to the recognition of tRNAIle2(UAU). We found that a chimericL. caseiIleRS in which the N‐terminal domain was replaced by the corresponding region ofL. plantatarumIleRS has very low aminoacylation activity towards both tRNAIle2(UAU) and tRNAIle1(GAU). The A18G mutant had barely detectable aminoacylation activity towards either of the tRNAsIle. However, a double point mutant of A18G and G19N aminoacylated tRNAIle1(GAU), but not tRNAIle2(UAU). Our results suggest that, forL. caseiIleRS, Ala18 and Gly19 also play a critical role in recognition of tRNAIle2(UAU).