Identity elements for specific aminoacylation of a tRNA by mammalian lysyl-tRNA synthetase bearing a nonspecific tRNA-interacting factor
Identity elements for specific aminoacylation of a tRNA by mammalian lysyl-tRNA synthetase bearing a nonspecific tRNA-interacting factor
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DOI：
10.1021/bi0606905
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发表时间：
2006-08-22
期刊：
影响因子：
2.9
通讯作者：
Mirande, Marc
中科院分区：
文献类型：
--
作者：
Francin, Mathilde;Mirande, Marc
哺乳动物赖氨酰-tRNA合成酶（LysRS）具有附加至原核样合成酶结构域的N-末端多肽链延伸。这种延伸，称为tRNA相互作用因子（tIF），具有RNA结合基序[KxxxK（K/R）xxK]，其非特异性结合tRNA的受体T Psi C茎环结构域，并提供与该酶有效的tRNA结合能力。因此，天然LysRS使RNA微螺旋氨基酰化，模拟tRNA（3）（Lys）的氨基酸受体茎环结构域。在这里，微螺旋识别的检查表明，哺乳动物LysRS aminoacylates RNA微螺旋没有特异性的序列，揭示了没有一个核苷酸从受体T Psi C茎环结构域的tRNA（赖氨酸）受体身份的重要决定因素。为了测试tIF结构域是否降低合成酶对完整tRNA分子的特异性，检查了野生型和突变型非同源tRNA通过野生型或N-末端截短的LysRS的氨酰化。tRNA（Lys）的UUU反密码子的存在似乎是将酵母tRNA（Asp）或tRNA（i）（Met）转化为有效的赖氨酸受体tRNA所必需和充分的。因此，tRNA的受体茎和tIF结构域之间的非特异性RNA-蛋白质相互作用不会放松哺乳动物LysRS的tRNA特异性。全长同源tRNA与合成酶的相互作用的可能性，需要诱导的催化中心的酶成生产构象进行了讨论。
Mammalian lysyl-tRNA synthetase (LysRS) has an N-terminal polypeptide chain extension appended to a prokaryotic-like synthetase domain. This extension, termed a tRNA-interacting factor (tIF), possesses a RNA-binding motif [KxxxK(K/R)xxK] that binds nonspecifically the acceptor T Psi C stem-loop domain of tRNA and provides a potent tRNA binding capacity to this enzyme. Consequently, native LysRS aminoacylates a RNA minihelix mimicking the amino acid acceptor stem-loop domain of tRNA(3)(Lys). Here, examination of minihelix recognition showed that mammalian LysRS aminoacylates RNA minihelices without specificity of sequence, revealing that none of the nucleotides from the acceptor T Psi C stem-loop domain are essential determinants of tRNA(Lys) acceptor identity. To test whether the tIF domain reduces the specificity of the synthetase with regard to complete tRNA molecules, aminoacylation of wild-type and mutant noncognate tRNAs by wild-type or N-terminally truncated LysRS was examined. The presence of the UUU anticodon of tRNA(Lys) appeared to be necessary and sufficient to transform yeast tRNA(Asp) or tRNA(i)(Met) into potent lysine acceptor tRNAs. Thus, nonspecific RNA-protein interactions between the acceptor stem of tRNA and the tIF domain do not relax the tRNA specificity of mammalian LysRS. The possibility that interaction of the full-length cognate tRNA with the synthetase is required to induce the catalytic center of the enzyme into a productive conformation is discussed.