Frugier M, Florentz C, Giegé R
Unité Structure des Macromolécules Biologiques et Mécanismes de Reconnaissance, Centre National de la Recherche Scientifique, Strasbourg, France.
EMBO J. 1994 May 1;13(9):2218-26. doi: 10.1002/j.1460-2075.1994.tb06499.x.
We show here that small RNA helices which recapitulate part or all of the acceptor stem of yeast aspartate tRNA are efficiently aminoacylated by cognate class II aspartyl-tRNA synthetase. Aminoacylation is strongly dependent on the presence of the single-stranded G73 'discriminator' identity nucleotide and is essentially insensitive to the sequence of the helical region. Substrates which contain as few as 3 bp fused to G73CCAOH are aspartylated. Their charging is insensitive to the sequence of the loop closing the short helical domains. Aminoacylation of the aspartate mini-helix is not stimulated by a hairpin helix mimicking the anticodon domain and containing the three major anticodon identity nucleotides. A thermodynamic analysis demonstrates that enzyme interactions with G73 in the resected RNA substrates and in the whole tRNA are the same. Thus, if the resected RNA molecules resemble in some way the earliest substrates for aminoacylation with aspartate, then the contemporary tRNA(Asp) has quantitatively retained the influence of the major signal for aminoacylation in these substrates.
我们在此表明，模拟酵母天冬氨酸tRNA受体茎部分或全部的小RNA螺旋，能被同源的II类天冬氨酰 - tRNA合成酶高效氨酰化。氨酰化强烈依赖于单链G73“判别”识别核苷酸的存在，并且对螺旋区域的序列基本不敏感。含有与G73CCAOH融合的低至3个碱基对的底物会被天冬氨酰化。它们的负载对封闭短螺旋结构域的环的序列不敏感。模拟反密码子结构域并包含三个主要反密码子识别核苷酸的发夹螺旋不会刺激天冬氨酸小螺旋的氨酰化。热力学分析表明，酶与切除的RNA底物以及完整tRNA中G73的相互作用是相同的。因此，如果切除的RNA分子在某种程度上类似于最早的天冬氨酸氨酰化底物，那么当代的tRNA(Asp)在数量上保留了这些底物中氨酰化主要信号的影响。
