Variant minihelix RNAs reveal sequence‐specific recognition of the helical tRNA(Ser) acceptor stem by E.coli seryl‐tRNA synthetase.

Variant minihelix RNAs reveal sequence‐specific recognition of the helical tRNA(Ser) acceptor stem by E.coli seryl‐tRNA synthetase.
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变体微型螺旋 RNA 揭示了大肠杆菌丝氨酰-tRNA 合成酶对螺旋 tRNA(Ser) 受体茎的序列特异性识别。

DOI:
10.1002/j.1460-2075.1996.tb00645.x
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发表时间:
1996
期刊:
The EMBO Journal
影响因子:
--
通讯作者:
Jeffrey R. Sampson
Jeffrey R. Sampson
中科院分区:
--
文献类型:
--
作者:
M. Saks;Jeffrey R. Sampson

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对一系列变体 RNA 小螺旋底物的氨酰化率测定表明,大肠杆菌丝氨酰 tRNA 合成酶 (SerRS) 识别大肠杆菌 tRNA(Ser) 受体茎的 1-72 至 5-68 个碱基对,主要识别元件聚集在位置 2-71 和 4-69 之间。 kcat/Km 上每个位置的规范碱基对取代的影响的排序用于评估主要沟功能组在识别中的参与。基于生化数据的结论与 Cusack 及其同事在随附论文中报告的同源嗜热栖热菌 tRNA(Ser)-SerRS 复合物的精细结构所揭示的相互作用基本一致。嘧啶 69 的主沟 C5(H) 与 SerRS 的芳香族侧链之间的端对疏水相互作用的破坏被证明会显着降低微型螺旋底物的 kcat/Km。这种类型的相互作用提供了一种方法,使蛋白质能够识别“简并”序列的二元信息,例如 tRNA(Ser) 的嘌呤-嘧啶碱基对。 3-70 个碱基对被证明有助于 SerRS 的识别,即使它没有与蛋白质特异性接触。后一种效应源自 SerRS 与邻近的 2-71 和 4-69 受体茎碱基对的特定接触的组织。
Aminoacylation rate determinations for a series of variant RNA minihelix substrates revealed that Escherichia coli seryl‐tRNA synthetase (SerRS) recognizes the 1–72 through 5–68 base pairs of the E.coli tRNA(Ser) acceptor stem with the major recognition elements clustered between positions 2–71 and 4–69. The rank order of effects of canonical base pair substitutions at each position on kcat/Km was used to assess the involvement of major groove functional groups in recognition. Conclusions based on the biochemical data are largely consistent with the interactions revealed by the refined structure of the homologous Thermus thermophilus tRNA(Ser)‐SerRS complex that Cusack and colleagues report in the accompanying paper. Disruption of an end‐on hydrophobic interaction between the major groove C5(H) of pyrimidine 69 and an aromatic side chain of SerRS is shown to significantly decrease kcat/Km of a minihelix substrate. This type of interaction provides a means by which proteins can recognize the binary information of ‘degenerate’ sequences, such as the purine‐pyrimidine base pairs of tRNA(Ser). The 3–70 base pair is shown to contribute to recognition by SerRS even though it is not contacted specifically by the protein. The latter effect derives from the organization of the specific contacts that SerRS makes with the neighboring 2–71 and 4–69 acceptor stem base pairs.
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