PROBING STRUCTURAL DIFFERENCES BETWEEN NATIVE AND IN-VITRO TRANSCRIBED ESCHERICHIA-COLI VALINE TRANSFER-RNA - EVIDENCE FOR STABLE BASE MODIFICATION-DEPENDENT CONFORMERS
PROBING STRUCTURAL DIFFERENCES BETWEEN NATIVE AND IN-VITRO TRANSCRIBED ESCHERICHIA-COLI VALINE TRANSFER-RNA - EVIDENCE FOR STABLE BASE MODIFICATION-DEPENDENT CONFORMERS
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DOI：
10.1093/nar/21.21.4948
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发表时间：
1993-10-25
影响因子：
14.9
通讯作者：
HOROWITZ, J
中科院分区：
文献类型：
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作者：
DERRICK, WB;HOROWITZ, J
天然（修改）和体外转录（未修改）的大肠杆菌tRNA（瓦尔）之间的结构差异进行了探讨，通过比较其温度-吸光度曲线作为镁离子浓度的函数，并通过探测其溶液构象与单链和双链特异性核酸内切酶。体外转录的tRNA（瓦尔）具有较不有序的结构，如通过热解链曲线监测的;明显低于天然tRNA（瓦尔）。用核酸酶S1和核糖核酸酶V1进行的结构探测实验表明，未修饰的tRNA（瓦尔）转录物在低Mg 2+浓度下更容易受到核酸酶攻击，特别是在D-环和T-环中，这表明D-环/T-环相互作用至少部分破坏。这些实验还提供了天然tRNA（瓦尔）反密码子环的温度依赖性替代构象的证据。修饰的核苷对于这些构象异构体的稳定性是必不可少的;它们不能在未修饰的体外转录物中检测到。观察结果表明，tRNA的转录后修饰允许采用独特的构象，并起到稳定那些具有生物活性的构象的作用。
Structural differences between native (modified) and in vitro transcribed (unmodified) Escherichia coli tRNA(Val) were explored by comparing their temperature-absorbance profiles as a function of magnesium ion concentration and by probing their solution conformation with single- and double-strand-specific endonucleases. In vitro transcribed tRNA(Val) has a less ordered structure as monitored by thermal melting profiles; its T. is appreciably lower than that of native tRNA(Val) at all Mg2+ concentrations. Structure probing experiments with nuclease S1 and ribonuclease V1 show that the unmodified tRNA(Val) transcript is more susceptible to nuclease attack at low Mg2+ concentrations, particularly in the D- and T-loops, indicative of at least a partial disruption of D-loop/T-loop interactions. These experiments also provide evidence for temperature-dependent alternative conformations of the anticodon loop of native tRNA(Val). Modified nucleosides are essential for the stability of these conformers; they cannot be detected in the unmodified in vitro transcript. The observations suggest that post-transcriptional modifications in tRNA allow the adoption of unique conformations and act to stabilize those that are biologically active.