Conformational transitions of an unmodified tRNA: Implications for RNA folding
Conformational transitions of an unmodified tRNA: Implications for RNA folding
复制标题
DOI：
10.1021/bi981722u
复制
发表时间：
1998-11-17
期刊：
影响因子：
2.9
通讯作者：
Glick, GD
中科院分区：
文献类型：
--
作者：
Maglott, EJ;Deo, SS;Glick, GD
未修饰的tRNA是研究转录后修饰和定点突变对这些核糖核酸的结构和功能的影响的强大系统。为了确定合成构建体作为天然tRNA模型的一般限制，有必要阐明未修饰的tRNA作为溶液条件的函数的构象状态。在这里，我们报告的构象特性的未修饰的酵母tRNA（苯丙氨酸）作为离子强度的函数，[Mg 2 +]，和温度使用光谱测量沿着与化学和酶的探针的组合。我们发现，在低[Naf]缓冲液在低温下，天然酵母tRNAPhe采用三级结构的Mg 2+的情况下。相比之下，未修饰的酵母tRNA（Phe）的三级折叠对Mg 2+有绝对的需求。低于苜蓿叶的熔化温度，未修饰的酵母tRNAPhe存在于二级和三级结构之间的Mg 2+依赖性平衡中。综上所述，我们的研究结果表明，虽然tRNA的三级结构是广泛可比的，三级折叠的内在稳定性，中间状态的构象特性，和中间状态的稳定性可以显着不同的tRNA序列。因此，使用未修饰的tRNA作为天然构建体的模型可能具有显著的局限性。关于“tRNA折叠”的广泛结论必须谨慎看待，特别是在结构发生变化的情况下，如在蛋白质合成过程中。
Unmodified tRNAs are powerful systems to study the effects of posttranscriptional modifications and site-directed mutations on both the structure and function of these ribonucleic acids. To define the general limitations of synthetic constructs as models for native tRNAs, it is necessary to elucidate the conformational states of unmodified tRNAs as a function of solution conditions. Here we report the conformational properties of unmodified yeast tRNA(Phe) as a function of ionic strength, [Mg2+], and temperature using a combination of spectroscopic measurements along with chemical and enzymatic probes. We find that in low [Naf] buffer at low temperature, native yeast tRNAPhe adopts tertiary structure in the absence of Mg2+. By contrast, tertiary folding of unmodified yeast tRNA(Phe) has an absolute requirement for Mg2+. Below the melting temperature of the cloverleaf, unmodified yeast tRNAPhe exists in a Mg2+ dependent equilibrium between secondary and tertiary structure. Taken together, our findings suggest that although the tertiary structures of tRNAs are broadly comparable, the intrinsic stability of the tertiary fold, the conformational properties of intermediate states, and the stability of intermediate states can differ significantly between tRNA sequences. Thus, the use of unmodified tRNAs as models for native constructs can have significant limitations. Broad conclusions regarding "tRNA folding" as a whole must be viewed cautiously, particularly in cases where structural changes occur, such as during protein synthesis.