Caron M, Dugas H
Nucleic Acids Res. 1976 Jan;3(1):35-47. doi: 10.1093/nar/3.1.35.
The molecular mechanism of thermal unfolding of E. coli tRNAGlu, tRNAfMet and tRNAPhe (in 0.02M Tris-HC1, pH 7.5. 10 MM Mg C12) has been examined by the spin-labeling technique. The rate of tumbling of the spin label has been measured as a function of temperature for ten different selectively spin-labeled tRNAs. Only spin labels at position s4U-8 were able to probe the tertiary structure. Evidences are presented which support the hypothesis that the thermal denaturation of the three species of tRNAs studied is sequential. The unfolding process occurs in three discrete stages. The first step (30 degrees-32 degrees) could either be assigned to a localized reorganization of the cold-denatured structure or to a "transient" melting, followed by the simultaneous disruption of the tertiary structure and part of the hU helix. This transition is observed even in the absence of magnesium. The second step (50 degrees-54 degrees) involves the melting of the anticodon and miniloop regions. The last step occurs above 65 degrees where the t psi c and amino acid acceptor stems, forming one continuous double helix, melt. A simple dynamic model is considered for tRNA function in protein biosynthesis.
通过自旋标记技术研究了大肠杆菌tRNAGlu、tRNAfMet和tRNAPhe（在0.02M Tris-HCl，pH 7.5，10mM MgCl2中）热解折叠的分子机制。对于十种不同的选择性自旋标记tRNA，测量了自旋标记的翻转速率随温度的变化。只有位于s4U-8位置的自旋标记能够探测三级结构。提供的证据支持了所研究的三种tRNA热变性是有序的这一假设。解折叠过程发生在三个离散阶段。第一步（30℃-32℃）要么归因于冷变性结构的局部重组，要么归因于“瞬时”解链，随后是三级结构和部分hU螺旋的同时破坏。即使在没有镁的情况下也能观察到这种转变。第二步（50℃-54℃）涉及反密码子和小环区域的解链。最后一步发生在65℃以上，此时tψc和氨基酸接受茎形成一个连续的双螺旋并解链。考虑了一个关于tRNA在蛋白质生物合成中功能的简单动力学模型。
