Jank P, Riesner D, Gross H J
Nucleic Acids Res. 1977 Jun;4(6):2009-200. doi: 10.1093/nar/4.6.2009.
In contrast to all other known tRNAs, mammalian tRNA1Val contains two adenosines A59 and A60, opposite to U54 and psi 55 in the U psi CG sequence of the T psi C loop, which could form unusual A:U (or A: psi pairs in addition to the five "normal" G:C pairs. In order to measure the number of G:C and A:U (A: psi) pairs in the T psi C stem, we prepared the 30 nucleotide long 3'-terminal fragment of this tRNA by "m7G-cleavage". From differentiated melting curves and temperature jump experiments it was concluded that the T psi C stem in this fragment is in fact extended by an additional A60:U54 pair. A dimer of this fragment with 14 base pairs was characterized by gel electrophoresis and by the same physical methods. An additional A:U pair in the tRNA1Val fragment does not necessarily mean that this is also true for intact tRNA. However, we showed that U54 is far less available for enzymatic methylation in mammalian tRNA1Val compared to tRNA from T-E. coli. This clear difference in U54 reactivity, together with the identification of an extra A60:U54 pair in the U psi CG containing fragment suggests the presence of a 6 base pair T psi C stem and a 5 nucleotide T psi C loop in this tRNA.
与所有其他已知的转运RNA（tRNA）不同，哺乳动物的tRNA1Val在TψCG序列的TψC环中，与U54和ψ55相对应的位置含有两个腺苷A59和A60，除了五个“正常”的G:C碱基对外，这两个腺苷还可形成异常的A:U（或A:ψ）碱基对。为了测定TψC茎中G:C和A:U（A:ψ）碱基对的数量，我们通过“m7G切割”制备了该tRNA 30个核苷酸长的3'末端片段。从解链曲线和温度跳跃实验可以得出结论，该片段中的TψC茎实际上因额外的A60:U54碱基对而延长。通过凝胶电泳和相同的物理方法对该具有14个碱基对的片段二聚体进行了表征。tRNA1Val片段中额外的A:U碱基对并不一定意味着完整的tRNA也是如此。然而，我们发现与来自大肠杆菌的tRNA相比，哺乳动物tRNA1Val中的U54在酶促甲基化方面的可用性要低得多。U54反应性的这种明显差异，以及在含有UψCG的片段中鉴定出额外的A60:U54碱基对，表明该tRNA中存在一个6碱基对的TψC茎和一个5核苷酸的TψC环。
