Pallanck L, Schulman L H
Department of Developmental Biology and Cancer, Albert Einstein College of Medicine, Bronx, NY 10461.
Proc Natl Acad Sci U S A. 1991 May 1;88(9):3872-6. doi: 10.1073/pnas.88.9.3872.
An assay based on the initiation of protein synthesis in Escherichia coli has been used to explore the role of the anticodon in tRNA identity in vivo. Mutations were introduced into the initiator tRNA to change the wild-type anticodon from CAU (methionine) to GAU (isoleucine), GAC (valine), and GAA (phenylalanine), where each derivative differs from the preceding by a single base change in the anticodon (underlined). These changes were sufficient to cause the mutant tRNAs to be aminoacylated by the corresponding aminoacyl-tRNA synthetases based on the amino acid inserted into protein from complementary initiation codons. Construction of additional single base anticodon variants (GUU, GGU, GCC, GUC, GCA, and UAA) showed that all three anticodon bases specify isoleucine and phenylalanine identity and that both the middle and the third anticodon bases are important for valine identity in vivo.
一种基于大肠杆菌中蛋白质合成起始的检测方法已被用于探索反密码子在体内tRNA特异性识别中的作用。将突变引入起始tRNA，把野生型反密码子从CAU（甲硫氨酸）改变为GAU（异亮氨酸）、GAC（缬氨酸）和GAA（苯丙氨酸），其中每个衍生物的反密码子（下划线部分）与前一个仅相差一个碱基变化。这些变化足以使突变的tRNA根据从互补起始密码子插入蛋白质中的氨基酸，被相应的氨酰-tRNA合成酶进行氨酰化。构建额外的单碱基反密码子变体（GUU、GGU、GCC、GUC、GCA和UAA）表明，所有三个反密码子碱基都决定异亮氨酸和苯丙氨酸的特异性识别，并且反密码子的中间碱基和第三个碱基在体内对缬氨酸的特异性识别都很重要。
