Role of the 1-72 base pair in tRNAs for the activity of Escherichia coli peptidyl-tRNA hydrolase.
Role of the 1-72 base pair in tRNAs for the activity of Escherichia coli peptidyl-tRNA hydrolase.
复制标题
tRNA 中 1-72 碱基对对大肠杆菌肽基-tRNA 水解酶活性的作用。
DOI：
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发表时间：
1993
影响因子：
14.9
通讯作者：
Sylvain Blanquet
中科院分区：
文献类型：
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作者：
Sophie Dutka;Thierry Meinnel;C. Lazennec;Y. Mechulam;Sylvain Blanquet
Schulman和Pelka（1975）J.Biol.Chem.250，542-547的先前工作表明，起始物tRNA（fMet）中碱基1和72之间不存在配对解释了肽基-tRNA水解酶对N-乙酰基-甲硫氨酰-tRNA（fMet）的相对小的活性。在本研究中，N-乙酰基-氨酰-tRNA对大肠杆菌肽基-tRNA水解酶活性的敏感性的结构要求已被进一步研究。已经产生了10个衍生物的tRNA（fMet）与受体茎中的位置1和72处的碱基的各种组合，氨酰化和化学乙酰化。从这些tRNA衍生物的氨酰基部分的释放进行了测定的存在下，从一个过度生产的菌株纯化的肽基-tRNA水解酶。与具有C1 G72、U1 A72、G1 C72、A1 U 72或G1 U 72的tRNA（fMet）衍生物相比，具有C1 A72、C1 C72、U1 G72、U1 C72或A1 C72的tRNA（fMet）衍生物表现为较差的酶底物。除了U1 G72之外，因此可以得出结论，tRNA（fMet）对肽基-tRNA水解酶的相对抗性不取决于位置1和72处的核苷酸的特定组合，而是反映了在这些位置处不存在碱基配对。在第二系列实验中，用C-A或A-C碱基而不是甲硫氨酰-tRNA（mMet）或缬氨酰-tRNA（Val 1）中的G-C产生的1和72碱基的解配对显示出显著降低肽酰-tRNA水解酶催化的水解速率。总之，数据表明1-72对的稳定性决定了肽基-tRNA对肽基-tRNA水解酶的敏感程度。
Previous work by Schulman and Pelka (1975) J. Biol. Chem. 250, 542-547, indicated that the absence of a pairing between the bases 1 and 72 in initiator tRNA(fMet) explained the relatively small activity of peptidyl-tRNA hydrolase towards N-acetyl-methionyl-tRNA(fMet). In the present study, the structural requirements for the sensitivity of an N-acetyl-aminoacyl-tRNA to Escherichia coli peptidyl-tRNA hydrolase activity have been further investigated. Ten derivatives of tRNA(fMet) with various combinations of bases at positions 1 and 72 in the acceptor stem have been produced, aminoacylated and chemically acetylated. The release of the aminoacyl moiety from these tRNA derivatives was assayed in the presence of peptidyl-tRNA hydrolase purified from an overproducing strain. tRNA(fMet) derivatives with either C1A72, C1C72, U1G72, U1C72 or A1C72 behaved as poor substrates of the enzyme, as compared to those with C1G72, U1A72, G1C72, A1U72 or G1U72. With the exception of U1G72, it could be therefore concluded that the relative resistance of tRNA(fMet) to peptidyl-tRNA hydrolase did not depend on a particular combination of nucleotides at positions 1 and 72, but rather reflected the absence of a base pairing at these positions. In a second series of experiments, the unpairing of the 1 and 72 bases, created with C-A or A-C bases, instead of G-C in methionyl-tRNA(mMet) or in valyl-tRNA(Val1), was shown to markedly decrease the rate of hydrolysis catalysed by peptidyl-tRNA hydrolase. Altogether, the data indicate that the stability of the 1-72 pair governs the degree of sensitivity of a peptidyl-tRNA to peptidyl-tRNA hydrolase.