Hagervall T G, Tuohy T M, Atkins J F, Björk G R
Department of Microbiology, University of Umeå, Sweden.
J Mol Biol. 1993 Aug 5;232(3):756-65. doi: 10.1006/jmbi.1993.1429.
The trmD gene encodes the tRNA(m1G37)methyltransferase, which methylates guanosine (G) to 1-methylguanosine (m1G) at position 37 of tRNAs that read CUN (leucine), CCN (proline), and CGG (arginine) codons. A mutant, trmD3, has previously been isolated, which at high temperature lacks m1G in tRNA, and this deficiency was correlated with a +1 frameshifting activity. In this study, the mechanism of this trmD3-induced frameshift involving mutant tRNA(Pro) and tRNA(Leu) species has been investigated. Potential frameshifting sites for proline tRNAs, CCC-N, were efficiently suppressed in the mutant strain. Hybrid beta-galactosidases encoded by plasmid constructs containing the sites CCC-U and CCC-A were subjected to amino-terminal sequencing. The protein sequences demonstrated that a quadruplet translocation had occurred and that a proline was inserted at these sites, suggesting that a tRNA(Pro) deficient in m1G is the frameshifting agent. Therefore, a mechanism involving a quadruplet codon-anticodon interaction is favoured for trmD3-dependent +1 frameshifting. Of the four potential sites for tRNA(Leu) (CCU-N), two, CCU-U and CCU-C, were significantly suppressed in the trmD3 mutant. Thus, species of tRNA(Leu) may also act as +1 frameshift suppressors. No -1 frameshifting activity was found with the trmD3 mutant.
trmD基因编码tRNA（m1G37）甲基转移酶，该酶可将读取CUN（亮氨酸）、CCN（脯氨酸）和CGG（精氨酸）密码子的tRNA第37位的鸟苷（G）甲基化为1-甲基鸟苷（m1G）。此前已分离出一个突变体trmD3，其在高温下tRNA中缺乏m1G，这种缺陷与+1移码活性相关。在本研究中, 对这种由trmD3诱导的涉及突变型tRNA（Pro）和tRNA（Leu）的移码机制进行了研究。脯氨酸tRNA的潜在移码位点CCC-N在突变菌株中被有效抑制。对含有CCC-U和CCC-A位点的质粒构建体编码的杂合β-半乳糖苷酶进行氨基末端测序。蛋白质序列表明发生了四联体易位，并且在这些位点插入了脯氨酸，这表明缺乏m1G的tRNA（Pro）是移码因子。因此，trmD3依赖的+1移码倾向于涉及四联体密码子-反密码子相互作用的机制。在tRNA（Leu）的四个潜在位点（CCU-N）中，有两个位点CCU-U和CCU-C在trmD3突变体中被显著抑制。因此，tRNA（Leu）的种类也可能作为+1移码抑制因子。未发现trmD3突变体具有-1移码活性。
