Strassler Sarah E, Bowles Isobel E, Krishnamohan Aiswarya, Kim Hyejeong, Edgington Catherine B, Kuiper Emily G, Hancock Clio J, Comstock Lindsay R, Jackman Jane E, Conn Graeme L
Department of Biochemistry, Emory University School of Medicine, 1510 Clifton Road NE, Atlanta GA, 30322, USA.
Graduate Program in Biochemistry, Cell and Developmental Biology, Graduate Division of Biological and Biomedical Sciences, Emory University.
bioRxiv. 2023 Oct 19:2023.02.01.526536. doi: 10.1101/2023.02.01.526536.
The methyltransferase Trm10 modifies a subset of tRNAs on the base N1 position of the 9th nucleotide in the tRNA core. Trm10 is conserved throughout Eukarya and Archaea, and mutations in the human gene () have been linked to neurological disorders such as microcephaly and intellectual disability, as well as defects in glucose metabolism. Of the 26 tRNAs in yeast with guanosine at position 9, only 14 are substrates for Trm10. However, no common sequence or other posttranscriptional modifications have been identified among these substrates, suggesting the presence of some other tRNA feature(s) which allow Trm10 to distinguish substrate from nonsubstrate tRNAs. Here, we show that substrate recognition by Trm10 is dependent on both intrinsic tRNA flexibility and the ability of the enzyme to induce specific tRNA conformational changes upon binding. Using the sensitive RNA structure-probing method SHAPE, conformational changes upon binding to Trm10 in tRNA substrates, but not nonsubstrates, were identified and mapped onto a model of Trm10-bound tRNA. These changes may play an important role in substrate recognition by allowing Trm10 to gain access to the target nucleotide. Our results highlight a novel mechanism of substrate recognition by a conserved tRNA modifying enzyme. Further, these studies reveal a strategy for substrate recognition that may be broadly employed by tRNA-modifying enzymes which must distinguish between structurally similar tRNA species.
甲基转移酶Trm10修饰tRNA核心区域第9个核苷酸的N1位上的一部分tRNA。Trm10在整个真核生物和古细菌中都保守存在，人类基因（）中的突变与小头畸形和智力残疾等神经疾病以及葡萄糖代谢缺陷有关。在酵母中第9位为鸟苷的26种tRNA中，只有14种是Trm10的底物。然而，在这些底物中尚未鉴定出共同序列或其他转录后修饰，这表明存在一些其他tRNA特征，使得Trm10能够区分底物tRNA和非底物tRNA。在这里，我们表明Trm10对底物的识别既依赖于tRNA固有的灵活性，也依赖于该酶在结合时诱导特定tRNA构象变化的能力。使用灵敏的RNA结构探测方法SHAPE，鉴定了tRNA底物（而非非底物）与Trm10结合时的构象变化，并将其映射到Trm10结合tRNA的模型上。这些变化可能通过使Trm10能够接近靶核苷酸而在底物识别中发挥重要作用。我们的结果突出了一种保守的tRNA修饰酶识别底物的新机制。此外，这些研究揭示了一种底物识别策略，可能被必须区分结构相似tRNA种类的tRNA修饰酶广泛采用。
