Fudan University Pudong Medical Center, and Shanghai Key Laboratory of Medical Epigenetics, International Laboratory of Medical Epigenetics and Metabolism, Ministry of Science and Technology, Institutes of Biomedical Sciences, Fudan University, Shanghai, China
Key Laboratory of Metabolism and Molecular Medicine, Ministry of Education, School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai, China
Shanghai Public Health Clinical Center, State Key Laboratory of Genetic Engineering, Collaborative Innovation Center of Genetics and Development, Department of Biochemistry and Biophysics, School of Life Sciences, Fudan University, Shanghai, China
RNA 甲基化有助于广泛的细胞功能。细胞 RNA 通常被蛋白质甲基转移酶甲基化，使用S-腺苷-1-蛋氨酸(SAM) 作为辅因子。在这里，我们报告了从随机序列中体外选择 33 个核苷酸的 SAM 依赖性甲基转移酶核酶 RNA。RNA 上甲基的检测和作图表明N 7的位点特异性甲基化SAM的鸟嘌呤位置。核酶在很宽的 pH 值和温度范围内都具有活性，并且与蛋白质酶相比更加稳定。在存在和不存在 SAM 的情况下，核酶结构显示出与辅因子结合相关的剧烈局部构象变化。发现核酶基序分布在自然界中，并且候选序列在体外被证明是有活性的。这种使用辅因子 SAM 特异性甲基化 RNA 的核酶的发现开启了甲基转移酶核酶可能有助于细胞 RNA 甲基化的可能性。
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The identification and characterization of a selected SAM-dependent methyltransferase ribozyme that is present in natural sequences
RNA methylations contribute to a wide range of cellular functions. Cellular RNAs are usually methylated by protein methyltransferases using S-adenosyl-l-methionine (SAM) as a cofactor. Here we report the in vitro selection of a 33-nucleotide SAM-dependent methyltransferase ribozyme RNA from a randomized sequence. Detection and mapping of the methyl group on the RNA demonstrates site-specific methylation of the N7 position of guanine by SAM. The ribozyme is active over a wide range of pH and temperatures and is robust compared to protein enzymes. The ribozyme structures in the presence and absence of SAM show a dramatic local conformational change associated with cofactor binding. The ribozyme motif was found to be distributed in nature, and candidate sequences were shown to be active in vitro. The discovery of this ribozyme that uses the cofactor SAM to specifically methylate RNA opens up the possibility that methyltransferase ribozymes may contribute to cellular RNA methylation.
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