High-Throughput-Methyl-Reading (HTMR) assay: a solution based on nucleotide methyl-binding proteins enables large-scale screening for DNA/RNA methyltransferases and demethylases

doi:10.1093/nar/gkab989

	High-Throughput-Methyl-Reading (HTMR) assay: a solution based on nucleotide methyl-binding proteins enables large-scale screening for DNA/RNA methyltransferases and demethylases
	Xiao, Senhao1,2,3 ; Guo, Siqi 1,5; Han, Jie 1; Sun, Yanli 1,6,7; Wang, Mingchen1,2,3 ; Chen, Yantao 1; Fang, Xueyu1,2,3 ; Yang, Feng 1; Mu, Yajuan 8; Zhang, Liang 8; Ding, Yiluan 3,4; Zhang, Naixia 3,4; Jiang, Hualiang1,2,3 ; Chen, Kaixian1,2,3 ; Zhao, Kehao 6,7; Luo, Cheng 1,2,3,9; Chen, Shijie 1,3,9,10
	2022-01-25
发表期刊	NUCLEIC ACIDS RESEARCH (IF:16.6[JCR-2023],16.1[5-Year])
ISSN	0305-1048
EISSN	1362-4962
卷号	50 期号:2
发表状态	已发表
DOI	10.1093/nar/gkab989
摘要	Epigenetic therapy has significant potential for cancer treatment. However, few small potent molecules have been identified against DNA or RNA modification regulatory proteins. Current approaches for activity detection of DNA/RNA methyltransferases and demethylases are time-consuming and labor-intensive, making it difficult to subject them to high-throughput screening. Here, we developed a fluorescence polarization-based 'High-Throughput Methyl Reading' (HTMR) assay to implement large-scale compound screening for DNA/RNA methyltransferases and demethylases-DNMTs, TETs, ALKBH5 and METTL3/METTL14. This assay is simple to perform in a mix-and-read manner by adding the methyl-binding proteins MBD1 or YTHDF1. The proteins can be used to distinguish FAM-labelled substrates or product oligonucleotides with different methylation statuses catalyzed by enzymes. Therefore, the extent of the enzymatic reactions can be coupled with the variation of FP binding signals. Furthermore, this assay can be effectively used to conduct a cofactor competition study. Based on the assay, we identified two natural products as candidate compounds for DNMT1 and ALKBH5. In summary, this study outlines a powerful homogeneous approach for high-throughput screening and evaluating enzymatic activity for DNA/RNA methyltransferases and demethylases that is cheap, easy, quick, and highly sensitive.
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收录类别	SCI ; SCIE
语种	英语
资助项目	National Natural Science Foundation of China[81703415,91853205,81625022,81821005,21820102008,81728020] ; Science and Technology Commission of Shanghai Municipality["21ZR1474700","19XD1404700",18431907100]
WOS研究方向	Biochemistry & Molecular Biology
WOS类目	Biochemistry & Molecular Biology
WOS记录号	WOS:000763001100003
出版者	OXFORD UNIV PRESS
引用统计	正在获取...
文献类型	期刊论文
条目标识符	https://kms.shanghaitech.edu.cn/handle/2MSLDSTB/165025
专题	生命科学与技术学院_博士生生命科学与技术学院_特聘教授组_陈凯先组免疫化学研究所_特聘教授组_蒋华良组生命科学与技术学院_硕士生
通讯作者	Chen, Shijie
作者单位	1.Chinese Acad Sci, Shanghai Inst Mat Med, State Key Lab Drug Res, Ctr Chem Biol,Drug Discovery & Design Ctr, Shanghai 201203, Peoples R China 2.ShanghaiTech Univ, Sch Life Sci & Technol, Shanghai 201210, Peoples R China 3.Univ Chinese Acad Sci, 19A Yuquan Rd, Beijing 100049, Peoples R China 4.Chinese Acad Sci, Analyt Res Ctr Organ & Biol Mol, Shanghai Inst Mat Med, Shanghai 201203, Peoples R China 5.Nanchang Univ, Sch Pharm, Nanchang 330006, Jiangxi, Peoples R China 6.Yantai Univ, Sch Pharm, Key Lab Mol Pharmacol & Drug Evaluat, Minist Educ, Yantai 264005, Peoples R China 7.Yantai Univ, Collaborat Innovat Ctr Adv Drug Delivery Syst & B, Yantai 264005, Peoples R China 8.Shanghai Jiao Tong Univ, Dept Pharmacol & Chem Biol, State Key Lab Oncogenes & Related Genes, Sch Med, Shanghai 200025, Peoples R China 9.UCAS, Hangzhou Inst Adv Study, Sch Pharmaceut Sci & Technol, Hangzhou 310024, Peoples R China 10.Chinese Acad Sci, Shanghai Inst Mat Med, Drug Discovery & Design Ctr, Ctr Chem Biol, Zuchongzhi Rd 555, Shanghai 201203, Peoples R China
第一作者单位	生命科学与技术学院
推荐引用方式 GB/T 7714	Xiao, Senhao,Guo, Siqi,Han, Jie,et al. High-Throughput-Methyl-Reading (HTMR) assay: a solution based on nucleotide methyl-binding proteins enables large-scale screening for DNA/RNA methyltransferases and demethylases[J]. NUCLEIC ACIDS RESEARCH,2022,50(2).
APA	Xiao, Senhao.,Guo, Siqi.,Han, Jie.,Sun, Yanli.,Wang, Mingchen.,...&Chen, Shijie.(2022).High-Throughput-Methyl-Reading (HTMR) assay: a solution based on nucleotide methyl-binding proteins enables large-scale screening for DNA/RNA methyltransferases and demethylases.NUCLEIC ACIDS RESEARCH,50(2).
MLA	Xiao, Senhao,et al."High-Throughput-Methyl-Reading (HTMR) assay: a solution based on nucleotide methyl-binding proteins enables large-scale screening for DNA/RNA methyltransferases and demethylases".NUCLEIC ACIDS RESEARCH 50.2(2022).