Department of Chemistry, Institute of Biochemistry, University of Münster, Corrensstr. 36, 48149, Münster, Germany.
Current address: ETH Zürich, Department of Chemistry and Applied Biosciences, Laboratory of Organic Chemistry, Vladimir-Prelog-Weg 1-5/10, 8093, Zürich, Switzerland.
Angew Chem Int Ed Engl. 2021 Jan 4;60(1):480-485. doi: 10.1002/anie.202012623. Epub 2020 Nov 13.
Methylation and demethylation of DNA, RNA and proteins has emerged as a major regulatory mechanism. Studying the function of these modifications would benefit from tools for their site-specific inhibition and timed removal. S-Adenosyl-L-methionine (AdoMet) analogs in combination with methyltransferases (MTases) have proven useful to map or block and release MTase target sites, however their enzymatic generation has been limited to aliphatic groups at the sulfur atom. We engineered a SAM synthetase from Cryptosporidium hominis (PC-ChMAT) for efficient generation of AdoMet analogs with photocaging groups that are not accepted by any WT MAT reported to date. The crystal structure of PC-ChMAT at 1.87 Å revealed how the photocaged AdoMet analog is accommodated and guided engineering of a thermostable MAT from Methanocaldococcus jannaschii. PC-MATs were compatible with DNA- and RNA-MTases, enabling sequence-specific modification ("writing") of plasmid DNA and light-triggered removal ("erasing").
DNA、RNA 和蛋白质的甲基化和去甲基化已成为主要的调控机制。研究这些修饰的功能将受益于其特异性抑制和定时去除的工具。S-腺苷甲硫氨酸（AdoMet）类似物与甲基转移酶（MTases）结合已被证明可用于绘制或阻断和释放 MTase 靶标位点，但它们的酶促生成仅限于硫原子上的脂肪族基团。我们从隐孢子虫（Cryptosporidium hominis）（PC-ChMAT）中设计了一种 SAM 合酶，用于有效生成带有光笼基团的 AdoMet 类似物，这些基团迄今尚未被任何 WT MAT 接受。PC-ChMAT 的 1.87 Å 晶体结构揭示了光笼 AdoMet 类似物是如何被容纳的，并指导了来自产甲烷球菌（Methanocaldococcus jannaschii）的热稳定 MAT 的工程设计。PC-MAT 与 DNA 和 RNA-MTases 兼容，能够对质粒 DNA 进行序列特异性修饰（“写入”）和光触发去除（“擦除”）。
