Danna Dong, Mingyu Xia, Sili Wang, Pengfei Fang, Wen Liu
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引用次数: 0
摘要
丝裂霉素是由链霉菌产生的一类天然分子,具有很强的抗菌和抗肿瘤活性。MitM 是一种参与丝裂霉素生物合成的关键后mitosane修饰酶。此前曾有研究认为,该蛋白作为一种 N-甲基转移酶,可催化丝裂霉素 A 和丝裂霉素中间体 9a-demethyl-mitomycin A 的氮丙啶甲基化。MitM 识别辅助因子 S-腺苷-l-蛋氨酸(SAM)和底物丝裂霉素 A 的结构基础尚不清楚。在这里,我们测定了 apo-MitM 和 MitM-mitomycin A-S-adenosylhomocysteine (SAH) 三元复合物的晶体结构,分辨率分别为 2.23 和 2.80 Å。我们发现 MitM 采用一类 SAM 依赖性甲基转移酶折叠,并在溶液中形成同源二聚体。一系列参与形成活性口袋的残基发生了构象变化,这有助于 MitM 与 SAH 和丝裂霉素 A 结合。当丝裂霉素 A 结合时,该环路的弯曲方向发生逆转,使活性位点的入口从开放变为封闭。这项研究从结构上揭示了 MitM 参与丝裂霉素生物合成的后mitosane 阶段,并为甲基转移酶的工程设计提供了模板。
Structural Basis of Substrate Recognition by the Postmitosane Modification Enzyme MitM in Mitomycin Biosynthesis.
Mitomycins make up a class of natural molecules produced by Streptomyces with strong antibacterial and antitumor activities. MitM is a key postmitosane modification enzyme involved in mitomycin biosynthesis in Streptomyces caespitosus. This protein was previously suggested to catalyze the aziridinium methylation of mitomycin A and the mitomycin intermediate 9a-demethyl-mitomycin A as an N-methyltransferase. The structural basis for MitM to recognize cofactor S-adenosyl-l-methionine (SAM) and substrate mitomycin A is unknown. Here, we determined the crystal structures of apo-MitM and MitM-mitomycin A-S-adenosylhomocysteine (SAH) ternary complexes with resolutions of 2.23 and 2.80 Å, respectively. We found that MitM adopts a class I SAM-dependent methyltransferase fold and forms a homodimer in solution. Conformational changes in a series of residues involved in the formation of active pockets assist MitM in binding SAH and mitomycin A. In particular, the 28ALGAASLGE36 loop changes most significantly. When mitomycin A binds, the bending direction of this loop is reversed, changing the entrance of the active site from open to closed. This study provides structural insights into MitM's involvement in the postmitosane stage of mitomycin biosynthesis and provides a template for the engineering of methyltransferases.
期刊介绍:
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