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首页> 外文期刊>IUCrJ >A cytosine modification mechanism revealed by the structure of a ternary complex of deoxycytidylate hydroxymethylase from bacteriophage T4 with its cofactor and substrate
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A cytosine modification mechanism revealed by the structure of a ternary complex of deoxycytidylate hydroxymethylase from bacteriophage T4 with its cofactor and substrate

机译：通过噬菌体T4的脱氧胞苷酸羟甲基酶三元复合物及其辅因子和底物的结构揭示了胞嘧啶修饰机制

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To protect viral DNA against the host bacterial restriction system, bacterio­phages utilize a special modification system – hydroxymethylation – in which dCMP hydroxymethylase (dCH) converts dCMP to 5-hydroxymethyl-dCMP (5hm-dCMP) using N5,N10-methylenetetrahydrofolate as a cofactor. Despite shared similarity with thymidylate synthase (TS), dCH catalyzes hydroxylation through an exocyclic methylene intermediate during the last step, which is different from the hydride transfer that occurs with TS. In contrast to the extensively studied TS, the hydroxymethylation mechanism of a cytosine base is not well understood due to the lack of a ternary complex structure of dCH in the presence of both its substrate and cofactor. This paper reports the crystal structure of the ternary complex of dCH from bacteriophage T4 (T4dCH) with dCMP and tetrahydrofolate at 1.9 Å resolution. The authors found key residues of T4dCH for accommodating the cofactor without a C-terminal tail, an optimized network of ordered water molecules and a hydrophobic gating mechanism for cofactor regulation. In combination with biochemical data on structure-based mutants, key residues within T4dCH and a substrate water molecule for hydroxymethylation were identified. Based on these results, a complete enzyme mechanism of dCH and signature residues that can identify dCH enzymes within the TS family have been proposed. These findings provide a fundamental basis for understanding the pyrimidine modification system.
机译：为了保护病毒DNA不受宿主细菌限制系统的侵害，噬菌体利用一种特殊的修饰系统-羟甲基化-在其中dCMP羟甲基酶（dCH）使用N5，N10-亚甲基四氢叶酸作为辅助因子将dCMP转化为5-羟甲基-dCMP（5hm-dCMP）。尽管与胸苷酸合酶（TS）具有共同的相似性，但dCH在最后一步中通过环外亚甲基中间体催化羟基化，这与TS发生的氢化物转移不同。与广泛研究的TS相反，由于在其底物和辅因子均存在的情况下，dCH缺乏三元复合结构，因此胞嘧啶碱基的羟甲基化机理尚未得到很好的理解。本文报道了噬菌体T4（T4dCH）与dCMP和四氢叶酸的dCH三元复合物的晶体结构，分辨率为1.9Å。作者发现T4dCH的关键残基可容纳无C末端尾部的辅因子，优化的有序水分子网络和用于辅因子调节的疏水门控机制。结合基于结构的突变体的生化数据，鉴定了T4dCH内的关键残基和羟甲基化的底物水分子。基于这些结果，已经提出了dCH的完整酶机制和可以鉴定TS家族内dCH酶的特征残基。这些发现为理解嘧啶修饰系统提供了基础。

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