Doukov T, Seravalli J, Stezowski J J, Ragsdale S W
Department of Chemistry, University of Nebraska-Lincoln, 68503, USA.
Structure. 2000 Aug 15;8(8):817-30. doi: 10.1016/s0969-2126(00)00172-6.
Methyltetrahydrofolate, corrinoid iron-sulfur protein methyltransferase (MeTr), catalyzes a key step in the Wood-Ljungdahl pathway of carbon dioxide fixation. It transfers the N5-methyl group from methyltetrahydrofolate (CH3-H4folate) to a cob(I)amide center in another protein, the corrinoid iron-sulfur protein. MeTr is a member of a family of proteins that includes methionine synthase and methanogenic enzymes that activate the methyl group of methyltetra-hydromethano(or -sarcino)pterin. We report the first structure of a protein in this family.
We determined the crystal structure of MeTr from Clostridium thermoaceticum at 2.2 A resolution using multiwavelength anomalous diffraction methods. The overall architecture presents a new functional class of the versatile triose phosphate isomerase (TIM) barrel fold. The MeTr tertiary structure is surprisingly similar to the crystal structures of dihydropteroate synthetases despite sharing less than 20% sequence identity. This homology permitted the methyl-H4folate binding site to be modeled. The model suggests extensive conservation of the pterin ring binding residues in the polar active sites of the methyltransferases and dihydropteroate synthetases. The most significant structural difference between these enzymes is in a loop structure above the active site. It is quite open in MeTr, where it can be modeled as the cobalamin binding site.
The MeTr structure consists of a TIM barrel that embeds methyl-H4folate and cobamide. All related methyltransferases are predicted to fold into a similar TIM barrel pattern and have a similar pterin and cobamide binding site. The observed structure is consistent with either a 'front' (N5) or 'back' (C8a) side protonation of CH3-H4folate, a key step that enhances the electrophilic character of the methyl group, activating it for nucleophilic attack by Co(I).
甲基四氢叶酸、类咕啉铁硫蛋白甲基转移酶（MeTr）催化二氧化碳固定的伍德-Ljungdahl途径中的关键步骤。它将甲基四氢叶酸（CH3-H4folate）中的N5-甲基基团转移到另一种蛋白质类咕啉铁硫蛋白中的钴胺素（I）酰胺中心。MeTr是包括甲硫氨酸合成酶和激活甲基四氢甲（或-肌）蝶呤甲基基团的产甲烷酶在内的蛋白质家族的成员。我们报道了该家族中一种蛋白质的首个结构。
我们使用多波长反常衍射方法，以2.2埃的分辨率确定了嗜热醋梭菌中MeTr的晶体结构。整体结构呈现出一种多功能的磷酸丙糖异构酶（TIM）桶状折叠的新功能类别。尽管序列同一性低于20%，但MeTr的三级结构与二氢蝶酸合酶的晶体结构惊人地相似。这种同源性使得能够对甲基-H4folate结合位点进行建模。该模型表明，甲基转移酶和二氢蝶酸合酶的极性活性位点中蝶呤环结合残基具有广泛的保守性。这些酶之间最显著的结构差异在于活性位点上方的一个环结构。在MeTr中它相当开放，可被建模为钴胺素结合位点。
MeTr结构由一个嵌入甲基-H4folate和钴胺酰胺的TIM桶组成。预计所有相关的甲基转移酶都将折叠成类似的TIM桶模式，并具有相似的蝶呤和钴胺酰胺结合位点。观察到的结构与CH3-H4folate的“前”（N5）或“后”（C8a）侧质子化一致，这是增强甲基亲电特性的关键步骤，使其能够被Co（I）进行亲核攻击。
