嗜热丁烷氧化古菌Candidatus Syntrophoarchaeum烷基转移酶MtaA催化丁基辅酶M的分子机制研究

Molecular mechanism of the alkyl-transferase MtaA catalyzing butyl-CoM in the thermophilic butane-oxidizing archaea Candidatus Syntrophoarchaeum

摘要:

[背景] 嗜热古菌Candidatus Syntrophoarchaeum可以与硫酸盐还原细菌共生，通过逆转产甲烷途径进行正丁烷的氧化，但在该过程中负责催化丁基辅酶M氧化的酶尚未确定。[目的] 利用分子动力学模拟证明Ca.Syntrophoarchaeum中mtaA基因编码的蛋白可以特异性催化丁基辅酶M中丁基的转移，并非转移甲基。[方法] 使用Methanosarcina mazei辅酶M甲基转移酶MtaA的晶体结构（PDB ID：4ay8）作为模板，对MtaA_1（GenBank登录号OFV65993.1）和MtaA_2（GenBank登录号OFV65678.1）进行同源建模。使用分子对接得到两者分别结合CH₃-CoM和C₄H₉-CoM时的结构，并用AMBER18进行分子动力学模拟。[结果] 当MtaA_1和MtaA_2分别结合C₄H₉-CoM时，表现出与4ay8晶体结构类似的TIM-Barrel折叠三维结构，但在活性中心形状、Zn²⁺与底物距离以及活性位点附近氨基酸配位方式等方面存在差异，这可能是导致Ca.Syntrophoarchaeum中mtaA基因编码的蛋白催化丁基辅酶M氧化的原因。其中MtaA_2与4ay8结构更相似，活性中心氨基酸配位更完整，暗示其更可能具备催化活性。然而当MtaA_1和MtaA_2分别结合CH₃-CoM时，整体结构不合实际，活性中心Zn²⁺与底物距离过远，表明底物几乎不可能与酶结合。[结论] Ca.Syntrophoarchaeum中的MtaA_1和MtaA_2很可能是特异性的丁基转移酶，而非催化甲基的转移，其中MtaA_2具备活性的可能性更高。

Abstract:

[Background] Thermophilic archaea Candidatus Syntrophoarchaeum was found to coexist with sulfate-reducing bacteria and oxidize n-butane by the reverse methanogenesis pathway. However, in this process, the enzyme responsible for catalyzing the oxidation of butyl-CoM has not been determined yet. [Objective] Using molecular dynamics simulation to prove that the enzymes encoded by mtaA genes found in Ca. Syntrophoarchaeum can specifically catalyze the transfer of butyl in butyl-CoM, and they are not methyltransferases. [Methods] Using the crystal structure of Methanosarcina mazei coenzyme M methyltransferase MtaA (PDB ID:4ay8) as a template, homology modeling of MtaA_1 (GenBank ID:OFV65993.1) and MtaA_2 (GenBank ID:OFV65678.1) was performed. Molecular docking was used to obtain the structure when they were combined with CH₃-CoM and C₄H₉-CoM respectively, and AMBER18 was used for molecular dynamics simulation. [Results] When combined with C₄H₉-CoM, MtaA_1 and MtaA_2 exhibited a TIM-barrel-like three-dimensional structure similar to the fold of 4ay8. However, there are differences in the shape of the active site, the distance between Zn²⁺ and the substrate, and the coordination of amino acid residues around the active site. These differences may be the reason why the enzymes encoded by mtaA genes found in Ca. Syntrophoarchaeum catalyze the oxidation of butyl-CoM. The overall structure of MtaA_2 is more similar to 4ay8, and the coordination of residues around the active site is complete, suggesting that MtaA_2 is more likely to be active. When MtaA_1 and MtaA_2 bind to CH₃-CoM respectively, their overall structures are unrealistic, and the coordinated Zn²⁺ is too far away from the substrate, indicating that CH₃-CoM is almost impossible to bind to the enzyme. [Conclusion] MtaA_1 and MtaA_2 of Ca. Syntrophoarchaeum are likely to be specific butyl-transferases rather than catalyzing the transfer of methyl groups, and MtaA_2 is more likely to be active.