Combined quantum mechanics/molecular mechanics study on the reversible isomerization of glucose and fructose catalyzed by Pyrococcus furiosus phosphoglucose isomerase.
Combined quantum mechanics/molecular mechanics study on the reversible isomerization of glucose and fructose catalyzed by Pyrococcus furiosus phosphoglucose isomerase.
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DOI：
10.1021/ja710633c
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发表时间：
2008-05
影响因子：
15
通讯作者：
Ruibo Wu;Hujun Xie;Z. Cao;Y. Mo
中科院分区：
文献类型：
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作者：
Ruibo Wu;Hujun Xie;Z. Cao;Y. Mo
磷酸葡萄糖异构酶 (PGI) 催化葡萄糖 6-磷酸 (G6P) 和果糖 6-磷酸 (F6P) 的可逆相互转化，由两个进化上不同的蛋白质家族代表。一种是真细菌、真核生物和少数古细菌中的常规类型，其中活性位点不含金属离子，反应通过顺式烯二醇中间机制进行。第二种是最近在广古菌中发现的，属于金属酶，对于催化异构化是通过顺式烯二醇中间体机制还是通过氢化物转移机制发生存在争议。我们通过 QM(B3LYP)/MM 单点优化和 QM(PM3)/MM 分子动力学模拟研究了含金属激烈热球菌 PGI 催化的开链形式 G6P 和 F6P 的可逆相互转化。提出了一种基于两性离子中间体的机制，涉及质子和氢化物转移。该机制中关键两性离子中间体的存在可以有效地调和有争议的机制并使酶促反应合理化。计算表明，整个异构化过程在动力学和热力学上都非常容易。基于对其丙氨酸突变体的计算，已经阐明了保守残基的关键作用。特别是，Tyr152 通过氢化物转移机制推动 H1 转移，并主导氢转移的立体化学选择性。其余的保守残基基本上将底物维持在近攻击反应构象并介导质子转移。尽管Zn(2+)不直接参与反应，但金属离子作为结构锚构建了氢键线，将基底与外部区域连接起来，为基底和溶剂之间的氢交换提供了潜在的通道。
Phosphoglucose isomerase (PGI), which catalyzes the reversible interconversion of glucose 6-phosphate (G6P) and fructose 6-phosphate (F6P), is represented by two evolutionarily distinct protein families. One is a conventional type in eubacteria, eukaryotes, and a few archaea, where the active sites contain no metal ions and reactions proceed via the cis-enediol intermediate mechanism. The second type, found recently in euryarchaeota species, belongs to metalloenzymes, and controversies exist over whether the catalyzed isomerization occurs via the cis-enediol intermediate mechanism or a hydride shift mechanism. We studied the reversible interconversion of the open-chain form G6P and F6P catalyzed by the metal-containing Pyrococcus furiosus PGI by performing QM(B3LYP)/MM single-point optimizations and QM(PM3)/MM molecular dynamics simulations. A zwitterion intermediate-based mechanism, which involves both proton and hydride transfers, has been put forward. The presence of the key zwitterionic intermediate in this mechanism can effectively reconcile the controversial mechanisms and rationalize the enzymatic reaction. Computations show that the overall isomerization process is quite facile, both dynamically and thermodynamically. The crucial roles of conserved residues have been elucidated on the basis of computations on their alanine mutants. In particular, Tyr152 pushes the H1 transfer through a hydride-shift mechanism and dominates the stereochemistry selectivity of the hydrogen transfer. The rest of the conserved residues basically maintain the substrate in the near-attack reactive conformation and mediate the proton transfer. Although Zn(2+) is not directly involved in the reaction, the metal ion as a structural anchor constructs a hydrogen bond wire to connect the substrate to the outer region, providing a potential channel for hydrogen exchange between the substrate and solvent.