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Combined quantum mechanics/molecular mechanics study on the reversible isomerization of glucose and fructose catalyzed by Pyrococcus furiosus phosphoglucose isomerase.

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.