Theoretical analysis of the catalytic mechanism of Helicobacter pylori glutamate racemase.

One of the most challenging open key questions behind the stereoinversion of D-glutamate and L-glutamate catalyzed by glutamate racemases is how those enzymes manage to generate the thermodynamically unfavorable reverse protonation state of the catalytic residue cysteine required for the proton abstraction from the α-carbon of glutamate. In this paper, we have used molecular dynamics (MD) simulations with a molecular mechanics force field along with QM/MM calculations starting from the crystal structure and from different MD snapshots to study the enantiomeric conversion of D-glutamate to L-glutamate catalyzed by the Helicobacter pylori glutamate racemase. Our results show that structural fluctuations of the enzyme-substrate complex, represented by the different snapshots, lead to reaction paths with different features and fates. The whole reaction, when it occurs, involves four successive proton transfers in three or four different steps. In the first step, Asp7 assists the deprotonation of D-glutamate by participating in general base catalysis with neutral Cys70 thiol. An analogous mechanism was previously found by some of us for the case of Bacillus subtilis glutamate racemase. This fact explains why that aspartate belongs to the group of strictly conserved residues.