Broken-Symmetry DFT Computations for the ReactionPathway of IspH an Iron–Sulfur Enzyme in Pathogenic Bacteria

摘要

The recently discovered methylerythritol phosphate (MEP) pathway provides new targets for the development of antibacterial and antimalarial drugs. In the final step of the MEP pathway, the [4Fe–4S] IspH protein catalyzes the 2e^–/2H⁺ reductive dehydroxylation of (E)-4-hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP) to afford the isoprenoid precursors isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP). Recent experiments have attempted to elucidate the IspH catalytic mechanism to drive inhibitor development. Two competing mechanisms have recently emerged, differentiated by their proposed HMBPP binding modes upon 1e^– reduction of the [4Fe–4S] cluster: (1) a Birch reduction mechanism, in which HMBPP remains bound to the [4Fe–4S] cluster through its terminal C4–OH group (ROH-bound) until the −OH is cleaved as water; and (2) an organometallic mechanism, in which the C4–OH group rotates away from the [4Fe–4S] cluster, allowing the HMBPP olefin group to form a metallacycle complex with the apicaliron (η²-bound). We perform broken-symmetry densityfunctional theory computations to assess the energies and reductionpotentials associated with the ROH- and η²-boundstates implicated by these competing mechanisms. Reduction potentialsobtained for ROH-bound states are more negative (−1.4 to −1.0V) than what is typically expected of [4Fe–4S] ferredoxin proteins.Instead, we find that η²-bound states are lower inenergy than ROH-bound states when the [4Fe–4S] cluster is 1e^– reduced. Furthermore, η²-bound states can already be generated in the oxidized state, yieldingreduction potentials of ca. −700 mV when electron additionoccurs after rotation of the HMBPP C4–OH group.We demonstrate that such η²-bound states are kineticallyaccessible both when the IspH [4Fe–4S] cluster is oxidizedand 1e^– reduced. The energeticallypreferred pathway gives 1e^– reductionof the cluster after substrate conformational change, generating the1e^– reduced intermediate proposedin the organometallic mechanism.