The reaction mechanism of UDP-GlcNAc 5,6-dehydratase: a quantum mechanical/molecular mechanical (QM/MM) study

Abstract

UDP-GlcNAc 5,6-dehydratase (TunA), a particular member of the short-chain dehydrogenase/reductase family, catalyzes the bioconversion of UDP-GlcNAc to UDP-6-deoxy-GlcNAc-5,6-ene. To elucidate the detailed mechanism of TunA, combined quantum mechanical/molecular mechanical calculations have been performed. The calculation results reveal that the TunA-catalyzed process follows a three-step sequential mechanism, including oxidation, dehydration and reduction. The oxidation step is calculated to be rate limiting and undergoes a concerted but asynchronous mechanism with an energy barrier of 21.6 kcal/mol. The dehydration step proceeds via the E1cB mechanism involving proton transfer and departure of hydroxyl group. Our calculation results show that the key residue Glu121 is ideally situated in the active site and serves as the general base and then as acid to take part in the elimination of water. The residue Cys120 is confirmed to play an important role in maintaining the active pocket arrangement. As the reverse process of the oxidation reaction, the reduction step follows the same concerted asynchronous mechanism, but the hydride transfer is prior to proton transfer.