Molecular modelling of xylose isomerase catalysis: the role of electrostatics and charge transfer to metals

The two main steps of the mechanism of xylose-xylulose conversioncatalysed by D-xylose isomerase, the ring opening of xyloseand the isomerization of the opened product by hydride transfer,were investigated by molecular mechanical and molecular orbitaltechniques. The activation energies calculated for these reactionsclearly showed that hydrogen transfer is the rate-determiningstep of the enzymatic isomerization and that Mg²⁺ ions activatewhereas Zn²⁺ ions inhibit the reaction, in agreement with theexperiments. The remarkable differences between the net chargesof these ions found by molecular orbital calculations and theinspection of the protein electrostatic potential around thereaction intermediates indicate that the main role of bivalentmetal ions should be the electrostatic stabilization of thesubstrate transition states. In order to propose a more detailedmechanism, an attempt was made to clarify the effects of nearbyresidues (e.g. His54, Asp57, Lysl83, Asp257) in the reaction.Different isomerization mechanisms, such as through an enediolintermediate, were examined and could be excluded, in additionto the charge-relay mechanism during the ring opening.