L-Arabinose Binding, Isomerization, and Epimerization by D-Xylose Isomerase: X-Ray/Neutron Crystallographic and Molecular Simulation Study

D-xylose isomerase (XI) is capable of sugar isomerization and slow conversion of some monosaccharides into their C2-epimers. We present X-ray and neutron crystallographic studies to locate H and D atoms during the respective isomerization and epimerization of L-arabinose to L-ribulose and L-ribose, respectively. Neutron structures in complex with cyclic and linear L-arabinose have demonstrated that the mechanism of ring-opening is the same as for the reaction with D-xylose. Structural evidence and QM/MM calculations show that in the reactive Michaelis complex L-arabinose is distorted to the high-energy 5S1 conformation; this may explain the apparent high KM for this sugar. MD-FEP simulations indicate that amino acid substitutions in a hydrophobic pocket near C5 of L-arabinose can enhance sugar binding. L-ribulose and L-ribose were found in furanose forms when bound to XI. We propose that these complexes containing Ni2+ cofactors are Michaelis-like and the isomerization between these two sugars proceeds via a cis-ene-diol mechanism. [Display omitted] •Joint X-ray/neutron structures of D-xylose isomerase with L-arabinose•Structures and QM/MM reveal L-arabinose conformation is 5S1 in the active site•MD simulations suggest mutations to improve enzyme affinity to L-arabinose•Isomerization between L-ribulose and L-ribose may proceed by cis-ene-diol pathway Using X-ray/neutron crystallography and QM/MM, Langan et al. show that L-arabinose has a high-energy 5S1 conformation in complex with reactive D-xylose isomerase. They propose that L-arabinose isomerizes through hydride shift, whereas isomerization between L-ribulose and L-ribose may involve a cis-ene-diol.