Human lysosomal alpha-mannosidase (hLAMAN) is a paradigmatic example of how few missense mutations can critically affect normal catabolism in the lysosome and cause the severe condition named alpha-mannosidosis. Here we have made use of computational chemistry methods to unveil the molecular basis of 4 missense mutations in hLAMAN with pathological consequences. We have simulated for the first time the all-atom catalytic reaction mechanism of hLAMAN by means of quantum mechanics/molecular mechanics metadynamics. Second, we show how the catalytically inactive variant D74E presents a significant increase of the free energy barrier. Third, we have identified that the D159N and E402K mutations are connected with the active site movement. Finally, we show that mutation R229W does not alter the balance between the hydrophilic and hydrophobic solvent accessible surface area in the protein, but does affect the active site dynamics. Our findings open up new opportunities for treatment of mannosidosis.
This PDF file contains the description of the computational methods for the modeling of the hLAMAN, MD simulations, exploration of the Cremer-Pople puckering parameters, QM/MM metadynamics simulations and free energy landscape calculations, and NEB method and QM/MM optimizations + supporting figures and tables.
QM/MM metadynamics and NEB-optimized structures for wild-type and D74E hLAMAN variants. The all-atom 3D-models for wild-type, D74E and R229W variants are included too.