Recent discovery of huge magnesium isotope effect in the rate of enzymatic synthesis of ATP offers a new insight on the mechanochemistry of enzymes as the molecular machines. The activity of magnesium-dependent enzymes (ATPase, phosphocreatine, phosphoglycerate kinases), in which Mg2+ ion has a magnetic isotope nucleus 25Mg, was found to be two or three times higher than that of enzymes in which the Mg2+ ion has nonmagnetic, spinless isotopic nuclei 24Mg or 26Mg. This isotope effect demonstrates unambiguously that the ATP synthesis is a spin-dependent process in which the Mg2+ ion is a reagent. It transforms mechanical energy of unequilibrium conformations of enzyme macromolecule into the energy of the P–O bond in ATP.