The effect of ionic strength on PETase enzymes: An experimental and computational study.

Over recent decades, various enzymes capable of breaking down polyethylene terephthalate (PET) have emerged as sustainable tools for plastic waste management. Among them, IsPETase from Ideonella sakaiensis 201-f6 stands out for its high catalytic activity at low temperatures. However, the discovery of the PETase-like enzyme from the marine sponge Streptomyces sp. SM14 (PETaseSM14) has introduced a new class of biocatalysts active at high-salt concentrations, whose structural and catalytic properties remain poorly understood. This study explores the structural and catalytic behavior of both IsPETase and PETaseSM14 under varying ionic strength (from 150 to 900 mM of NaCl concentration) using all-atom molecular dynamics simulations and in vitro assays. Results reveal that the flexible, enlarged binding site of IsPETase improves substrate accommodation but also causes catalytic residue displacement and rapid deactivation, particularly under high-salt conditions. In contrast, PETaseSM14 has a smaller, more rigid binding pocket that undergoes moderate widening upon salt concentration increasing, thus promoting water and substrate recruitment. Additionally, active forms of both enzymes bind PET chains in conformations similar to those found in amorphous PET. These findings offer key structural insights that can inform future enzyme engineering efforts for effective PET degradation tailored to diverse environmental conditions.