Yin_2022_Bioresour.Technol_364_128026

The serious environmental pollution that came up with the continuously growing demand for polyethylene terephthalate (PET) has attracted global concern. The IsPETase which has shown the highest PET degradation activity under ambient temperature is a promising enzyme for PET biodegradation, while poor thermostability limited its practical application. Herein, an electrostatic interaction-based strategy was applied for rational design of IsPETase towards enhanced thermostability. The IsPETase(I139R) variant displayed the highest T(m) value of 56.4s degreesC and 3.6-times higher PET degradation activity. Molecular simulations demonstrated that the introduction of salt bridges stabilized the local structures, resulting in robust thermostability. Meanwhile, the IsPETase(S92K/D157E/R251A) not only exhibited higher thermostability but also showed a 1.74-fold k(cat) increase towards mono-(2-hydroxyethyl) terephthalate, which ultimately achieved PET depolymerization to complete monomer TPA. Collectively, the electrostatic interaction-based strategy, together with the derived IsPETase variants, could help promote the bio-recycle of PET, reducing the severe global burden of PET waste.