Processes, Vol. 13, Pages 3320: Engineered Mors1 Enzyme from the Antarctic Bacterium Moraxella TA144 for Enhanced Thermal Stability and Activity for Polyethylene Terephthalate Degradation

Processes doi: 10.3390/pr13103320

Authors:
Satyam Satyam
Sanjukta Patra

Plastic pollution, particularly from polyethylene terephthalate (PET), poses significant environmental concerns due to ecosystem persistence and extensive packaging use. Conventional recycling methods face inefficiencies, high costs, and limited scalability, necessitating sustainable alternatives. Biodegradation via PET hydrolases offers promising eco-friendly solutions, although most natural PET-degrading enzymes are thermophilic and require energy-intensive high temperatures. In contrast, psychrophilic enzymes function efficiently at extremely low temperatures but often lack stability under moderate conditions. Therefore, this study aimed to enhance the ability of the Mors1 enzyme from Moraxella TA144 to operate effectively under mesophilic conditions, which is closer to the optimal conditions for environmental application. Three strategic hydrophobic substitutions (K93I, E221I, and R235F) were introduced in loop regions, generating the mutant variant Mors1MUT. Comparative characterization revealed that Mors1MUT retained 98% of its activity at pH 9 and displayed greater resilience across both acidic and alkaline conditions than did the wild-type enzyme. Thermal stability assays revealed that Mors1MUT preserved 61% of its activity at 40 °C and 14% at 50 °C, whereas the wild-type enzyme was fully inactivated at these temperatures. The enzymatic hydrolysis of PET films significantly improved with Mors1MUT. Gravimetric analysis revealed weight losses of 0.83% for Mors1WT and 3.46% for Mors1MUT after a 12-day incubation period. This corresponds to a 4.16-fold increase in hydrolysis efficiency, confirming the enhanced catalytic performance of the mutant variant. The improvement was further validated by scanning electron microscopy (SEM), atomic force microscopy (AFM), and attenuated total reflectance–Fourier transform infrared (ATR-FTIR) analysis. Optimization of the reaction parameters through response surface methodology (enzyme load, time, pH, temperature, and agitation) confirmed increased PET hydrolysis under mild mesophilic conditions. These findings establish Mors1MUT as a robust mesophilic PETase with enhanced catalytic efficiency and thermal stability, representing a promising candidate for sustainable PET degradation under environmentally relevant conditions.

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