Fig. 5: Transplant of loop reconstruction mutations to create the highly active PET hydrolases LCC-ICCG-NM and Kubu-P^M12-NM, and their experimental characterization under industrial conditions.

a Structure alignment between Bhr-PETase (PDB ID:7EOA, colored in purple) and LCC-ICCG (PDB ID:6THT, colored in orange). The catalytic residue serine (S165) was mutated to alanine in the crystal structure of LCC-ICCG. b Structure alignment between Bhr-PETase (PDB ID:7EOA, colored in purple) and Kubu-P^M12 (PDB ID:8YTY, colored in green). c, d Comparison of PET depolymerization by LCC-ICCG-NM and LCC-ICCG under high PET substrate concentrations in a bioreactor. The reaction system had a total volume of 50 ml, containing 16.5% (w/v) amorphized Pc-PET powder and 0.5 mg_enzyme g_PET⁻¹ (or 0.3 mg_enzyme g_PET⁻¹) in 100 mM phosphate buffer at pH 8.0. e, f Comparison of PET depolymerization by Kubu-P^M12-NM and Kubu-P^M12 under high PET substrate concentrations in a bioreactor. The reaction system had a total volume of 50 ml, containing 16.5% (w/v) amorphized Pc-PET powder and 0.5 mg_enzyme g_PET⁻¹ (or 0.3 mg_enzyme g_PET⁻¹) in 100 mM phosphate buffer at pH 8.0. The circles represent the conversion measured by the consumption of NaOH, while the crosses indicate the conversion calculated by HPLC analyses.