Designed CYP450 Scaffold That Possesses a Local Electric\nField Biodegrades Polyethylene Terephthalate

In\nenzymes, protein residues themselves carry charges and produce\na preorganized local electric field (LEF). Such LEFs can be modified\nby strategically mutating the charged and polar residues to create\na designed LEF (D-LEF) to project on the reaction axis to modulate\nthe reactivity of the enzyme. We investigated the enzymatic degradation\nof polyethylene terephthalate (PET) using in silico engineering of\nCYP450 enzymes, which possess D-LEFs. We show that PET degradation\ncan, in principle, be catalyzed using a CYP450 scaffold through oxidative\ncleavage of the ester bond. The PET degradation occurs in two crucial\nsteps; the first step is the usual hydroxylation reaction initiated\nby compound I (Cpd I), while the second step is the dealkylation of\nthe <i>gem</i>-hydroxy moiety, which is driven by the preorganized\nLEF of the CYP450 enzyme. Using molecular-level analyses for three\ndifferent enzymes, we found that each of the three can efficiently\ncatalyze the HAT reaction. However, only CYP450GcoA performs an efficient\ndealkylation reaction since it possesses the only scaffold among the\nthree enzymes that has preorganized LEF properly oriented for dealkylation.\nWe show that a strategic mutation based on the designed LEF along\nthe reaction axis and the binding site architecture can evolve the\nenzyme for the PET degradation reaction. Our study further provides\na key lesson that intuiting the LEF of the enzyme in the direction\nof the reaction axis could be crucial in selecting the most suitable\nscaffold for the desired reaction.