TY - JOUR
T1 - The reaction mechanism of the Ideonella sakaiensis PETase enzyme
AU - Burgin, Tucker
AU - Pollard, Benjamin C.
AU - Knott, Brandon C.
AU - Mayes, Heather, B.
AU - Crowley, Michael F.
AU - McGeehan, John
AU - Beckham, Gregg T.
AU - Woodcock, H. Lee
PY - 2024/3/27
Y1 - 2024/3/27
N2 - Polyethylene terephthalate (PET), the most abundantly produced polyester plastic, can be depolymerized by the Ideonella sakaiensis PETase enzyme. Based on multiple PETase crystal structures, the reaction has been proposed to proceed via a two-step serine hydrolase mechanism mediated by a serine-histidine-aspartate catalytic triad. However, the details and rate-limiting step remained heretofore unknown, with speculation regarding the role of ancillary residues along the catalytic itinerary. To elucidate the multi-step PETase catalytic mechanism, we use path sampling and likelihood maximization to identify optimal reaction coordinates. We predict that acylation is rate-limiting and the reaction coordinates for both steps include elements describing nucleophilic attack, ester cleavage, and the "moving-histidine" mechanism. We find the flexibility of Trp185 is critical to the reaction, providing an explanation for decreased activity observed in mutations restricting Trp185 motion. Overall, this study reveals the detailed reaction mechanism necessary towards further engineering an important class of enzymes for plastics bioconversion.
AB - Polyethylene terephthalate (PET), the most abundantly produced polyester plastic, can be depolymerized by the Ideonella sakaiensis PETase enzyme. Based on multiple PETase crystal structures, the reaction has been proposed to proceed via a two-step serine hydrolase mechanism mediated by a serine-histidine-aspartate catalytic triad. However, the details and rate-limiting step remained heretofore unknown, with speculation regarding the role of ancillary residues along the catalytic itinerary. To elucidate the multi-step PETase catalytic mechanism, we use path sampling and likelihood maximization to identify optimal reaction coordinates. We predict that acylation is rate-limiting and the reaction coordinates for both steps include elements describing nucleophilic attack, ester cleavage, and the "moving-histidine" mechanism. We find the flexibility of Trp185 is critical to the reaction, providing an explanation for decreased activity observed in mutations restricting Trp185 motion. Overall, this study reveals the detailed reaction mechanism necessary towards further engineering an important class of enzymes for plastics bioconversion.
U2 - 10.1038/s42004-024-01154-x
DO - 10.1038/s42004-024-01154-x
M3 - Article
SN - 2399-3669
VL - 7
JO - Communications Chemistry
JF - Communications Chemistry
M1 - 65
ER -