0428同步年報-2021-全

Life Science 041 P olyethylene Terephthalate (PET) is an extensively produced plastic, with annual production approaching 70 metric tons. A huge amount of PET wastes that enter and accumulate in the ecosystem has caused a serious threat to the global environment. PET wastes are mainly incinerated or landfilled, but these methods result in secondary pollution. Developing environmentally compatible measures to eliminate PET wastes is thus an urgent task. PET is composed of ester bond-linked terephthalate (TPA) and ethandiol (EG), which are presumably susceptible to enzyme-mediated hydrolysis. Several types of esterases, including lipases, carboxylesterases and cutinases, have been reported to exhibit PET-hydrolytic activity, 1 but all these enzymes show poor PET hydrolytic efficacy because PET contains aromatic moieties and compact polymeric structures in a large ratio. 2 Cutinases are the most effective PET-hydrolytic enzymes known so far. Furthermore, only enzymes that can operate at a temperature above the glass-transition temperature of PET (60−70 o C), at which the crystalline structure of PET can be resolved, show notable hydrolytic activity. In 2016, a PET- assimilating bacterium named Ideonella sakaiensis isolated from a PET recycling factory revealed a naturally evolved PET degradation machinery. 3 This bacterium secretes a cutinase-like enzyme, denoted Is PETase, to hydrolyze PET into smaller compounds mono (2-hydroxyethyl) terephthalic acid (MHET) and TPA at ambient temperature (30−35 o C) ( Fig. 1(a) ). These small compounds are then transported into bacterial cells and metabolized. To uncover the mechanism of action of Is PETase, a research team led by Rey-Ting Guo (Hubei University, China) solved the crystal structure of Is PETase and its complex with substrate/product analogues and identified several unique features. 4 The X-ray diffraction data were collected at TLS 15A1 and TPS 05A of NSRRC. First, the enzyme has an extra disulfide bond in addition to the one that is strictly found in all known cutinases. This extra disulfide bond stabilizes a loop region, also unique to Is PETase, which constitutes the substrate- binding pocket ( Fig. 1(b) ). Depleting this extra disulfide bond leads to active site collapse and severely impacts the enzyme activity. 5 Distinct from canonical cutinases that are found to show PET hydrolytic activity, Is PETase exhibits a substrate preference towards PET over fatty acids (a component of cutin). This condition indicates that Is PETase features a substrate-binding pocket architecture that is Fig. 1 : Overall structure of PETase and comparison with Cutinase. (a) Structures of PETase hydrolysis products (boxed). (b) The PETase structure is presented as a cartoon model. The catalytic triad (red dashed-line circle) and disulfide bridges (red labels) are shown as sticks. (c) The crystal structures of Is PETase in complex with substrate analogue 1-(2-hydroxyethyl) 4-methyl TPA (left) and a PET-hydrolyzing cutinase from Thermobifida fusca ( Tf Cutinase) with the catalytic Ser-OH group converted to sulfonate ester-OS(O) 2 Bn (right) shown at the surface. Bound ligands in structures are shown as pink (PETase) and purple ( Tf Cut) sticks. [Reproduced from Ref. 2 and Ref. 4] Structural Studies Revealed the Evolution and Development of PETase Various plastic products made from polyethylene terephthalate (PET) are widely used in various industries. PET products cannot be decomposed by themselves and cause plastic pollution. A combination of structural and biochemical analysis shows how IsPETase has greater hydrolytic activity than other PETase.