PT - JOURNAL ARTICLE AU - Cui, Yinglu AU - Chen, Yanchun AU - Liu, Xinyue AU - Dong, Saijun AU - Tian, Yu’e AU - Qiao, Yuxin AU - Han, Jing AU - Li, Chunli AU - Han, Xu AU - Liu, Weidong AU - Chen, Quan AU - Du, Wenbin AU - Tang, Shuangyan AU - Xiang, Hua AU - Liu, Haiyan AU - Wu, Bian TI - Computational redesign of PETase for plastic biodegradation by GRAPE strategy AID - 10.1101/787069 DP - 2019 Jan 01 TA - bioRxiv PG - 787069 4099 - http://biorxiv.org/content/early/2019/09/30/787069.short 4100 - http://biorxiv.org/content/early/2019/09/30/787069.full AB - The excessive use of plastics has been accompanied by severe ecologically-damaging effects. The recent discovery of PETase from Ideonalla sakaiensis that decomposes poly(ethylene terephthalate) (PET) under mild conditions provides an attractive avenue for biodegradation of plastics. However, the inherent instability of the enzyme limits its practical utilization. Here, we devised a novel computational strategy (Greedy Accumulated strategy for Protein Engineering, GRAPE). A systematic clustering analysis combined with greedy accumulation of the beneficial mutations in a computationally derived library enabled a redesigned variant, DuraPETase, with drastically elevated apparent melting temperature by 31 °C and strikingly enhanced degradation activity toward high-crystallinity PET film (49.2%) at moderate temperatures (product concentration raised from μM range to mM levels). The underlying mechanism of the robust promotion of enzyme performance has been demonstrated via crystal structure and molecular dynamics simulations. Collectively, this work promotes the methodological capabilities of computational enzyme design to circumvent antagonistic epistatic effects and provides a valuable tool for further understanding and advancing polyester hydrolysis in the natural environment.