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.