Secretory expression in Bacillus subtilis and biochemical characterization of a highly thermostable polyethylene terephthalate hydrolase from bacterium HR29

• Published in: Enzyme and microbial technology Vol. 143; p. 109715
• Main Authors: Xi, Xingxiang, Ni, Kefeng, Hao, Helong, Shang, Yuepeng, Zhao, Bo, Qian, Zhen
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.02.2021
• Subjects: Bacillus subtilis; bacteria; Biochemical characterization; bioremediation; composts; cutinase; enzymatic treatment; PET degradation; PET hydrolyzing enzyme; polyethylene terephthalates; Secretory expression; sequence analysis; temperature; thermal stability; Biochemical characterization; PET degradation; PET hydrolyzing enzyme; Bacillus subtilis; Secretory expression
• ISSN: 0141-0229; 1879-0909; 1879-0909
• DOI: 10.1016/j.enzmictec.2020.109715

•High level extracellular expression of bacterial PET hydrolyzing enzymes in Bacillus subtilis.•Boosted expression titer by employing the chaperone-overexpression mutant strain.•First report on biochemical properties of BhrPETase, a highly active and super thermostable PET hydrolyzing enzyme. The environmental threat posed by disposal of plastic wastes has drawn extensive attention in recent years wherein polyethylene terephthalate (PET) constitutes one of the major plastic materials in the wastes. Recycling of PET wastes into reusable materials effectively overcomes its accumulation in the environment and can be achieved by mechanical, chemical, and biological processes. In comparison to the other methods, enzymatic treatment utilizing PET hydrolyzing enzymes (PETases) is environmental-friendly which avoids the use of hazardous chemicals. In this study, we report on the secretory expression in Bacillus subtilis a PETase (BhrPETase) from the bacterium HR29, a close homologue of the leaf-branch compost cutinase (LCC) with 94 % sequence identity. The expression titer of BhrPETase reached 0.66 g/L in an engineered chaperone-overexpression Bacillus subtilis strain, and the biochemical characterization of BhrPETase for the first time revealed its high hydrolyzing activity towards amorphous PET in comparison to two reported PET hydrolyzing enzymes LCC and IsPETase, which were expressed under the same expression conditions in Bacillus subtilis in our study. Most intriguingly, purified BhrPETase displayed a melting temperature as high as 101 °C. To our knowledge it is the most thermostable bacterial PETase characterized so far. The superior activity and thermostability of BhrPETase rendered it one of the most promising PETases for plastic waste recycling and bioremediation applications in the future.