Synergistic precipitant powered self-assembly of papain for cross-linked enzyme crystals preparation--颗粒学报PARTICUOLOGY

Hao, Q., Guo, P., Wang, H., Su, M., Li, C., Hu, Y., & Su, W. (2024). Synergistic precipitant powered self-assembly of papain for cross-linked enzyme crystals preparation. Particuology, 85, 102-112. https://doi.org/10.1016/j.partic.2023.03.023

Synergistic precipitant powered self-assembly of papain for cross-linked enzyme crystals preparation

Qi Hao ^{a b 1}, Pan Guo ^{a b 1}, Honghai Wang ^{a b}, Min Su ^a, Chunli Li ^{a b}, Yuqi Hu ^{a b}, Weiyi Su ^{a b *}

a School of Chemical Engineering and Technology, Hebei University of Technology, Tianjin, 300130, China
b National-Local Joint Engineering Laboratory for Energy Conservation of Chemical Process Integration and Resources Utilization, Tianjin, 300130, China

10.1016/j.partic.2023.03.023

Volume 85, February 2024, Pages 102-112

Received 14 November 2022, Revised 13 March 2023, Accepted 29 March 2023, Available online 17 April 2023, Version of Record 25 April 2023.

E-mail: suweiyi@hebut.edu.cn

Highlights

• Papain crystals was obtained by the synergistic precipitate.

• l-cysteine increased the enzyme activity by affecting the conformation of papain.

• Three types of forces facilitated the binding between papain and l-cysteine.

• Papain CLECs were successful prepared with higher stability and wider application.

Abstract

In this study, we prepared cross-linked enzyme crystals (CLECs) of papain to further broaden the application of the enzyme with high activity in extreme environments. Initially, papain crystals were successfully obtained based on the micro-batch, batch, and expanded batch crystallization experiments. Specifically, ammonium sulfate and polyethylene glycol 6000 (PEG6000) were synergistically used as the precipitants, while l-cysteine was applied to enhance the activity of papain. Furthermore, the interaction between l-cysteine and papain was modeled by molecular docking technique. It was found that l-cysteine could form a hydrogen bond with aspartic acid residue (Asp) at site 158, and the electrostatic attraction with lysine residue (Lys) at site 156 was also quite obvious. Then the enzyme crystals were cross-linked by glutaraldehyde at optimized conditions. The papain CLECs were identified by various methods, and it was found that the thermal stability and enzymatic activity both increased compared to the raw enzyme. More importantly, it could be applied at more rigorous conditions, for example, pH of 4.

Graphical abstract

Keywords

Papain; Cross-linked enzyme crystals; l-cysteine; Molecular docking; Crystallization

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