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. 2021 Aug;43(8):1625-1635.
doi: 10.1007/s10529-021-03139-5. Epub 2021 May 16.

A temperature-induced chitosanase bacterial cell-surface display system for the efficient production of chitooligosaccharides

Qianqian Li  1 , Tuantuan Wang  1 , Yangzhi Ye  1 , Shimin Guan  1 , Baoguo Cai  1 , Shuo Zhang  1 , Shaofeng Rong  2
Affiliations

A temperature-induced chitosanase bacterial cell-surface display system for the efficient production of chitooligosaccharides

Qianqian Li et al. Biotechnol Lett. 2021 Aug.

Abstract

Objective: To establish a temperature-induced chitosanase bacterial cell-surface display system to produce chitooligosaccharides (COSs) efficiently for industrial applications.

Results: Temperature-inducible chitosanase CSN46A bacterial surface display systems containing one or two copies of ice nucleation protein (InaQ-N) as anchoring motifs were successfully constructed on the basis of Escherichia coli and named as InaQ-N-CSN46A (1 copy) and 2InaQ-N-CSN46A (2 copies). The specific enzyme activity of 2InaQ-N-CSN46A reached 761.34 ± 0.78 U/g cell dry weight, which was 45.6% higher than that of InaQ-N-CSN46A. However, few proteins were detected in the 2InaQ-N-CSN46A hydrolysis system. Therefore, 2InaQ-N-CSN46A had higher hydrolysis efficiency and stability than InaQ-N-CSN46A. Gel permeation chromatography revealed that under the optimum enzymatic hydrolysis temperature, the final products were mainly chitobiose and chitotriose. Chitopentaose accumulated (77.62%) when the hydrolysis temperature reached 60 °C. FTIR and NMR analysis demonstrated that the structures of the two hydrolysis products were consistent with those of COSs.

Conclusions: In this study, chitosanase was expressed on the surfaces of E. coli by increasing the induction temperature, and chitosan was hydrolysed directly without enzyme purification steps. This study provides a novel strategy for industrial COS production.

Keywords: Cell-surface display; Chitooligosaccharides; Chitosanases; Escherichia coli; Temperature-inducible.

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