Structural and functional analyses of microcystinase A: optimized heterologous expression, stability, and degradability

Siyu YANG; Jun ZUO; Licheng HUANG; Jin LIU; Nanqin GAN

doi:10.1007/s00343-024-4047-3

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Structural and functional analyses of microcystinase A: optimized heterologous expression, stability, and degradability

Special Section: Scientific control of cyanobacterial blooms and their secondary hazards | Updated：2024-12-24

- Structural and functional analyses of microcystinase A: optimized heterologous expression, stability, and degradability
- Journal of Oceanology and Limnology Vol. 42, Issue 6, Pages: 1805-1816(2024)
- Affiliations：
  
  1.Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan430072, China
  2.National and Local Joint Engineering Research Center of Ecological Treatment Technology for Urban Water Pollution, Zhejiang Provincial Key Lab for Water Environment and Marine Biological Resources Protection, Institute for Eco-Environmental Research of Sanyang Wetland, Wenzhou University, Wenzhou325035, China
  3.University of Chinese Academy of Sciences, Beijing100190, China
  4.Kunming Dianchi and Plateau Lakes Institute, Kunming650228, China
- Author bio：
  
  jzuo@wzu.edu.cn
  gannq@ihb.ac.cn
- Funds：
- DOI：10.1007/s00343-024-4047-3
  CLC：
- Received：17 February 2024，
  
  Published：01 November 2024
- Accepted：
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YANG Siyu,ZUO Jun,HUANG Licheng,et al.Structural and functional analyses of microcystinase A: optimized heterologous expression, stability, and degradability[J].Journal of Oceanology and Limnology,2024,42(06):1805-1816.
DOI：

YANG Siyu,ZUO Jun,HUANG Licheng,et al.Structural and functional analyses of microcystinase A: optimized heterologous expression, stability, and degradability[J].Journal of Oceanology and Limnology,2024,42(06):1805-1816. DOI： 10.1007/s00343-024-4047-3.

摘要

Abstract

Microcystinase (MlrA) is a key endopeptidase that catalyzes microcystin degradation without generating harmful byproduct. However

the application of MlrA in the field is primarily impeded by its limited productivity and short lifespan. Therefore

the MlrA’s function was studied by modelling its structure

which subsequently increased its heterologous expression and high-temperature stability. Results demonstrate that after the irregular sequence at the C-terminus of MlrA was removed

enzyme solubility was significantly decreased. In addition

three fusion tags

namely maltose-binding protein

glutathione S-transferase (GST)

and N-utilization substance A (NusA) were used to enhance the overexpression of soluble recombinant MlrA

among which NusA-MlrA exhibited the highest solubility. Moreover

NusA-MlrA was active in pH 4–10 at 20–80 °‍‍C; even at 80 °‍‍C

approximately 35.8% of fusion protein remained active. NusA-MlrA retained 89% of MlrA’s activity even after 7 d of storage at 50 °‍‍C; and on day 7

the protein retained

90% of its activity at pH 7. Finally

a stable

soluble

and long-lasting heterologous MlrA was successfully constructed th

at could eliminate microcystins in

Escherichia

coli

C43 (DE3). This study enriched the comprehension of MlrA’s structure and enzymatic properties

by particularly addressing the endopeptidase’s low expression and short lifespan

which improved its suitability for future applications.

关键词

Keywords

references

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