Efficient production of L-asparaginase in <i>Bacillus licheniformis</i> by optimizing expression elements and host.

Xinyuan Yang; Yi Rao; Mengxi Zhang; Jiaqi Wang; Wenyuan Liu; Dongbo Cai; Shouwen Chen

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Efficient production of L-asparaginase in Bacillus licheniformis by optimizing expression elements and host.

Author: Xinyuan YANG ¹ ; Yi RAO ¹ ; Mengxi ZHANG ¹ ; Jiaqi WANG ¹ ; Wenyuan LIU ¹ ; Dongbo CAI ¹ ; Shouwen CHEN ¹
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1. State Key Laboratory of Biocatalysis and Enzyme Engineering, Environmental Microbial Technology Center of Hubei Province, College of Life Sciences, Hubei University, Wuhan 430062, Hubei, China.
Publication Type:Journal Article
Keywords: Bacillus licheniformis; L-asparaginase; expression host; promoter; signal peptide
MeSH: Bacillus licheniformis/metabolism*; Asparaginase/genetics*; Bacillus/genetics*; Protein Sorting Signals; Promoter Regions, Genetic/genetics*; Bacillus subtilis/genetics*; Bacterial Proteins
From: Chinese Journal of Biotechnology 2023;39(3):1096-1106
CountryChina
Language:Chinese
Abstract: L-asparaginase (L-ASN) is widely applied in the treatment of malignant tumor and low-acrylamide food production, however, the low expression level hampers its application. Heterologous expression is an effective strategy to increase the expression level of target enzymes, and Bacillus is generally used as the host for efficient production of enzymes. In this study, the expression level of L-asparaginase in Bacillus was enhanced through optimization of expression element and host. Firstly, five signal peptides (SP_SacC, SP_AmyL, SP_AprE, SP_YwbN and SP_WapA) were screened, among which SP_SacC showed the best performance, reaching an activity of 157.61 U/mL. Subsequently, four strong promoters (P43, P_ykzA-P43, P_Ubay and P_bacA) from Bacillus were screened, and tandem promoter P_ykzA-P43 showed the highest yield of L-asparaginase, which was 52.94% higher than that of control strain. Finally, three Bacillus expression hosts (B. licheniformis Δ0F3 and BL10, B. subtilis WB800) were investigated, and the maximum L-asparaginase activity, 438.3 U/mL, was reached by B. licheniformis BL10, which was an 81.83% increase compared with that of the control. This is also the highest level of L-asparaginase in shake flask reported to date. Taken together, this study constructed a B. licheniformis strain BL10/P_ykzA-P43-SP_SacC-ansZ capable of efficiently producing L-asparaginase, which laid the foundation for industrial production of L-asparaginase.