Establishing a Novel <i>E. coli</i> Heterologous Secretion Expression System Mediated by mScarlet3 for the Expression of a Novel Lipolytic Enzyme
Our previous study demonstrated that an <i>Escherichia coli</i> heterologous secretion expression system, mediated by superfolder green fluorescent protein (sfGFP) mutants, significantly enhances recombinant lipase yield and reduces large-scale production costs. In this study, we identif...
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2025-06-01
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| author | Jun Yang Mingjun Yang Huichen Liu Xinyu Liu Fei Wang Wenqiang Li Yang Liu Chao Zhai Lixin Ma |
| author_facet | Jun Yang Mingjun Yang Huichen Liu Xinyu Liu Fei Wang Wenqiang Li Yang Liu Chao Zhai Lixin Ma |
| author_sort | Jun Yang |
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| description | Our previous study demonstrated that an <i>Escherichia coli</i> heterologous secretion expression system, mediated by superfolder green fluorescent protein (sfGFP) mutants, significantly enhances recombinant lipase yield and reduces large-scale production costs. In this study, we identified mScarlet3, a fast-folding fluorescent protein, as another effective mediator of secretion expression in <i>E. coli</i>. A novel lipolytic enzyme, named LipHu6, was identified through sequence alignment. Secretion expression of LipHu6 was achieved by fusing mScarlet3 to either its N- or C-terminus. The specific activity of mScarlet3-LipHu6 reached 669,151.75 U/mmol, slightly surpassing that of LipHu6 alone (646,682.69 U/mmol) and markedly exceeding that of sfGFP<sub>(-15)</sub>-LipHu6 (492,432.39 U/mmol). Notably, N-terminal mScarlet3 fusion had no impact on LipHu6 hydrolytic activity toward short-chain <i>p</i>-nitrophenyl fatty acyl esters (C2–C8). In contrast, mScarlet3-LipHu6 exhibited approximately 1.5- and 1.7-fold increases in hydrolytic activity toward <i>p</i>-nitrophenyl palmitate (p-NPP, C16) and <i>p</i>-nitrophenyl stearate (p-NPS, C18), respectively. In conclusion, this study establishes a novel <i>E. coli</i> heterologous secretion expression system mediated by mScarlet3, offering a highly efficient and cost-effective strategy for the large-scale production of lipolytic enzymes. |
| format | Article |
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| spelling | doaj-art-9dd5c3062d5a48208634cb737da932e82025-08-20T02:24:42ZengMDPI AGBiomolecules2218-273X2025-06-0115684210.3390/biom15060842Establishing a Novel <i>E. coli</i> Heterologous Secretion Expression System Mediated by mScarlet3 for the Expression of a Novel Lipolytic EnzymeJun Yang0Mingjun Yang1Huichen Liu2Xinyu Liu3Fei Wang4Wenqiang Li5Yang Liu6Chao Zhai7Lixin Ma8State Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan 430062, ChinaState Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan 430062, ChinaState Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan 430062, ChinaState Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan 430062, ChinaState Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan 430062, ChinaState Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan 430062, ChinaState Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan 430062, ChinaState Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan 430062, ChinaState Key Laboratory of Biocatalysis and Enzyme Engineering, Hubei Key Laboratory of Industrial Biotechnology, School of Life Sciences, Hubei University, Wuhan 430062, ChinaOur previous study demonstrated that an <i>Escherichia coli</i> heterologous secretion expression system, mediated by superfolder green fluorescent protein (sfGFP) mutants, significantly enhances recombinant lipase yield and reduces large-scale production costs. In this study, we identified mScarlet3, a fast-folding fluorescent protein, as another effective mediator of secretion expression in <i>E. coli</i>. A novel lipolytic enzyme, named LipHu6, was identified through sequence alignment. Secretion expression of LipHu6 was achieved by fusing mScarlet3 to either its N- or C-terminus. The specific activity of mScarlet3-LipHu6 reached 669,151.75 U/mmol, slightly surpassing that of LipHu6 alone (646,682.69 U/mmol) and markedly exceeding that of sfGFP<sub>(-15)</sub>-LipHu6 (492,432.39 U/mmol). Notably, N-terminal mScarlet3 fusion had no impact on LipHu6 hydrolytic activity toward short-chain <i>p</i>-nitrophenyl fatty acyl esters (C2–C8). In contrast, mScarlet3-LipHu6 exhibited approximately 1.5- and 1.7-fold increases in hydrolytic activity toward <i>p</i>-nitrophenyl palmitate (p-NPP, C16) and <i>p</i>-nitrophenyl stearate (p-NPS, C18), respectively. In conclusion, this study establishes a novel <i>E. coli</i> heterologous secretion expression system mediated by mScarlet3, offering a highly efficient and cost-effective strategy for the large-scale production of lipolytic enzymes.https://www.mdpi.com/2218-273X/15/6/842lipolytic enzymemScarlet3<i>E. coli</i> secretion expression |
| spellingShingle | Jun Yang Mingjun Yang Huichen Liu Xinyu Liu Fei Wang Wenqiang Li Yang Liu Chao Zhai Lixin Ma Establishing a Novel <i>E. coli</i> Heterologous Secretion Expression System Mediated by mScarlet3 for the Expression of a Novel Lipolytic Enzyme Biomolecules lipolytic enzyme mScarlet3 <i>E. coli</i> secretion expression |
| title | Establishing a Novel <i>E. coli</i> Heterologous Secretion Expression System Mediated by mScarlet3 for the Expression of a Novel Lipolytic Enzyme |
| title_full | Establishing a Novel <i>E. coli</i> Heterologous Secretion Expression System Mediated by mScarlet3 for the Expression of a Novel Lipolytic Enzyme |
| title_fullStr | Establishing a Novel <i>E. coli</i> Heterologous Secretion Expression System Mediated by mScarlet3 for the Expression of a Novel Lipolytic Enzyme |
| title_full_unstemmed | Establishing a Novel <i>E. coli</i> Heterologous Secretion Expression System Mediated by mScarlet3 for the Expression of a Novel Lipolytic Enzyme |
| title_short | Establishing a Novel <i>E. coli</i> Heterologous Secretion Expression System Mediated by mScarlet3 for the Expression of a Novel Lipolytic Enzyme |
| title_sort | establishing a novel i e coli i heterologous secretion expression system mediated by mscarlet3 for the expression of a novel lipolytic enzyme |
| topic | lipolytic enzyme mScarlet3 <i>E. coli</i> secretion expression |
| url | https://www.mdpi.com/2218-273X/15/6/842 |
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