Optimization Expression and Enzymatic Properties Research of Mannanase in Aspergillus niger
To achieve higher expression levels and enhanced enzymatic properties of mannanase, this study employed homologous recombination and site-specific integration technology to construct a recombinant mannanase Aspergillus niger strain. Subsequently, the expression characteristics and enzymatic properti...
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The editorial department of Science and Technology of Food Industry
2025-08-01
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| Series: | Shipin gongye ke-ji |
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| Online Access: | http://www.spgykj.com/cn/article/doi/10.13386/j.issn1002-0306.2024080210 |
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| author | Hui ZHANG Zhu LI Jiaxin MA Junbo FAN Zhiwei SHAO Yujun JIANG Jie LI |
| author_facet | Hui ZHANG Zhu LI Jiaxin MA Junbo FAN Zhiwei SHAO Yujun JIANG Jie LI |
| author_sort | Hui ZHANG |
| collection | DOAJ |
| description | To achieve higher expression levels and enhanced enzymatic properties of mannanase, this study employed homologous recombination and site-specific integration technology to construct a recombinant mannanase Aspergillus niger strain. Subsequently, the expression characteristics and enzymatic properties of the mannanase were systematically analyzed. This study successfully obtained the pure mannanase transformant MG of Aspergillus niger, which underwent homologous recombination at the glaA locus. Additionally, the pure mannanase transformant MD of Aspergillus niger was selected that not only exhibited homologous recombination at the glaA locus but also overexpressed the endoplasmic reticulum co-chaperone DnaJ1 gene at the asAA locus. Both strains MG and MD exhibited a distinct mannanase Man protein band at approximately 37 kDa, with the highest enzyme activities recorded at 6909.861 U·mL−1 and 7890.083 U·mL−1, respectively. Additionally, their colony sizes were marginally smaller than that of the wild-type strain TH-2 (ΔasAA::pyrG). After adding 1% CPPs, the maximum mannanase enzyme activities of strains MG and MD increased by factors of 1.07 and 1.14, respectively, while the colony sizes exhibited a significant increase. The transcription level of the mannanase man gene in strain MD was 1.01 times greater than that in strain MG. Furthermore, the transcription levels of the endoplasmic reticulum UPR marker genes hacA and bipA were elevated in strain MD compared to strain MG. The optimal temperature and pH of the recombinant mannanase Man were 70 ℃ and 4.5, respectively, and it showed high tolerance to metal ions and chemical reagents. Overexpression of the DnaJ1 gene and addition of 1% CPPs could effectively increase the expression level of mannanase Man. This study holds significant importance for the development and screening of mannanases with enhanced enzymatic properties and improved yield. |
| format | Article |
| id | doaj-art-9a06071fa3334ddb810111e1301002f9 |
| institution | Kabale University |
| issn | 1002-0306 |
| language | zho |
| publishDate | 2025-08-01 |
| publisher | The editorial department of Science and Technology of Food Industry |
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| series | Shipin gongye ke-ji |
| spelling | doaj-art-9a06071fa3334ddb810111e1301002f92025-08-20T04:03:21ZzhoThe editorial department of Science and Technology of Food IndustryShipin gongye ke-ji1002-03062025-08-01461521822710.13386/j.issn1002-0306.20240802102024080210-15Optimization Expression and Enzymatic Properties Research of Mannanase in Aspergillus nigerHui ZHANG0Zhu LI1Jiaxin MA2Junbo FAN3Zhiwei SHAO4Yujun JIANG5Jie LI6School of Life Sciences, Northeast Agricultural University, Harbin 150030, ChinaSchool of Life Sciences, Northeast Agricultural University, Harbin 150030, ChinaSchool of Life Sciences, Northeast Agricultural University, Harbin 150030, ChinaSchool of Life Sciences, Northeast Agricultural University, Harbin 150030, ChinaSchool of Life Sciences, Northeast Agricultural University, Harbin 150030, ChinaSchool of Life Sciences, Northeast Agricultural University, Harbin 150030, ChinaSchool of Life Sciences, Northeast Agricultural University, Harbin 150030, ChinaTo achieve higher expression levels and enhanced enzymatic properties of mannanase, this study employed homologous recombination and site-specific integration technology to construct a recombinant mannanase Aspergillus niger strain. Subsequently, the expression characteristics and enzymatic properties of the mannanase were systematically analyzed. This study successfully obtained the pure mannanase transformant MG of Aspergillus niger, which underwent homologous recombination at the glaA locus. Additionally, the pure mannanase transformant MD of Aspergillus niger was selected that not only exhibited homologous recombination at the glaA locus but also overexpressed the endoplasmic reticulum co-chaperone DnaJ1 gene at the asAA locus. Both strains MG and MD exhibited a distinct mannanase Man protein band at approximately 37 kDa, with the highest enzyme activities recorded at 6909.861 U·mL−1 and 7890.083 U·mL−1, respectively. Additionally, their colony sizes were marginally smaller than that of the wild-type strain TH-2 (ΔasAA::pyrG). After adding 1% CPPs, the maximum mannanase enzyme activities of strains MG and MD increased by factors of 1.07 and 1.14, respectively, while the colony sizes exhibited a significant increase. The transcription level of the mannanase man gene in strain MD was 1.01 times greater than that in strain MG. Furthermore, the transcription levels of the endoplasmic reticulum UPR marker genes hacA and bipA were elevated in strain MD compared to strain MG. The optimal temperature and pH of the recombinant mannanase Man were 70 ℃ and 4.5, respectively, and it showed high tolerance to metal ions and chemical reagents. Overexpression of the DnaJ1 gene and addition of 1% CPPs could effectively increase the expression level of mannanase Man. This study holds significant importance for the development and screening of mannanases with enhanced enzymatic properties and improved yield.http://www.spgykj.com/cn/article/doi/10.13386/j.issn1002-0306.2024080210mannanaseaspergillus nigerdnaj1cppsenzymatic properties |
| spellingShingle | Hui ZHANG Zhu LI Jiaxin MA Junbo FAN Zhiwei SHAO Yujun JIANG Jie LI Optimization Expression and Enzymatic Properties Research of Mannanase in Aspergillus niger Shipin gongye ke-ji mannanase aspergillus niger dnaj1 cpps enzymatic properties |
| title | Optimization Expression and Enzymatic Properties Research of Mannanase in Aspergillus niger |
| title_full | Optimization Expression and Enzymatic Properties Research of Mannanase in Aspergillus niger |
| title_fullStr | Optimization Expression and Enzymatic Properties Research of Mannanase in Aspergillus niger |
| title_full_unstemmed | Optimization Expression and Enzymatic Properties Research of Mannanase in Aspergillus niger |
| title_short | Optimization Expression and Enzymatic Properties Research of Mannanase in Aspergillus niger |
| title_sort | optimization expression and enzymatic properties research of mannanase in aspergillus niger |
| topic | mannanase aspergillus niger dnaj1 cpps enzymatic properties |
| url | http://www.spgykj.com/cn/article/doi/10.13386/j.issn1002-0306.2024080210 |
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