Heterologous Expression and Functional Analysis of <i>Exiguobacterium</i> Algin Lyase Gene by <i>Pichia pastoris</i>
Algin is the most abundant substance in alga. Alginate lyase degrades algin and produces algin monosaccharides, disaccharides, and oligosaccharides, which are widely used in bioenergy, food, medicine, and other fields. In this study, one <i>Exiguobacterium</i> strain isolated from rotten...
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2025-01-01
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| author | Hanwen Wu Kai Hou Yutong Jiang Mingjian Luan Yuxia Sun Xi He Xiangzhong Zhao |
| author_facet | Hanwen Wu Kai Hou Yutong Jiang Mingjian Luan Yuxia Sun Xi He Xiangzhong Zhao |
| author_sort | Hanwen Wu |
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| description | Algin is the most abundant substance in alga. Alginate lyase degrades algin and produces algin monosaccharides, disaccharides, and oligosaccharides, which are widely used in bioenergy, food, medicine, and other fields. In this study, one <i>Exiguobacterium</i> strain isolated from rotten kelp exhibited a robust ability to degrade the alga. The sequencing of this strain revealed the presence of three different types of algin alginate lyase. Nevertheless, the expression of three genes in <i>Escherichia coli</i> revealed a lower alginate lyase activity compared to that of the original strain. After codon optimization, the gene with the highest activity of the three was successfully expressed in <i>Pichia pastoris</i> to produce recombinant EbAlg664. The activity of the recombinant enzyme in 5 L high-density fermentation reached 1306 U/mg protein, 3.9 times that of the original <i>Exiguobacterium</i> strain. The results of the enzymatic analysis revealed that the optimal temperature and the pH range of recombinant EbAlg664 were narrower compared to the original strain. Additionally, the presence of Cu<sup>2+</sup> and Co<sup>2+</sup> enhanced the enzymatic activity, whereas Mg<sup>2+</sup> and Fe<sup>3+</sup> exhibited inhibitory effects on the recombinant alginate lyase. The study offers a theoretical and practical foundation for the industrial-scale production of engineered <i>Pichia pastoris</i> with high alginate lyase activity. |
| format | Article |
| id | doaj-art-69c60bdfd854478bb01d9373bde8825c |
| institution | Kabale University |
| issn | 2311-5637 |
| language | English |
| publishDate | 2025-01-01 |
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| series | Fermentation |
| spelling | doaj-art-69c60bdfd854478bb01d9373bde8825c2025-01-24T13:32:08ZengMDPI AGFermentation2311-56372025-01-011113410.3390/fermentation11010034Heterologous Expression and Functional Analysis of <i>Exiguobacterium</i> Algin Lyase Gene by <i>Pichia pastoris</i>Hanwen Wu0Kai Hou1Yutong Jiang2Mingjian Luan3Yuxia Sun4Xi He5Xiangzhong Zhao6School of Biologic Engineering, Qi Lu University of Technology (Shandong Academy of Sciences), Jinan 250353, ChinaSchool of Biologic Engineering, Qi Lu University of Technology (Shandong Academy of Sciences), Jinan 250353, ChinaSchool of Biologic Engineering, Qi Lu University of Technology (Shandong Academy of Sciences), Jinan 250353, ChinaSchool of Biologic Engineering, Qi Lu University of Technology (Shandong Academy of Sciences), Jinan 250353, ChinaShandong Academy of Grape/Winegrape and Wine Technological Innovation Center of Shandong Province, Jinan 250199, ChinaSchool of Biologic Engineering, Qi Lu University of Technology (Shandong Academy of Sciences), Jinan 250353, ChinaSchool of Biologic Engineering, Qi Lu University of Technology (Shandong Academy of Sciences), Jinan 250353, ChinaAlgin is the most abundant substance in alga. Alginate lyase degrades algin and produces algin monosaccharides, disaccharides, and oligosaccharides, which are widely used in bioenergy, food, medicine, and other fields. In this study, one <i>Exiguobacterium</i> strain isolated from rotten kelp exhibited a robust ability to degrade the alga. The sequencing of this strain revealed the presence of three different types of algin alginate lyase. Nevertheless, the expression of three genes in <i>Escherichia coli</i> revealed a lower alginate lyase activity compared to that of the original strain. After codon optimization, the gene with the highest activity of the three was successfully expressed in <i>Pichia pastoris</i> to produce recombinant EbAlg664. The activity of the recombinant enzyme in 5 L high-density fermentation reached 1306 U/mg protein, 3.9 times that of the original <i>Exiguobacterium</i> strain. The results of the enzymatic analysis revealed that the optimal temperature and the pH range of recombinant EbAlg664 were narrower compared to the original strain. Additionally, the presence of Cu<sup>2+</sup> and Co<sup>2+</sup> enhanced the enzymatic activity, whereas Mg<sup>2+</sup> and Fe<sup>3+</sup> exhibited inhibitory effects on the recombinant alginate lyase. The study offers a theoretical and practical foundation for the industrial-scale production of engineered <i>Pichia pastoris</i> with high alginate lyase activity.https://www.mdpi.com/2311-5637/11/1/34alginate lyase<i>Exiguobacterium</i>heteroexpression<i>Pichia pastoris</i>high-density fermentation |
| spellingShingle | Hanwen Wu Kai Hou Yutong Jiang Mingjian Luan Yuxia Sun Xi He Xiangzhong Zhao Heterologous Expression and Functional Analysis of <i>Exiguobacterium</i> Algin Lyase Gene by <i>Pichia pastoris</i> Fermentation alginate lyase <i>Exiguobacterium</i> heteroexpression <i>Pichia pastoris</i> high-density fermentation |
| title | Heterologous Expression and Functional Analysis of <i>Exiguobacterium</i> Algin Lyase Gene by <i>Pichia pastoris</i> |
| title_full | Heterologous Expression and Functional Analysis of <i>Exiguobacterium</i> Algin Lyase Gene by <i>Pichia pastoris</i> |
| title_fullStr | Heterologous Expression and Functional Analysis of <i>Exiguobacterium</i> Algin Lyase Gene by <i>Pichia pastoris</i> |
| title_full_unstemmed | Heterologous Expression and Functional Analysis of <i>Exiguobacterium</i> Algin Lyase Gene by <i>Pichia pastoris</i> |
| title_short | Heterologous Expression and Functional Analysis of <i>Exiguobacterium</i> Algin Lyase Gene by <i>Pichia pastoris</i> |
| title_sort | heterologous expression and functional analysis of i exiguobacterium i algin lyase gene by i pichia pastoris i |
| topic | alginate lyase <i>Exiguobacterium</i> heteroexpression <i>Pichia pastoris</i> high-density fermentation |
| url | https://www.mdpi.com/2311-5637/11/1/34 |
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