Biochemical Characterization of a Novel Alkaline-Tolerant Xaa-Pro Dipeptidase from <i>Aspergillus phoenicis</i>
Xaa-Pro dipeptidase (XPD, EC 3.4.13.9; also known as prolidase) catalyzes the hydrolysis of the iminopeptide bond in the trans-Xaa-Pro dipeptides (Xaa represents any amino acid except proline), which makes it find wide applications in food, medical and environmental protection fields. In the present...
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MDPI AG
2023-11-01
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| Series: | Fermentation |
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| Online Access: | https://www.mdpi.com/2311-5637/9/11/978 |
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| author | Zixing Dong Shuangshuang Yang Kun Zhang Cunduo Tang Yunchao Kan Lunguang Yao |
| author_facet | Zixing Dong Shuangshuang Yang Kun Zhang Cunduo Tang Yunchao Kan Lunguang Yao |
| author_sort | Zixing Dong |
| collection | DOAJ |
| description | Xaa-Pro dipeptidase (XPD, EC 3.4.13.9; also known as prolidase) catalyzes the hydrolysis of the iminopeptide bond in the trans-Xaa-Pro dipeptides (Xaa represents any amino acid except proline), which makes it find wide applications in food, medical and environmental protection fields. In the present study, a novel Xaa-Pro dipeptidase from <i>Aspergillus phoenicis</i> ATCC 14332 (ApXPD) was heterologously expressed and biochemically characterized. Reclassification based on phylogenetic analysis and the version 12.5 MEROPS database showed that this enzyme was the only fungal XPD in the unassigned subfamily that shared the highest sequence identity with <i>Xanthomonas campestris</i> prolidase but not with that from the more related fungal species <i>A. niudulans</i>. As compared with other prolidases, ApXPD also contained a long N-terminal tail (residues 1–63) and an additional region (PAPARLREKL) and used a different arginine residue for dipeptide selectivity. After heterologous expression and partial purification, recombinant ApXPD was highly active and stable over the alkaline range from 8.5 to 10.0, with maximum activity at pH 9.0 and more than 80% activity retained after 1 h incubation at pHs of 8.5–10.0 (55 °C). It also had an apparent optimum temperature of 55 °C and remained stable at 20–30 °C. Moreover, this enzyme was a cobalt-dependent prolidase that only cleaved dipeptides Lys-Pro, Gly-Pro, and Ala-Pro rather than other dipeptides, tripeptides, and tetrapeptides. All these distinct features make <i>A. phoenicis</i> ATCC 14332 XPD unique among currently known prolidases, thus defining a novel Xaa-Pro dipeptidase subfamily. |
| first_indexed | 2024-03-09T16:50:49Z |
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| institution | Directory Open Access Journal |
| issn | 2311-5637 |
| language | English |
| last_indexed | 2024-03-09T16:50:49Z |
| publishDate | 2023-11-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Fermentation |
| spelling | doaj.art-5acdb610b86347d5980ead5042a6874c2023-11-24T14:41:25ZengMDPI AGFermentation2311-56372023-11-0191197810.3390/fermentation9110978Biochemical Characterization of a Novel Alkaline-Tolerant Xaa-Pro Dipeptidase from <i>Aspergillus phoenicis</i>Zixing Dong0Shuangshuang Yang1Kun Zhang2Cunduo Tang3Yunchao Kan4Lunguang Yao5Henan Provincial Engineering Laboratory of Insect Bio-Reactor, College of Life Science and Agricultural Engineering, Nanyang Normal University, Nanyang 473061, ChinaCollege of Physical Education, Nanyang Normal University, Nanyang 473061, ChinaHenan Provincial Engineering Laboratory of Insect Bio-Reactor, College of Life Science and Agricultural Engineering, Nanyang Normal University, Nanyang 473061, ChinaHenan Provincial Engineering Laboratory of Insect Bio-Reactor, College of Life Science and Agricultural Engineering, Nanyang Normal University, Nanyang 473061, ChinaHenan Provincial Engineering Laboratory of Insect Bio-Reactor, College of Life Science and Agricultural Engineering, Nanyang Normal University, Nanyang 473061, ChinaHenan Provincial Engineering Laboratory of Insect Bio-Reactor, College of Life Science and Agricultural Engineering, Nanyang Normal University, Nanyang 473061, ChinaXaa-Pro dipeptidase (XPD, EC 3.4.13.9; also known as prolidase) catalyzes the hydrolysis of the iminopeptide bond in the trans-Xaa-Pro dipeptides (Xaa represents any amino acid except proline), which makes it find wide applications in food, medical and environmental protection fields. In the present study, a novel Xaa-Pro dipeptidase from <i>Aspergillus phoenicis</i> ATCC 14332 (ApXPD) was heterologously expressed and biochemically characterized. Reclassification based on phylogenetic analysis and the version 12.5 MEROPS database showed that this enzyme was the only fungal XPD in the unassigned subfamily that shared the highest sequence identity with <i>Xanthomonas campestris</i> prolidase but not with that from the more related fungal species <i>A. niudulans</i>. As compared with other prolidases, ApXPD also contained a long N-terminal tail (residues 1–63) and an additional region (PAPARLREKL) and used a different arginine residue for dipeptide selectivity. After heterologous expression and partial purification, recombinant ApXPD was highly active and stable over the alkaline range from 8.5 to 10.0, with maximum activity at pH 9.0 and more than 80% activity retained after 1 h incubation at pHs of 8.5–10.0 (55 °C). It also had an apparent optimum temperature of 55 °C and remained stable at 20–30 °C. Moreover, this enzyme was a cobalt-dependent prolidase that only cleaved dipeptides Lys-Pro, Gly-Pro, and Ala-Pro rather than other dipeptides, tripeptides, and tetrapeptides. All these distinct features make <i>A. phoenicis</i> ATCC 14332 XPD unique among currently known prolidases, thus defining a novel Xaa-Pro dipeptidase subfamily.https://www.mdpi.com/2311-5637/9/11/978Xaa-Pro dipeptidase<i>Aspergillus phoenicis</i>reclassificationheterologous expressionbiochemical characterization |
| spellingShingle | Zixing Dong Shuangshuang Yang Kun Zhang Cunduo Tang Yunchao Kan Lunguang Yao Biochemical Characterization of a Novel Alkaline-Tolerant Xaa-Pro Dipeptidase from <i>Aspergillus phoenicis</i> Fermentation Xaa-Pro dipeptidase <i>Aspergillus phoenicis</i> reclassification heterologous expression biochemical characterization |
| title | Biochemical Characterization of a Novel Alkaline-Tolerant Xaa-Pro Dipeptidase from <i>Aspergillus phoenicis</i> |
| title_full | Biochemical Characterization of a Novel Alkaline-Tolerant Xaa-Pro Dipeptidase from <i>Aspergillus phoenicis</i> |
| title_fullStr | Biochemical Characterization of a Novel Alkaline-Tolerant Xaa-Pro Dipeptidase from <i>Aspergillus phoenicis</i> |
| title_full_unstemmed | Biochemical Characterization of a Novel Alkaline-Tolerant Xaa-Pro Dipeptidase from <i>Aspergillus phoenicis</i> |
| title_short | Biochemical Characterization of a Novel Alkaline-Tolerant Xaa-Pro Dipeptidase from <i>Aspergillus phoenicis</i> |
| title_sort | biochemical characterization of a novel alkaline tolerant xaa pro dipeptidase from i aspergillus phoenicis i |
| topic | Xaa-Pro dipeptidase <i>Aspergillus phoenicis</i> reclassification heterologous expression biochemical characterization |
| url | https://www.mdpi.com/2311-5637/9/11/978 |
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