Two Highly Similar Chitinases from Marine <i>Vibrio</i> Species have Different Enzymatic Properties
Chitinase, as one of the most important extracellular enzymes in the marine environment, has great ecological and applied values. In this study, two chitinases (Chi1557 and Chi4668) with 97.33% amino acid sequences identity were individually found in <i>Vibrio rotiferianus</i> and <i&...
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| Format: | Article |
| Language: | English |
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MDPI AG
2020-02-01
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| Series: | Marine Drugs |
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| Online Access: | https://www.mdpi.com/1660-3397/18/3/139 |
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| author | Xinxin He Min Yu Yanhong Wu Lingman Ran Weizhi Liu Xiao-Hua Zhang |
| author_facet | Xinxin He Min Yu Yanhong Wu Lingman Ran Weizhi Liu Xiao-Hua Zhang |
| author_sort | Xinxin He |
| collection | DOAJ |
| description | Chitinase, as one of the most important extracellular enzymes in the marine environment, has great ecological and applied values. In this study, two chitinases (Chi1557 and Chi4668) with 97.33% amino acid sequences identity were individually found in <i>Vibrio rotiferianus</i> and <i>Vibrio harveyi</i>. They both were encoding by 561 amino acids, but differed in 15 amino acids and showed different enzymatic properties. The optimal temperature and pH ranges were 45−50 °C and pH 5.0−7.0 for Chi1557, while ~50 °C and pH 3.0−6.0 for Chi4668. K<sup>+</sup>, Mg<sup>2+</sup>, and EDTA increased the enzymatic activity of Chi4668 significantly, yet these factors were inhibitory to Chi1557. Moreover, Chi1557 degraded colloidal chitin to produce (GlcNAc)<sub>2</sub> and minor GlcNAc, whereas Chi4668 produce (GlcNAc)<sub>2 </sub>with minor (GlcNAc)<sub>3</sub> and (GlcNAc)<sub>4</sub>. The <i>K</i>cat/<i>K</i>m of Chi4668 was ~4.7 times higher than that of Chi1557, indicating that Chi4668 had stronger catalytic activity than Chi1557. Furthermore, site-directed mutagenesis was performed on Chi1557 focusing on seven conserved amino acid residues of family GH18 chitinases. Chi1557 was almost completely inactive after Glu154, Gln219, Tyr221, or Trp312 was individually mutated, retained ~50% activity after Tyr37 was mutated, and increased two times activity after Asp152 was mutated, indicating that these six amino acids were key sites for Chi1557. |
| first_indexed | 2024-04-12T05:37:46Z |
| format | Article |
| id | doaj.art-681eddce6e9b4762a51c7093ae1c0010 |
| institution | Directory Open Access Journal |
| issn | 1660-3397 |
| language | English |
| last_indexed | 2024-04-12T05:37:46Z |
| publishDate | 2020-02-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Marine Drugs |
| spelling | doaj.art-681eddce6e9b4762a51c7093ae1c00102022-12-22T03:45:46ZengMDPI AGMarine Drugs1660-33972020-02-0118313910.3390/md18030139md18030139Two Highly Similar Chitinases from Marine <i>Vibrio</i> Species have Different Enzymatic PropertiesXinxin He0Min Yu1Yanhong Wu2Lingman Ran3Weizhi Liu4Xiao-Hua Zhang5College of Marine Life Sciences, Ocean University of China, Qingdao 266003, ChinaCollege of Marine Life Sciences, Ocean University of China, Qingdao 266003, ChinaCollege of Marine Life Sciences, Ocean University of China, Qingdao 266003, ChinaCollege of Marine Life Sciences, Ocean University of China, Qingdao 266003, ChinaCollege of Marine Life Sciences, Ocean University of China, Qingdao 266003, ChinaCollege of Marine Life Sciences, Ocean University of China, Qingdao 266003, ChinaChitinase, as one of the most important extracellular enzymes in the marine environment, has great ecological and applied values. In this study, two chitinases (Chi1557 and Chi4668) with 97.33% amino acid sequences identity were individually found in <i>Vibrio rotiferianus</i> and <i>Vibrio harveyi</i>. They both were encoding by 561 amino acids, but differed in 15 amino acids and showed different enzymatic properties. The optimal temperature and pH ranges were 45−50 °C and pH 5.0−7.0 for Chi1557, while ~50 °C and pH 3.0−6.0 for Chi4668. K<sup>+</sup>, Mg<sup>2+</sup>, and EDTA increased the enzymatic activity of Chi4668 significantly, yet these factors were inhibitory to Chi1557. Moreover, Chi1557 degraded colloidal chitin to produce (GlcNAc)<sub>2</sub> and minor GlcNAc, whereas Chi4668 produce (GlcNAc)<sub>2 </sub>with minor (GlcNAc)<sub>3</sub> and (GlcNAc)<sub>4</sub>. The <i>K</i>cat/<i>K</i>m of Chi4668 was ~4.7 times higher than that of Chi1557, indicating that Chi4668 had stronger catalytic activity than Chi1557. Furthermore, site-directed mutagenesis was performed on Chi1557 focusing on seven conserved amino acid residues of family GH18 chitinases. Chi1557 was almost completely inactive after Glu154, Gln219, Tyr221, or Trp312 was individually mutated, retained ~50% activity after Tyr37 was mutated, and increased two times activity after Asp152 was mutated, indicating that these six amino acids were key sites for Chi1557.https://www.mdpi.com/1660-3397/18/3/139chitinchitinases<i>vibrio</i>enzymatic propertiessite-directed mutagenesis |
| spellingShingle | Xinxin He Min Yu Yanhong Wu Lingman Ran Weizhi Liu Xiao-Hua Zhang Two Highly Similar Chitinases from Marine <i>Vibrio</i> Species have Different Enzymatic Properties Marine Drugs chitin chitinases <i>vibrio</i> enzymatic properties site-directed mutagenesis |
| title | Two Highly Similar Chitinases from Marine <i>Vibrio</i> Species have Different Enzymatic Properties |
| title_full | Two Highly Similar Chitinases from Marine <i>Vibrio</i> Species have Different Enzymatic Properties |
| title_fullStr | Two Highly Similar Chitinases from Marine <i>Vibrio</i> Species have Different Enzymatic Properties |
| title_full_unstemmed | Two Highly Similar Chitinases from Marine <i>Vibrio</i> Species have Different Enzymatic Properties |
| title_short | Two Highly Similar Chitinases from Marine <i>Vibrio</i> Species have Different Enzymatic Properties |
| title_sort | two highly similar chitinases from marine i vibrio i species have different enzymatic properties |
| topic | chitin chitinases <i>vibrio</i> enzymatic properties site-directed mutagenesis |
| url | https://www.mdpi.com/1660-3397/18/3/139 |
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