Cloning, expression and characterization of an aspartate aminotransferase gene from Lactobacillus brevis CGMCC 1306
An aspartate aminotransferase (AATase) gene from Lactobacillus brevis CGMCC 1306 was cloned, which contains a 1182-bp open reading frame coding for 393 amino acids (41.43 kDa). When expressed in Escherichia coli BL21 (DE3), the recombinant AATase was purified and subsequently characterized. The reco...
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| Format: | Article |
| Language: | English |
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Taylor & Francis Group
2017-05-01
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| Series: | Biotechnology & Biotechnological Equipment |
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| Online Access: | http://dx.doi.org/10.1080/13102818.2017.1304181 |
| _version_ | 1818152951980490752 |
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| author | Sheng Hu Xiang Zhang Yi Lu Yue-Cheng Lin Dong-Fang Xie Hui Fang Jun Huang Le-He Mei |
| author_facet | Sheng Hu Xiang Zhang Yi Lu Yue-Cheng Lin Dong-Fang Xie Hui Fang Jun Huang Le-He Mei |
| author_sort | Sheng Hu |
| collection | DOAJ |
| description | An aspartate aminotransferase (AATase) gene from Lactobacillus brevis CGMCC 1306 was cloned, which contains a 1182-bp open reading frame coding for 393 amino acids (41.43 kDa). When expressed in Escherichia coli BL21 (DE3), the recombinant AATase was purified and subsequently characterized. The recombinant AATase can catalyse the conversion of L-Asp to L-Glu, and the kcat/Km was determined to be 25.5 (mmol/L)−1 s−1 for L-Asp and 207.8 m(mol/L)−1 s−1 for α-ketoglutarate. With optimum temperature as 25 ˚C, the AATase may be a novel and special psychrophilic enzyme which exhibited a good thermal stability below 55 ˚C. The conserved active site residue of AATase was identified as Lys237 by phylogenetic analysis. Secondary structure of the enzyme includes α-helix (39.2%), β-sheet (5.5%), β-turn (8.8%), and random coil (36.5%) by circular dichroism spectral analysis. Phase diagram for the fluorescence data analysis showed that guanidinium chloride-induced unfolding of AATase involved at least one intermediate. |
| first_indexed | 2024-12-11T14:02:53Z |
| format | Article |
| id | doaj.art-0d351b41c70642b7a973d990653a4434 |
| institution | Directory Open Access Journal |
| issn | 1310-2818 1314-3530 |
| language | English |
| last_indexed | 2024-12-11T14:02:53Z |
| publishDate | 2017-05-01 |
| publisher | Taylor & Francis Group |
| record_format | Article |
| series | Biotechnology & Biotechnological Equipment |
| spelling | doaj.art-0d351b41c70642b7a973d990653a44342022-12-22T01:03:46ZengTaylor & Francis GroupBiotechnology & Biotechnological Equipment1310-28181314-35302017-05-0131354455310.1080/13102818.2017.13041811304181Cloning, expression and characterization of an aspartate aminotransferase gene from Lactobacillus brevis CGMCC 1306Sheng Hu0Xiang Zhang1Yi Lu2Yue-Cheng Lin3Dong-Fang Xie4Hui Fang5Jun Huang6Le-He Mei7Ningbo Institute of Technology, Zhejiang UniversityZhejiang University of Science and TechnologyZhejiang University of Science and TechnologyZhejiang University of Science and TechnologyZhejiang University of Science and TechnologyZhejiang University of Science and TechnologyZhejiang University of Science and TechnologyNingbo Institute of Technology, Zhejiang UniversityAn aspartate aminotransferase (AATase) gene from Lactobacillus brevis CGMCC 1306 was cloned, which contains a 1182-bp open reading frame coding for 393 amino acids (41.43 kDa). When expressed in Escherichia coli BL21 (DE3), the recombinant AATase was purified and subsequently characterized. The recombinant AATase can catalyse the conversion of L-Asp to L-Glu, and the kcat/Km was determined to be 25.5 (mmol/L)−1 s−1 for L-Asp and 207.8 m(mol/L)−1 s−1 for α-ketoglutarate. With optimum temperature as 25 ˚C, the AATase may be a novel and special psychrophilic enzyme which exhibited a good thermal stability below 55 ˚C. The conserved active site residue of AATase was identified as Lys237 by phylogenetic analysis. Secondary structure of the enzyme includes α-helix (39.2%), β-sheet (5.5%), β-turn (8.8%), and random coil (36.5%) by circular dichroism spectral analysis. Phase diagram for the fluorescence data analysis showed that guanidinium chloride-induced unfolding of AATase involved at least one intermediate.http://dx.doi.org/10.1080/13102818.2017.1304181Aspartate aminotransferasepsychrophilic enzymephase diagramphylogenetic analysis; Lactobacillus brevis |
| spellingShingle | Sheng Hu Xiang Zhang Yi Lu Yue-Cheng Lin Dong-Fang Xie Hui Fang Jun Huang Le-He Mei Cloning, expression and characterization of an aspartate aminotransferase gene from Lactobacillus brevis CGMCC 1306 Biotechnology & Biotechnological Equipment Aspartate aminotransferase psychrophilic enzyme phase diagram phylogenetic analysis; Lactobacillus brevis |
| title | Cloning, expression and characterization of an aspartate aminotransferase gene from Lactobacillus brevis CGMCC 1306 |
| title_full | Cloning, expression and characterization of an aspartate aminotransferase gene from Lactobacillus brevis CGMCC 1306 |
| title_fullStr | Cloning, expression and characterization of an aspartate aminotransferase gene from Lactobacillus brevis CGMCC 1306 |
| title_full_unstemmed | Cloning, expression and characterization of an aspartate aminotransferase gene from Lactobacillus brevis CGMCC 1306 |
| title_short | Cloning, expression and characterization of an aspartate aminotransferase gene from Lactobacillus brevis CGMCC 1306 |
| title_sort | cloning expression and characterization of an aspartate aminotransferase gene from lactobacillus brevis cgmcc 1306 |
| topic | Aspartate aminotransferase psychrophilic enzyme phase diagram phylogenetic analysis; Lactobacillus brevis |
| url | http://dx.doi.org/10.1080/13102818.2017.1304181 |
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