論文 - 詳細
| RRC ID | 15818 |
|---|---|
| 著者 | Li Y, Kawakami N, Ogola HJ, Ashida H, Ishikawa T, Shibata H, Sawa Y. |
| タイトル | A novel L-aspartate dehydrogenase from the mesophilic bacterium Pseudomonas aeruginosa PAO1: molecular characterization and application for L-aspartate production. |
| ジャーナル | Appl Microbiol Biotechnol |
| Abstract |
L-aspartate dehydrogenase (EC 1.4.1.21; L: -AspDH) is a rare member of amino acid dehydrogenase superfamily and so far, two thermophilic enzymes have been reported. In our study, an ORF PA3505 encoding for a putative L-AspDH in the mesophilic bacterium Pseudomonas aeruginosa PAO1 was identified, cloned, and overexpressed in Escherichia coli. The homogeneously purified enzyme (PaeAspDH) was a dimeric protein with a molecular mass of about 28 kDa exhibiting a very high specific activity for L-aspartate (L-Asp) and oxaloacetate (OAA) of 127 and 147 U mg(-1), respectively. The enzyme was capable of utilizing both nicotinamide adenine dinucleotide (NAD) and nicotinamide adenine dinucleotide phosphate (NADP) as coenzyme. PaeAspDH showed a T (m) value of 48°C for 20 min that was improved to approximately 60°C by the addition of 0.4 M NaCl or 30% glycerol. The apparent K (m) values for OAA, NADH, and ammonia were 2.12, 0.045, and 10.1 mM, respectively; comparable results were observed with NADPH. The L-Asp production system B consisting of PaeAspDH, Bacillus subtilis malate dehydrogenase and E. coli fumarase, achieved a high level of L-Asp production (625 mM) from fumarate in fed-batch process with a molar conversion yield of 89.4%. Furthermore, the fermentative production system C released 33 mM of L-Asp after 50 h by using succinate as carbon source. This study represented an extensive characterization of the mesophilic AspDH and its potential applicability for efficient and attractive production of L-Asp. Our novel production systems are also hopeful for developing the new processes for other compounds production. |
| 巻・号 | 90(6) |
| ページ | 1953-62 |
| 公開日 | 2011-6-1 |
| DOI | 10.1007/s00253-011-3208-4 |
| PMID | 21468714 |
| MeSH | Amino Acid Oxidoreductases / chemistry Amino Acid Oxidoreductases / genetics* Amino Acid Oxidoreductases / isolation & purification Amino Acid Oxidoreductases / metabolism* Aspartic Acid / metabolism* Cloning, Molecular Coenzymes / metabolism Enzyme Stability Escherichia coli / genetics Kinetics Molecular Weight NAD / metabolism NADP / metabolism Oxaloacetic Acid / metabolism Protein Multimerization Pseudomonas aeruginosa / enzymology* Pseudomonas aeruginosa / genetics* Recombinant Proteins / chemistry Recombinant Proteins / genetics Recombinant Proteins / isolation & purification Recombinant Proteins / metabolism Substrate Specificity Temperature |
| IF | 3.53 |
| 引用数 | 9 |
| WOS 分野 | BIOTECHNOLOGY & APPLIED MICROBIOLOGY |
| オルトメトリクス指標 |
オルトメトリクス指標項目
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| 最多言及媒体 | Patent(IFI CLAIMS) |
| 各媒体での言及数の合計 | 6 |
| 過去6か月間でのオルトメトリクス指標の変動値 | 0.0 |
| リソース情報 | |
| 一般微生物 | JCM 14847 |