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Hokkaido University Collection of Scholarly and Academic Papers >
Graduate School of Agriculture / Faculty of Agriculture >
Peer-reviewed Journal Articles, etc >
Purification, characterization and amino acid sequence of a novel enzyme, D-threo-3-hydroxyaspartate dehydratase, from Delftia sp. HT23
| Title: | Purification, characterization and amino acid sequence of a novel enzyme, D-threo-3-hydroxyaspartate dehydratase, from Delftia sp. HT23 |
| Authors: | Maeda, Takayuki Browse this author | | Takeda, Yuki Browse this author | | Murakami, Tomoko Browse this author | | Yokota, Atsushi Browse this author | | Wada, Masaru Browse this author →KAKEN DB |
| Keywords: | alanine racemase | | Delftia sp. HT23 | | D-threo-3-hydroxyaspartate dehydratase | | pyridoxal 5'-phosphate |
| Issue Date: | Dec-2010 |
| Publisher: | Oxford University Press |
| Journal Title: | The Journal of Biochemistry |
| Volume: | 148 |
| Issue: | 6 |
| Start Page: | 705 |
| End Page: | 712 |
| Publisher DOI: | 10.1093/jb/mvq106 |
| PMID: | 20843822 |
| Abstract: | D-threo-3-hydroxyaspartate dehydratase (D-THA DH) was purified from the cell-free extract of the soil-isolated bacterium Delftia sp. HT23. The enzyme exhibited dehydratase activity towards D-threo-3-hydroxyaspartate, L-threo-3-hydroxyaspartate, L-erythro-3-hydroxyaspartate and D-serine. Absorption of the purified enzyme at 412 nm suggests that it contains pyridoxal 5'-phosphate (PLP) as a cofactor. The NH2-terminal and internal amino acid sequences showed significant similarity to hypothetical alanine racemase of genome-sequenced Delftia acidovorans SPH-1; however, the purified enzyme showed no alanine racemase activity. Using the sequence information of Delftia acidovorans SPH-1, the gene encoding D-THA DH was cloned. The deduced amino acid sequence, which belongs to the alanine racemase family, shows significant (26-36%) similarity to D-serine dehydratase of both Saccharomyces cerevisiae and chicken. In order to obtain purified D-THA DH efficiently, the gene was expressed in Escherichia coli. The recombinant enzyme was highly activated by divalent cations, such as Mn2+, Co2+ and Ni2+. Site-directed mutagenesis experiment revealed that lysine 43 is an important residue involved in PLP binding and catalysis. This is the first reported enzyme that acts on D-THA. In addition, this enzyme is the first example of a prokaryotic dehydratase belonging to the fold-type III PLP-dependent enzyme family. |
| Rights: | This is a pre-copy-editing, author-produced PDF of an article accepted for publication in The Journal of Biochemistry following peer review. The definitive publisher-authenticated version, J Biochem (2010) 148 (6): 705-712, is available online at: http://jb.oxfordjournals.org/content/148/6/705 |
| Type: | article (author version) |
| URI: | http://hdl.handle.net/2115/47109 |
| Appears in Collections: | 農学院・農学研究院 (Graduate School of Agriculture / Faculty of Agriculture) > 雑誌発表論文等 (Peer-reviewed Journal Articles, etc)
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Submitter: 和田 大
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