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ツゲ ヒデアキ
TSUGE HIDEAKI
津下 英明 所属 京都産業大学 生命科学部 先端生命科学科 職種 教授 |
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| 言語種別 | 英語 |
| 発行・発表の年月 | 2009/10 |
| 形態種別 | 研究論文 |
| 査読 | 査読あり |
| 標題 | Refolding, characterization and crystal structure of (S)-malate dehydrogenase from the hyperthermophilic archaeon Aeropyrum pernix |
| 執筆形態 | その他 |
| 掲載誌名 | BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS |
| 出版社・発行元 | ELSEVIER SCIENCE BV |
| 巻・号・頁 | 1794(10),pp.1496-1504 |
| 著者・共著者 | Ryushi Kawakami,Haruhiko Sakuraba,Shuichiro Goda,Hideaki Tsuge,Toshihisa Ohshima |
| 概要 | Tartrate oxidation activity was found in the crude extract of an aerobic hyperthermophilic archaeon Aeropyrum pernix, and the enzyme was identified as (S)-malate dehydrogenase (MDH), which, when produced in Escherichia coli, was mainly obtained as an inactive inclusion body. The inclusion body was dissolved in 6 M guanidine-HCl and gradually refolded to the active enzyme through dilution of the denaturant. The purified recombinant enzyme consisted of four identical subunits with a molecular mass of about 110 kDa. NADP was preferred as a coenzyme over NAD for (S)-malate oxidation and, unlike MDHs from other sources, this enzyme readily catalyzed the oxidation of (2S,3S)-tartrate and (2S,3R)-tartrate. The tartrate oxidation activity was also observed in MDHs from the hyperthermophilic archaea Methanocaldococcus jannaschii and Archaeoglobus fulgidus, suggesting these hyperthermophilic MDHs loosely bind their substrates. The refolded A. pernix MDH was also crystallized, and the structure was determined at a resolution of 2.9 angstrom. Its overall structure was similar to those of the M. jannaschii, Chloraflexus aurantiacus, Chlorobium vibrioforme and Cryptosporidium parvum [lactate dehydrogenase-like] MDHs with root-mean-square-deviation values between 1.4 and 2.1 angstrom. Consistent with earlier reports, Ala at position 53 was responsible for coenzyme specificity, and the next residue, Arg, was important for NADP binding. Structural comparison revealed that the hyperthermostability of the A. pernix MDH is likely attributable to its smaller cavity volume and larger numbers of ion pairs and ion-pair networks, but the molecular strategy for thermostability may be specific for each enzyme. (C) 2009 Elsevier B.V. All rights reserved. |
| DOI | 10.1016/j.bbapap.2009.06.014 |
| ISSN | 1570-9639/0006-3002 |