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High-level expression of Myrothecium verrucaria bilirubin oxidase in Pichia pastoris, and its facile purification and characterization
http://hdl.handle.net/2297/1732
http://hdl.handle.net/2297/17327ec5bf12-5c58-4a99-a80e-370f108332a4
名前 / ファイル | ライセンス | アクション |
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SC-PR-KATAOKA-K-BO-PEP.pdf (1.9 MB)
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Item type | 学術雑誌論文 / Journal Article(1) | |||||
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公開日 | 2017-10-03 | |||||
タイトル | ||||||
タイトル | High-level expression of Myrothecium verrucaria bilirubin oxidase in Pichia pastoris, and its facile purification and characterization | |||||
言語 | ||||||
言語 | eng | |||||
資源タイプ | ||||||
資源タイプ識別子 | http://purl.org/coar/resource_type/c_6501 | |||||
資源タイプ | journal article | |||||
著者 |
片岡, 邦重
× 片岡, 邦重× Tanaka, Kazuhiro× Sakai, Yoko× Sakurai, Takeshi |
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書誌情報 |
Protein expression and purification 巻 41, 号 1, p. 77-83, 発行日 2005-05-01 |
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ISSN | ||||||
収録物識別子タイプ | ISSN | |||||
収録物識別子 | 1046-5928 | |||||
DOI | ||||||
関連タイプ | isVersionOf | |||||
識別子タイプ | DOI | |||||
関連識別子 | https://doi.org/10.1016/j.pep.2005.02.001 | |||||
出版者 | ||||||
出版者 | Elsevier | |||||
抄録 | ||||||
内容記述タイプ | Abstract | |||||
内容記述 | Bilirubin oxidase (BO) from Myrothecium verrucaria (authentic BO) catalyzing the oxidation of bilirubin to biliverdine was overexpressed in the methylotrophic yeast, Pichia pastoris. The cDNA encoding BO was cloned into the P. pastoris expression vector pPIC9K under the control of the alcohol oxidase 1 promoter and its protein product was secreted using the Saccharomyces cerevisiae α-mating factor signal sequence. The productivity of recombinant BO (rBO) in P. pastoris was approximately 5000 U/L of culture broth, being about 2.5- and 250-fold higher than rBO expressed in Aspergillus oryzae and S. cerevisiae, respectively. The calculated molecular mass of rBO consisting of 538 amino acids was 60,493 kDa, however, that of SDS–PAGE was 66 kDa because of non-native type N-linked sugar chains. The spectroscopic properties of rBO were typical of multicopper oxidase containing four Cu ions per protein molecule. The specific activity to oxidize bilirubin was 57 U/mg, having a value about twice that of authentic BO and rBO expressed in A. oryzae. Moreover, the thermostability of rBO expressed in P. pastoris was significantly high compared to the authentic BO previously reported. Accordingly, a heterologous expression system of rBO to meet clinical and industrial needs was constructed | |||||
著者版フラグ | ||||||
出版タイプ | AM | |||||
出版タイプResource | http://purl.org/coar/version/c_ab4af688f83e57aa |