| Title Alternative | Studies on l-Glutamate Oxidase with Strict Substrate Specificity from Streptomyces sp. |
|---|---|
| FullText URL | srfa_112_013_018.pdf |
| Author | Nakayama, Natsume| Inagaki, K.| |
| Abstract | l-glutamate oxidase (LGOX) from Streptomyces sp. is a heterohexameric flavin enzyme that catalyzes the oxidative deamination of l-glutamate to form α-ketoglutarate with ammonia and hydrogen peroxide. LGOX shows strict substrate specificity for l-Glu. In addition, it is highly thermostable and pH stable. Because of these properties, LGOX is currently used as a biosensor for the trace determination of l-Glu in the food industry and clinical laboratories. The full-length cDNA is 2103 bp and is encoded by a single polypeptide chain consisting of 701 residues including subunits α-γ-β. The LGOX gene was heterologously expressed in Escherichia coli JM109. The LGOX precursor expressed in E. coli is a homodimer with weak enzymatic activity and becomes a heterohexamer upon activation by protease treatment. X-ray crystallography and docking studies of purified recombinant LGOX suggest that the Arg305 residue is a key residue for substrate recognition. Mutant analysis showed that Arg305 is essential for substrate recognition, as the activity toward l-Glu was greatly reduced and substrate specificity was changed in some enzymes. The functional analysis of R305E-LGOX, which is an l-Arg oxidase, revealed that R305E-LGOX can be used as a enzyme biosensor for l-Arg. |
| Keywords | l-glutamate oxidase biosensor substrate recognition X-ray crystallography modification of substrate specificity |
| Publication Title | Scientific Reports of the Faculty of Agriculture, Okayama University |
| Published Date | 2023-02-01 |
| Volume | volume112 |
| Start Page | 13 |
| End Page | 18 |
| ISSN | 2186-7755 |
| language | Japanese |
| File Version | publisher |
| Title Alternative | Studies on antitumor enzyme l-lysine α-oxidase from Trichoderma viride |
|---|---|
| FullText URL | srfa_111_007_014.pdf |
| Author | Saito, Masaya| Inagaki, Kenji| |
| Abstract | L-Lysine α-oxidase (LysOX) from Trichoderma viride is a homodimeric flavoenzyme that catalyzes the oxidative deamination of L-Lysine to produce α-keto-ε-aminocaproate with ammonia and hydrogen per-oxide. LysOX inhibited the growth of cancer cells but showed relatively low toxicity for normal cells. The full-length cDNA consists of 2,119 bp, and encodes a long N-terminal propeptide composed of 77 resi-dues (Met1-Arg77) and the mature protein (Ala78-Ile617). The LysOX gene was heterologously expressed in Streptomyces lividans TK24 or Escherichia coli SoluBL21. The enzymatic properties of the purified recombinant LysOX, such as substrate specificity, kinetic parameters and thermal stability, are the same as those of the native LysOX. The LysOX precursor (prLysOX) expressed in E. coli shows weak enzymatic activity and is activated by proteolytic processing. The crystal structure of prLysOX revealed that the propeptide of prLysOX indirectly changes the active site structure to inhibit enzyme activity. Moreover, the crystal structures of LysOX and its L-Lysine complex revealed that the hydrogen bonding network formed by Asp212, Asp315 and Ala440 with two water molecules is responsible for the recogni-tion of the ε-amino group of L-Lysine. In addition, a narrow substrate-binding site and acidic surface at the active site entrance both contribute to strict substrate specificity. Mutational analysis demonstrated that Asp212 and Asp315 are essential for substrate recognition, and the D212A/D315A LysOX prefers aromatic amino acids. Furthermore, the structural basis of the substrate specificity change has also been revealed by the structural analysis of the D212A/D315A LysOX and its substrate complexes. |
| Keywords | L-lysine α-oxidase antitumor enzyme substrate recognition X-ray crystallography enzyme activity regulation |
| Publication Title | Scientific Reports of the Faculty of Agriculture, Okayama University |
| Published Date | 2022-02-01 |
| Volume | volume111 |
| Start Page | 7 |
| End Page | 14 |
| ISSN | 2186-7755 |
| language | Japanese |
| File Version | publisher |
| NAID | 120007190707 |
| FullText URL | fulltext20210530_4.pdf |
|---|---|
| Author | Kitagawa, Masaki| Ito, Nanako| Matsumoto, Yuya| Saito, Masaya| Tamura, Takashi| Kusakabe, Hitoshi| Inagaki, Kenji| Imada, Katsumi| |
| Keywords | L-Lysine α-oxidase Crystal structure Precursor Substrate recognition |
| Published Date | 2021-12-31 |
| Publication Title | Journal of Structural Biology: X |
| Volume | volume5 |
| Publisher | Elsevier |
| Start Page | 100044 |
| ISSN | 25901524 |
| Content Type | Journal Article |
| language | English |
| OAI-PMH Set | 岡山大学 |
| Copyright Holders | © 2021 The Author(s). |
| File Version | publisher |
| PubMed ID | 33554108 |
| DOI | 10.1016/j.yjsbx.2021.100044 |
| Related Url | isVersionOf https://doi.org/10.1016/j.yjsbx.2021.100044 |
| FullText URL | fulltext.pdf |
|---|---|
| Author | Nemoto, Michiko| Iwaki, Sayako| Moriya, Hisao| Monden, Yuki| Tamura, Takashi| Inagaki, Kenji| Mayama, Shigeki| Obuse, Kiori| |
| Keywords | Biomineralization Diatom Silica Transcriptome Proteome |
| Note | This is a post-peer-review, pre-copyedit version of an article published in Marine Biotechnology. The final authenticated version is available online at: http://dx.doi.org/10.1007/s10126-020-09976-1.| |
| Published Date | 2020-06-03 |
| Publication Title | Marine Biotechnology |
| Publisher | Springer |
| ISSN | 1436-2228 |
| NCID | AA11357643 |
| Content Type | Journal Article |
| language | French |
| OAI-PMH Set | 岡山大学 |
| File Version | author |
| PubMed ID | 32488507 |
| DOI | 10.1007/s10126-020-09976-1 |
| Web of Science KeyUT | 000537432000001 |
| Related Url | isVersionOf https://doi.org/10.1007/s10126-020-09976-1 |
| Title Alternative | Recombinant expression and characterization of quinone-containing novel glycine oxidase from Marinomonas mediterranea |
|---|---|
| FullText URL | srfa_109_001_006.pdf |
| Author | Kajiyama, Yuki| Mizobata, Satsuki| Akaji, Shusaku| Nemoto, Michiko| Tamura, Takashi| Inagaki, Kenji| |
| Abstract | Novel glycine oxidase (GlyOX) from Marinomonas mediterranea depends on cysteine tryptophilquinone (CTQ) and catalyzes the oxidative deamination of glycine to produce a glyoxylate, ammonia, and hydrogen peroxide. M. mediterranea GlyOX genes (goxA and goxB) were cloned and recombinant GlyOX was heterologously expressed by E. coli. The purification of recombinant GlyOX was carried out by metal affinity and DEAE-Toyopearl 650M column chromatographies. M. mediterranea GlyOX was homotetramic with a molecular mass of 76kDa and showed optimum activity around 30°C and at pH 5.0, and stability below 50°C and between pH 5.0 to 9.0. M. mediterranea GlyOX shows a strict substrate specificity toward glycine, and the Michaelis constant for glycine was 0.5mM. M. mediterranea GlyOX could determine the quantity of glycine in human serum and human blood plasma with high sensitivity. This study revealed the catalytic and structural properties of M. mediterranea GlyOX with high substrate specificity. |
| Keywords | glycine oxidase Marinomonas mediterranea cysteine tryptophilquinone recombinant expression enzymatic glycine assay |
| Publication Title | Scientific Reports of the Faculty of Agriculture, Okayama University |
| Published Date | 2020-02-01 |
| Volume | volume109 |
| Start Page | 1 |
| End Page | 6 |
| ISSN | 2186-7755 |
| language | Japanese |
| File Version | publisher |
| Author | Tamura, T.| Ibi, T.| Inagaki, K.| Kubo, Y.| Okuda, K.| |
|---|---|
| Published Date | 2016-02-01 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume105 |
| Content Type | Departmental Bulletin Paper |
| Author | Tamura, Takashi| Tsunekawa, Naoki| Nemoto, Michiko| Inagaki, Kenji| Hirano, Toshiyuki| Sato, Fumitoshi| |
|---|---|
| Published Date | 2016-01-28 |
| Publication Title | Scientific reports |
| Volume | volume6 |
| Content Type | Journal Article |
| Author | Oshima, Kenshiro| Hattori, Masahira| Shimizu, Hitomi| Fukuda, Koji| Nemoto, Michiko| Inagaki, Kenji| Tamura, Takashi| |
|---|---|
| Published Date | 2015-07-09 |
| Publication Title | Genome Announcements |
| Volume | volume3 |
| Issue | issue4 |
| Content Type | Journal Article |
| Author | Nakai, Ryuichiro| Fujino, Shihoko| Utsumi, Tomohiro| Tamura, Takashi| Kusakabea, Hitoshi| Inagaki, Kenji| |
|---|---|
| Published Date | 2014-02-01 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume103 |
| Content Type | Departmental Bulletin Paper |
| Author | Kobayashi, Fumiaki| Aomine, Hiroki| Mizunashi, Wataru| Yu, Fujio| Tamura, Takashi| Inagaki, Kenji| |
|---|---|
| Published Date | 2012-02-01 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume101 |
| Content Type | Departmental Bulletin Paper |
| Author | Maemura, Tomomi| Uchitomi, Kumiko| Kusaka, Chika| Inagaki, Junko| Tamura, Takashi| Soda, Kenji| Inagaki, Kenji| |
|---|---|
| Published Date | 2011-02-01 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume100 |
| Content Type | Departmental Bulletin Paper |
| Author | Takahata, Muneaki| Tamura, Takashi| Abe, Katsumasa| Mihara, Hisaaki| Kurokawa, Suguru| Yamamoto, Yoshihiro| Nakano, Ryuhei| Esaki, Nobuyoshi| Inagaki, Kenji| |
|---|---|
| Published Date | 2008-06-26 |
| Publication Title | Journal of Biochemistry |
| Volume | volume143 |
| Issue | issue4 |
| Content Type | Journal Article |
| Author | Shinozaki, Mai| Yanagitani, Masahiko| Kaneda, Shouichirou| Kudou, Daizou| Endou, Yuuichi| Tamura, Takashi| Kuramitsu, Seiki| Inagaki, Kenji| |
|---|---|
| Published Date | 2010-02-01 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume99 |
| Issue | issue1 |
| Content Type | Departmental Bulletin Paper |
| Author | Fukuda, Koji| Tamura, Takashi| Inagaki, Kenji| |
|---|---|
| Published Date | 2010-02-01 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume99 |
| Issue | issue1 |
| Content Type | Departmental Bulletin Paper |
| Author | Yanagitani, Masahiko| Uemae, Satoshi| Shiraga, Tomoyuki| Tamura, Takashi| Inagaki, Kenji| |
|---|---|
| Published Date | 2009-02 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume98 |
| Issue | issue1 |
| Content Type | Departmental Bulletin Paper |
| Author | Tachibana, Akiko| Tamura, Takashi| Imada, Katsumi| Kinoshita, Miki| Namba, Keiichi| Tsutsumi, Noriko| Hashida, Miyoko| Sakaguchi, Hiromichi| Inagaki, Kenji| |
|---|---|
| Published Date | 2008-02 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume97 |
| Issue | issue1 |
| Content Type | Departmental Bulletin Paper |
| Author | Tanimoto, Yasuhide| Inagaki, Kenji| Tanaka, Hidehiko| |
|---|---|
| Published Date | 1996-02-01 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume85 |
| Issue | issue1 |
| Content Type | Departmental Bulletin Paper |
| Author | Seow, Teck Keong| Inagaki, Kenji| Tamura, Takashi| Tanaka, Hidehiko| |
|---|---|
| Published Date | 1997-02 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume86 |
| Issue | issue1 |
| Content Type | Departmental Bulletin Paper |
| Author | Inoue, Hiroyuki| Tamura, Takashi| Inagaki, Kenji| Tanaka, Hidehiko| |
|---|---|
| Published Date | 1998-02 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume87 |
| Issue | issue1 |
| Content Type | Departmental Bulletin Paper |
| Author | Inagaki, Kenji| Kawaguchi, Masakazu| Taguchi, Takaaki| Tamura, Takashi| Tanaka, Hidehiko| |
|---|---|
| Published Date | 1998-02 |
| Publication Title | 岡山大学農学部学術報告 |
| Volume | volume87 |
| Issue | issue1 |
| Content Type | Departmental Bulletin Paper |
