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Hokkaido University Collection of Scholarly and Academic Papers >
Graduate School of Agriculture / Faculty of Agriculture >
Peer-reviewed Journal Articles, etc >
Replacement of the Catalytic Nucleophile Aspartyl Residue of Dextran Glucosidase by Cysteine Sulfinate Enhances Transglycosylation Activity
| Title: | Replacement of the Catalytic Nucleophile Aspartyl Residue of Dextran Glucosidase by Cysteine Sulfinate Enhances Transglycosylation Activity |
| Authors: | Saburi, Wataru Browse this author →KAKEN DB | | Kobayashi, Momoko Browse this author | | Mori, Haruhide Browse this author →KAKEN DB | | Okuyama, Masayuki Browse this author →KAKEN DB | | Kimura, Atsuo Browse this author →KAKEN DB |
| Keywords: | Enzyme Catalysis | | Enzyme Kinetics | | Glycoside Hydrolases | | Site-specific Mutagenesis | | Protein Engineering | | Catalytic Nucleophile | | Cysteine Sulfinate | | Dextran Glucosidase | | Glycoside Hydrolase Family 13 | | Transglycosylation |
| Issue Date: | 1-Nov-2013 |
| Publisher: | American Society for Biochemistry and Molecular Biology (ASBMB) |
| Journal Title: | Journal of Biological Chemistry |
| Volume: | 288 |
| Issue: | 44 |
| Start Page: | 31670 |
| End Page: | 31677 |
| Publisher DOI: | 10.1074/jbc.M113.491449 |
| PMID: | 24052257 |
| Abstract: | Dextran glucosidase from Streptococcus mutans (SmDG) catalyzes the hydrolysis of an α-1,6-glucosidic linkage at the nonreducing end of isomaltooligosaccharides and dextran. This enzyme has an Asp-194 catalytic nucleophile and two catalytically unrelated Cys residues, Cys-129 and Cys-532. Cys-free SmDG was constructed by replacement with Ser (C129S/C532S (2CS), the activity of which was the same as that of the wild type, SmDG). The nucleophile mutant of 2CS was generated by substitution of Asp-194 with Cys (D194C-2CS). The hydrolytic activity of D194C-2CS was 8.1 × 10⁻⁴ % of 2CS. KI-associated oxidation of D194C-2CS increased the activity up to 0.27% of 2CS, which was 330 times higher than D194C-2CS. Peptide-mapping mass analysis of the oxidized D194C-2CS (Ox-D194C-2CS) revealed that Cys-194 was converted into cysteine sulfinate. Ox-D194C-2CS and 2CS shared the same properties (optimum pH, pI, and substrate specificity), whereas Ox-D194C-2CS had much higher transglucosylation activity than 2CS. This is the first study indicating that a more acidic nucleophile (-SOO−) enhances transglycosylation. The introduction of cysteine sulfinate as a catalytic nucleophile could be a novel approach to enhance transglycosylation. |
| Rights: | This research was originally published in The Journal of biological chemistry. Wataru Saburi, Momoko Kobayashi, Haruhide Mori, Masayuki Okuyama, and Atsuo Kimura. "Replacement of the Catalytic Nucleophile Aspartyl Residue of Dextran Glucosidase by Cysteine Sulfinate Enhances Transglycosylation Activity". J Biol Chem. 2013; Vol288(44):pp31670-pp31677. © the American Society for Biochemistry and Molecular Biology. |
| Type: | article |
| URI: | http://hdl.handle.net/2115/67776 |
| 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: 佐分利 亘
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