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j.bbagen.2010.06.003.pdf | 699.64 kB | Adobe PDF | 見る/開く |
タイトル: | Bulky high-mannose-type N-glycan blocks the taste-modifying activity of miraculin. |
著者: | Ito, Keisuke Sugawara, Taishi Koizumi, Ayako Nakajima, Ken-Ichiro Shimizu-Ibuka, Akiko Shiroishi, Mitsunori Asada, Hidetsugu https://orcid.org/0000-0001-6255-4728 (unconfirmed) Yurugi-Kobayashi, Takami Shimamura, Tatsuro Asakura, Tomiko Masuda, Katsuyoshi Ishiguro, Masaji Misaka, Takumi Iwata, So Kobayashi, Takuya Abe, Keiko |
著者名の別形: | 小林, 拓也 |
キーワード: | Miraculin Sweet protein Taste Yeast expression system N-glycan Steric hindrance |
発行日: | Sep-2010 |
出版者: | Elsevier B.V. |
誌名: | Biochimica et biophysica acta |
巻: | 1800 |
号: | 9 |
開始ページ: | 986 |
終了ページ: | 992 |
抄録: | BACKGROUND: Miraculin (MCL) is a taste-modifying protein that converts sourness into sweetness. The molecular mechanism underlying the taste-modifying action of MCL is unknown. METHODS: Here, a yeast expression system for MCL was constructed to accelerate analysis of its structure-function relationships. The Saccharomyces cerevisiae expression system has advantages as a high-throughput analysis system, but compared to other hosts it is characterized by a relatively low level of recombinant protein expression. To alleviate this weakness, in this study we optimized the codon usage and signal-sequence as the first step. Recombinant MCL (rMCL) was expressed and purified, and the sensory taste was analyzed. RESULTS: As a result, a 2 mg/l yield of rMCL was successfully obtained. Although sensory taste evaluation showed that rMCL was flat in taste under all the pH conditions employed, taste-modifying activity similar to that of native MCL was recovered after deglycosylation. Mutagenetic analysis revealed that the N-glycan attached to Asn42 was bulky in rMCL. CONCLUSIONS: The high-mannose-type N-glycan attached in yeast blocks the taste-modifying activity of rMCL. GENERAL SIGNIFICANCE: The bulky N-glycan attached to Asn42 may cause steric hindrance in the interaction between active residues and the sweet taste receptor hT1R2/hT1R3. |
著作権等: | © 2010 Elsevier B.V. This is not the published version. Please cite only the published version. この論文は出版社版でありません。引用の際には出版社版をご確認ご利用ください。 |
URI: | http://hdl.handle.net/2433/128855 |
DOI(出版社版): | 10.1016/j.bbagen.2010.06.003 |
PubMed ID: | 20542090 |
出現コレクション: | 学術雑誌掲載論文等 |
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