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Effect of the Extra N-terminal Methionine Residue on the Stability and Folding of Recombinant α-Lactalbumin Expressed in Escherichia coli
https://ir.soken.ac.jp/records/4291
https://ir.soken.ac.jp/records/429139bfdb73-1d80-4008-a455-b5f0d5a48703
| Item type | 学術雑誌論文 / Journal Article(1) | |||||||||||
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| 公開日 | 2014-03-13 | |||||||||||
| タイトル | ||||||||||||
| タイトル | Effect of the Extra N-terminal Methionine Residue on the Stability and Folding of Recombinant α-Lactalbumin Expressed in Escherichia coli | |||||||||||
| タイトル | ||||||||||||
| タイトル | Effect of the Extra N-terminal Methionine Residue on the Stability and Folding of Recombinant α-Lactalbumin Expressed in Escherichia coli | |||||||||||
| 言語 | en | |||||||||||
| 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 | |||||||||||
| アクセス権 | ||||||||||||
| アクセス権 | metadata only access | |||||||||||
| アクセス権URI | http://purl.org/coar/access_right/c_14cb | |||||||||||
| 著者 |
CHAUDHURI, Tapan K
× CHAUDHURI, Tapan K
× KUWAJIMA, Kunihiro
× et, al.
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| 著者別名 |
桑島, 邦博
× 桑島, 邦博
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| 抄録 | ||||||||||||
| 内容記述タイプ | Abstract | |||||||||||
| 内容記述 | The structure, stability, and unfolding-refolding kinetics of Escherichia coli-expressed recombinant goat α-lactalbumin were studied by circular dichroism spectroscopy, X-ray crystallography, and stopped-flow measurements, and the results were compared with those of the authentic protein prepared from goat milk. The electric properties of the two proteins were also studied by gel electrophoresis and ion-exchange chromatography. Although the overall structures of the authentic and recombinant proteins are the same, the extra methionine residue at the N terminus of the recombinant protein remarkably affects the native-state stability and the electric properties. The native state of the recombinant protein was 3.5 kcal/mol less stable than the authentic protein, and the recombinant protein was more negatively charged than the authentic one. The recombinant protein unfolded 5.7 times faster than the authentic one, although there were no significant differences in the refolding rates of the two proteins. The destabilization of the recombinant protein can be fully interpreted in terms of the increased unfolding rate of the protein, indicating that the N-terminal region remains unorganized in the transition state of refolding, and hence is not involved in the folding initiation site of the protein. A comparison of the X-ray structures of recombinant α-lactalbumin determined here with that of the authentic protein shows that the structural differences between the proteins are confined to the N-terminal region. Theoretical considerations for the differences in the conformational and solvation free energies between the proteins show that the destabilization of the recombinant protein is primarily due to excess conformational entropy of the N-terminal methionine residue in the unfolded state, and also due to less exposure of hydrophobic surface on unfolding. The results suggest that when the N-terminal region of a protein has a rigid structure, expression of the protein by E. coli, which adds the extra methionine residue, destabilizes the native state through a conformational entropy effect. It also shows that differences in the electrostatic interactions of the N-terminal amino group with the side-chain atoms of Thr38, Asp37, and Asp83 bring about a difference in the pKa value of the N-terminal amino group between the proteins, resulting in a greater negative net charge of the recombinant protein at neutral pH. | |||||||||||
| 書誌情報 |
Journal of Molecular Biology en : Journal of Molecular Biology 巻 285, 号 3, p. 1179-1194, 発行日 1999-01-22 |
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| 出版者 | ||||||||||||
| 出版者 | Elsevier | |||||||||||
| ISSN | ||||||||||||
| 収録物識別子タイプ | ISSN | |||||||||||
| 収録物識別子 | 0022-2836 | |||||||||||
| DOI | ||||||||||||
| 識別子タイプ | DOI | |||||||||||
| 関連識別子 | https://doi.org/10.1006/jmbi.1998.2362 | |||||||||||
| 関連名称 | 10.1006/jmbi.1998.2362 | |||||||||||
| 権利 | ||||||||||||
| 権利情報 | © 1999 Academic Press | |||||||||||
