2006年11月
In-cell NMR spectroscopy of proteins inside Xenopus laevis oocytes
JOURNAL OF BIOMOLECULAR NMR
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- 巻
- 36
- 号
- 3
- 開始ページ
- 179
- 終了ページ
- 188
- 記述言語
- 英語
- 掲載種別
- 研究論文(学術雑誌)
- DOI
- 10.1007/s10858-006-9079-9
- 出版者・発行元
- SPRINGER
In-cell NMR is an application of solution NMR that enables the investigation of protein conformations inside living cells. We have measured in-cell NMR spectra in oocytes from the African clawed frog Xenopus laevis. N-15-labeled ubiquitin, its derivatives and calmodulin were injected into Xenopus oocytes and two-dimensional H-1-N-15 correlation spectra of the proteins were obtained. While the spectrum of wild-type ubiquitin in oocytes had rather fewer cross-peaks compared to its in vitro spectrum, ubiquitin derivatives that are presumably unable to bind to ubiquitin-interacting proteins gave a markedly larger number of cross-peaks. This observation suggests that protein-protein interactions between ubiquitin and ubiquitin-interacting proteins may cause NMR signal broadening, and hence spoil the quality of the in-cell HSQC spectra. In addition, we observed the maturation of ubiquitin precursor derivative in living oocytes using the in-cell NMR technique. This process was partly inhibited by pre-addition of ubiquitin aldehyde, a specific inhibitor for ubiquitin C-terminal hydrolase (UCH). Our work demonstrates the potential usefulness of in-cell NMR with Xenopus oocytes for the investigation of protein conformations and functions under intracellular environmental conditions.
- リンク情報
- ID情報
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- DOI : 10.1007/s10858-006-9079-9
- ISSN : 0925-2738
- eISSN : 1573-5001
- Web of Science ID : WOS:000241167500004