Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
J Biomol NMR
2006 Nov 01;363:179-88. doi: 10.1007/s10858-006-9079-9.
Show Gene links
Show Anatomy links
In-cell NMR spectroscopy of proteins inside Xenopus laevis oocytes.
Sakai T, Tochio H, Tenno T, Ito Y, Kokubo T, Hiroaki H, Shirakawa M.
???displayArticle.abstract???
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. (15)N-labeled ubiquitin, its derivatives and calmodulin were injected into Xenopus oocytes and two-dimensional (1)H-(15)N 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.
Bryant,
Protein dynamics in living cells.
2005, Pubmed
Bryant,
Protein dynamics in living cells.
2005,
Pubmed Burz,
Mapping structural interactions using in-cell NMR spectroscopy (STINT-NMR).
2006,
Pubmed Dedmon,
FlgM gains structure in living cells.
2002,
Pubmed Delaglio,
NMRPipe: a multidimensional spectral processing system based on UNIX pipes.
1995,
Pubmed Dumont,
Oogenesis in Xenopus laevis (Daudin). I. Stages of oocyte development in laboratory maintained animals.
1972,
Pubmed
,
Xenbase Hayashi,
An expression system of rat calmodulin using T7 phage promoter in Escherichia coli.
1998,
Pubmed Hicke,
Ubiquitin-binding domains.
2005,
Pubmed Hilgemann,
Giant membrane patches: improvements and applications.
1998,
Pubmed
,
Xenbase Hubbard,
Nuclear magnetic resonance spectroscopy reveals the functional state of the signalling protein CheY in vivo in Escherichia coli.
2003,
Pubmed Johnston,
Structural basis for the specificity of ubiquitin C-terminal hydrolases.
1999,
Pubmed Ohno,
Structure of the UBA domain of Dsk2p in complex with ubiquitin molecular determinants for ubiquitin recognition.
2005,
Pubmed Partington,
Human globin gene transcription in injected Xenopus oocytes: enhancement by sodium butyrate.
1984,
Pubmed
,
Xenbase Ranjani Varadan,
Using NMR spectroscopy to monitor ubiquitin chain conformation and interactions with ubiquitin-binding domains.
2005,
Pubmed Reardon,
Multidimensional NMR spectroscopy for protein characterization and assignment inside cells.
2005,
Pubmed Sehy,
Apparent diffusion of water, ions, and small molecules in the Xenopus oocyte is consistent with Brownian displacement.
2002,
Pubmed
,
Xenbase Serber,
Methyl groups as probes for proteins and complexes in in-cell NMR experiments.
2004,
Pubmed Serber,
High-resolution macromolecular NMR spectroscopy inside living cells.
2001,
Pubmed Serber,
In-cell NMR spectroscopy.
2001,
Pubmed Serber,
In-cell NMR spectroscopy.
2005,
Pubmed Serber,
Evaluation of parameters critical to observing proteins inside living Escherichia coli by in-cell NMR spectroscopy.
2001,
Pubmed Tenno,
Structural basis for distinct roles of Lys63- and Lys48-linked polyubiquitin chains.
2004,
Pubmed Zhang,
Molecular mechanisms of calmodulin's functional versatility.
1998,
Pubmed
