3D structure determination of a protein in living cells using paramagnetic NMR spectroscopy.

Determining the three-dimensional structure of a protein in living cells remains particularly challenging. We demonstrated that the integration of site-specific tagging proteins and GPS-Rosetta calculations provides a fast and effective way of determining the structures of proteins in living cells, and in principle the interactions and dynamics of protein-ligand complexes.

[1]  David Baker,et al.  Protein structure determination from pseudocontact shifts using ROSETTA. , 2012, Journal of molecular biology.

[2]  Thomas Huber,et al.  Pseudocontact Shift-Driven Iterative Resampling for 3D Structure Determinations of Large Proteins. , 2016, Journal of molecular biology.

[3]  Yutaka Ito,et al.  Cellular structural biology. , 2010, Current opinion in structural biology.

[4]  Roberta Pierattelli,et al.  In-cell 13C NMR spectroscopy for the study of intrinsically disordered proteins , 2014, Nature Protocols.

[5]  J. Ferrell,et al.  Investigating macromolecules inside cultured and injected cells by in-cell NMR spectroscopy , 2006, Nature Protocols.

[6]  G. Otting,et al.  Paramagnetic labelling of proteins and oligonucleotides for NMR , 2010, Journal of biomolecular NMR.

[7]  D. Cowburn,et al.  In-cell NMR for protein-protein interactions (STINT-NMR) , 2006, Nature Protocols.

[8]  Gerhard Wagner,et al.  Quantitative NMR analysis of the protein G B1 domain in Xenopus laevis egg extracts and intact oocytes , 2006, Proceedings of the National Academy of Sciences.

[9]  Jens Meiler,et al.  Expanding the utility of NMR restraints with paramagnetic compounds: background and practical aspects. , 2011, Progress in nuclear magnetic resonance spectroscopy.

[10]  Ivano Bertini,et al.  Magnetic susceptibility in paramagnetic NMR , 2002 .

[11]  Zeting Zhang,et al.  NMR studies of protein folding and binding in cells and cell-like environments. , 2015, Current opinion in structural biology.

[12]  G. Jeschke,et al.  Gd(III)-PyMTA label is suitable for in-cell EPR. , 2014, Journal of the American Chemical Society.

[13]  Hidekazu Hiroaki,et al.  High-resolution multi-dimensional NMR spectroscopy of proteins in human cells , 2009, Nature.

[14]  G. Clore,et al.  Theory, practice, and applications of paramagnetic relaxation enhancement for the characterization of transient low-population states of biological macromolecules and their complexes. , 2009, Chemical reviews.

[15]  Lucia Banci,et al.  A Unique Tool for Cellular Structural Biology: In-cell NMR* , 2015, The Journal of Biological Chemistry.

[16]  Benjamin J. Wylie,et al.  Crystal polymorphism of protein GB1 examined by solid-state NMR spectroscopy and X-ray diffraction. , 2007, The journal of physical chemistry. B.

[17]  Wei-Min Liu,et al.  The application of paramagnetic lanthanoid ions in NMR spectroscopy on proteins , 2014 .

[18]  Gottfried Otting,et al.  Prospects for lanthanides in structural biology by NMR , 2008, Journal of biomolecular NMR.

[19]  M. Ubbink,et al.  Structure determination of protein-protein complexes with long-range anisotropic paramagnetic NMR restraints. , 2014, Current opinion in structural biology.

[20]  P. Ortiz de Montellano,et al.  Methyl groups as probes for proteins and complexes in in-cell NMR experiments. , 2004, Journal of the American Chemical Society.

[21]  Conggang Li,et al.  Direct observation of Ca(2+) -induced calmodulin conformational transitions in intact Xenopus laevis oocytes by (19) F NMR spectroscopy. , 2015, Angewandte Chemie.

[22]  Thomas Huber,et al.  Three-dimensional protein fold determination from backbone amide pseudocontact shifts generated by lanthanide tags at multiple sites. , 2013, Structure.

[23]  D. Burz,et al.  Structure of proteins in eukaryotic compartments. , 2012, Journal of the American Chemical Society.

[24]  G. Otting,et al.  Capturing Conformational States in Proteins Using Sparse Paramagnetic NMR Data , 2015, PloS one.

[25]  G. Otting,et al.  Numbat: an interactive software tool for fitting Δχ-tensors to molecular coordinates using pseudocontact shifts , 2008, Journal of biomolecular NMR.

[26]  Y. Yang,et al.  Site-specific tagging proteins via a rigid, stable and short thiolether tether for paramagnetic spectroscopic analysis. , 2015, Chemical communications.

[27]  Y. Yang,et al.  Thiol-ene reaction: a versatile tool in site-specific labelling of proteins with chemically inert tags for paramagnetic NMR. , 2012, Chemical communications.

[28]  V. Dötsch,et al.  High-resolution macromolecular NMR spectroscopy inside living cells. , 2001, Journal of the American Chemical Society.

[29]  V. Dötsch,et al.  In-cell NMR and EPR spectroscopy of biomacromolecules. , 2014, Angewandte Chemie.

[30]  I. Bertini,et al.  Atomic-resolution monitoring of protein maturation in live human cells by NMR , 2013, Nature chemical biology.

[31]  Y. Yang,et al.  Mn(II) tags for DEER distance measurements in proteins via C-S attachment. , 2015, Dalton transactions.

[32]  G. Otting,et al.  A dipicolinic acid tag for rigid lanthanide tagging of proteins and paramagnetic NMR spectroscopy. , 2008, Journal of the American Chemical Society.

[33]  M. Shirakawa,et al.  In-cell NMR spectroscopy of proteins inside Xenopus laevis oocytes , 2006, Journal of biomolecular NMR.

[34]  Guifang Wang,et al.  Protein (19)F NMR in Escherichia coli. , 2010, Journal of the American Chemical Society.

[35]  Philipp Selenko,et al.  Structural disorder of monomeric α-synuclein persists in mammalian cells , 2016, Nature.

[36]  Zeting Zhang,et al.  (19) F NMR spectroscopy as a probe of cytoplasmic viscosity and weak protein interactions in living cells. , 2013, Chemistry.

[37]  Harald Schwalbe,et al.  Protein alignment by a coexpressed lanthanide-binding tag for the measurement of residual dipolar couplings. , 2003, Journal of the American Chemical Society.

[38]  C Kooperberg,et al.  Assembly of protein tertiary structures from fragments with similar local sequences using simulated annealing and Bayesian scoring functions. , 1997, Journal of molecular biology.

[39]  M. Shirakawa,et al.  High-resolution heteronuclear multidimensional NMR of proteins in living insect cells using a baculovirus protein expression system. , 2013, Journal of the American Chemical Society.

[40]  Guido Pintacuda,et al.  NMR structure determination of protein-ligand complexes by lanthanide labeling. , 2007, Accounts of chemical research.

[41]  Masaki Mishima,et al.  Protein structure determination in living cells by in-cell NMR spectroscopy , 2009, Nature.

[42]  Y. Yang,et al.  Site-specific labeling of proteins with a chemically stable, high-affinity tag for protein study. , 2013, Chemistry.

[43]  D. Goldfarb,et al.  Probing protein conformation in cells by EPR distance measurements using Gd3+ spin labeling. , 2014, Journal of the American Chemical Society.

[44]  Jeffrey M. Macdonald,et al.  Differential dynamical effects of macromolecular crowding on an intrinsically disordered protein and a globular protein: implications for in-cell NMR spectroscopy. , 2008, Journal of the American Chemical Society.