Abstract Obtaining sequence-specific assignments remains a major bottleneck in solution NMR investigations of supramolecular structure, dynamics and interactions. Here we demonstrate that resonance assignment of methyl probes in high molecular weight protein assemblies can be efficiently achieved by combining fast NMR experiments, residue-type-specific isotope-labeling and automated site-directed mutagenesis. The utility of this general and straightforward strategy is demonstrated through the characterization of intermolecular interactions involving a 468-kDa multimeric aminopeptidase, PhTET2.
Content Type Journal Article
Pages 1-8
DOI 10.1007/s10858-011-9513-5
Authors
Carlos Amero, Institut de Biologie Structurale Jean-Pierre Ebel, CNRS, Grenoble, France
M. Asunción Durá, Institut de Biologie Structurale Jean-Pierre Ebel, CNRS, Grenoble, France
Marjolaine Noirclerc-Savoye, Institut de Biologie Structurale Jean-Pierre Ebel, CEA, Grenoble, France
Arnaud Perollier, Institut de Biologie Structurale Jean-Pierre Ebel, CNRS, Grenoble, France
Benoit Gallet, Institut de Biologie Structurale Jean-Pierre Ebel, CEA, Grenoble, France
Michael J. Plevin, Institut de Biologie Structurale Jean-Pierre Ebel, CNRS, Grenoble, France
Thierry Vernet, Institut de Biologie Structurale Jean-Pierre Ebel, CNRS, Grenoble, France
Bruno Franzetti, Institut de Biologie Structurale Jean-Pierre Ebel, CNRS, Grenoble, France
A systematic mutagenesis-driven strategy for site-resolved NMR studies of supramolecular assemblies.
A systematic mutagenesis-driven strategy for site-resolved NMR studies of supramolecular assemblies.
A systematic mutagenesis-driven strategy for site-resolved NMR studies of supramolecular assemblies.
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Obtaining sequence-specific assignments remains a major bottleneck in solution NMR investigations of supramolecular structure, dynamics and interactions. Here we demonstrate that resonance...
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Site-resolved measurement of water-protein interactions by solution NMR.
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Nat Struct Mol Biol. 2011 Jan 2;
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The interactions of biological macromolecules with water are fundamental to their structure, dynamics and function. Historically, characterization of the location and residence times of hydration waters of proteins in solution has been quite difficult. Confining proteins within the nanoscale interior of a reverse micelle slows water...
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[NMR paper] Site-directed mutagenesis, fluorescence, and two-dimensional NMR studies on microenvi
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A systematic mutagenesis-driven strategy for site-resolved NMR studies of supramolecular assemblies
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