Articles by Clément Danis in JoVE
Nuclear Magnetic Resonance Spectroscopy for the Identification of Multiple Phosphorylations of Intrinsically Disordered Proteins Clément Danis*1,2, Clément Despres*1, Luiza M. Bessa1, Idir Malki1, Hamida Merzougui1, Isabelle Huvent1, Haoling Qi1, Guy Lippens1, François-Xavier Cantrelle1, Robert Schneider1, Xavier Hanoulle1, Caroline Smet-Nocca1, Isabelle Landrieu1 1UMR8576, CNRS, Lille University, 2UMR-S1172, INSERM CNRS, Lille University We describe here a method to identify multiple phosphorylations of an intrinsically disordered protein by Nuclear Magnetic Resonance Spectroscopy (NMR), using Tau protein as a case study. Recombinant Tau is isotopically enriched and modified in vitro by a kinase prior to data acquisition and analysis.
Other articles by Clément Danis on PubMed
Cell-free Expression, Purification, and Membrane Reconstitution for NMR Studies of the Nonstructural Protein 4B from Hepatitis C Virus Journal of Biomolecular NMR. Jun, 2016 | Pubmed ID: 27233794 We describe the expression of the hepatitis C virus nonstructural protein 4B (NS4B), which is an integral membrane protein, in a wheat germ cell-free system, the subsequent purification and characterization of NS4B and its insertion into proteoliposomes in amounts sufficient for multidimensional solid-state NMR spectroscopy. First spectra of the isotopically [(2)H,(13)C,(15)N]-labeled protein are shown to yield narrow (13)C resonance lines and a proper, predominantly α-helical fold. Clean residue-selective leucine, isoleucine and threonine-labeling is demonstrated. These results evidence the suitability of the wheat germ-produced integral membrane protein NS4B for solid-state NMR. Still, the proton linewidth under fast magic angle spinning is broader than expected for a perfect sample and possible causes are discussed.