Lesovoy DM, Georgoulia PS, Diercks T, Matečko-Burmann I, Burmann BM, Bocharov EV, Bermel W, Orekhov VY
Angew. Chem. Int. Ed. Engl. - (-) - [2021-06-18; online 2021-06-18]
Dysregulation of post-translational modifications (PTMs) like phosphorylation is often involved in disease. NMR may elucidate exact loci and time courses of PTMs at atomic resolution and near-physiological condition, but requires signal assignment to individual atoms. Conventional NMR methods for this base on tedious global signal assignment that may often fail, as for large intrinsically disordered proteins (IDPs). We present a sensitive, robust alternative to rapidly obtain only the local assign-ment near affected signals, based on FOcused SpectroscopY (FOSY) experiments using selec-tive polarisation transfer (SPT). We prove its efficiency by identifying two phosphorylation sites of glycogen synthase kinase 3 beta (GSK3β) in human Tau40, an IDP of 441 residues, where the extreme spectral dispersion in FOSY revealed unprimed phosphorylation also of Ser409. FOSY may broadly benefit NMR studies of PTMs and other hotspots in IDPs, including sites involved in molecular interactions.
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