Optimized precursor to simplify assignment transfer between backbone resonances and stereospecifically labelled valine and leucine methyl groups: application to human Hsp90 N-terminal domain
Résumé
Methyl moieties are highly valuable probes for quantitative NMR studies of large proteins. Hence, their assignment is of the utmost interest to obtain information on both interactions and dynamics of proteins in solution. Here, we present the synthesis of a new precursor that allows connection of leucine and valine pro-S methyl moieties to backbone atoms by linear 13C-chains. This optimized 2H/13C-labelled acetolactate precursor can be combined with existing 13C/2H-alanine and isoleucine precursors in order to directly transfer backbone assignment to the corresponding methyl groups. Using this simple approach leucine and valine pro-S methyl groups can be assigned using a single sample without requiring correction of 1H/2H isotopic shifts on 13C resonances. The approach was demonstrated on the N-terminal domain of human HSP90, for which complete assignment of Ala-β, Ile-δ1, Leu-δ2, Met-ε, Thr-γ and Val-γ2 methyl groups was obtained.
Domaines
Biologie structurale [q-bio.BM]| Origine | Fichiers produits par l'(les) auteur(s) |
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