NMR spectroscopy is as a versatile emerging tool to study posttranslational modifications (PTMs) beyond the ability to detect/quantify PTM sites. The same sample can be employed to map structural changes and functional interactions with other molecules. We use NMR to study at the molecular level acetylation and O-GlcNAcylation (O-ß-linked N-acetylglucosaminylation) of the neuronal Tau protein, two PTMs that are involved either in the regulation of phosphorylation or in the control of Tau function/aggregation. We develop strategies to monitor these PTMs, i.e. detect them and identify/quantify the modification sites in a full protein context. An in vitro system of acetylation involving a functional domain of CBP/p300 (CREB-binding protein/p300) recombinantly expressed in E. coli has been set up to achieve Tau acetylation on the NMR scale. Impact on diverse phosphorylation patterns (see phospho-Tau section) as well as interactions with other proteins or small-molecule compounds that could interfere with the aggregation processes are further investigated both at the structural and functional levels. On the other hand, it has been shown that O-GlcNAc linkage on Ser/Thr residues can modulate site-specific phosphorylation of Tau and prevent from aggregation in a phosphorylation-independent manner. We want to characterize Tau O-GlcNAc patterns generated by O-GlcNAc transferase (OGT) activities and investigate their impact on Tau function. NMR can also be used to directly map the effect of O-GlcNAcylation on specific phosphorylation patterns obtained from different kinases.
Contact: caroline.smet-nocca@univ-lille.fr
