Abstract
Nuclear magnetic resonance (NMR) spectroscopy can be used as an analytical tool to investigate posttranslational modifications of protein. NMR is a valuable tool to map the interaction regions of protein partners. Here, we present protocols that have been developed in the course of our studies of the neuronal Tau protein. Tau is found aggregated in the neurons of Alzheimer’s disease patients. Development of the disease is accompanied by increased, abnormal phosphorylation and acetylation of Tau. We have used NMR to investigate how these posttranslational modifications of Tau affect the interactions with its partners. We present here detailed protocols of in vitro phosphorylation of Tau by recombinant kinase, ERK2, or kinase activity of rat brain extracts, and acetylation by recombinant Creb-binding protein (CBP) acetyltransferase. The analytical characterization of the modified Tau by NMR spectroscopy is additionally described.
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Acknowledgment
The NMR facilities were funded by the Région Nord, CNRS, Pasteur Institute of Lille, European Community (FEDER), French Research Ministry and the University of Sciences and Technologies of Lille I. We acknowledge support from the TGE RMN THC (FR-3050, France) and the Research Federation FRABio (Univ. Lille, CNRS, FR 3688, FRABio, “Structural & Functional Biochemistry of Biomolecular Assemblies”) for providing the scientific and technical environment. This study was supported by a grant from the LabEx (Laboratory of Excellence) DISTALZ (Development of Innovative Strategies for a Transdisciplinary approach to Alzheimer’s disease), and in part by the French government funding agency Agence Nationale de la Recherche TAF. S.P. and J.G. were partially supported on NIH R01-GM08157.
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Qi, H. et al. (2017). The Study of Posttranslational Modifications of Tau Protein by Nuclear Magnetic Resonance Spectroscopy: Phosphorylation of Tau Protein by ERK2 Recombinant Kinase and Rat Brain Extract, and Acetylation by Recombinant Creb-Binding Protein. In: Smet-Nocca, C. (eds) Tau Protein. Methods in Molecular Biology, vol 1523. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6598-4_11
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DOI: https://doi.org/10.1007/978-1-4939-6598-4_11
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