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Biomolecular NMR Assignments

, Volume 12, Issue 1, pp 149–154 | Cite as

1H, 15N, and 13C chemical shift assignments of the micelle immersed FAT C-terminal (FATC) domains of the human protein kinases ataxia-telangiectasia mutated (ATM) and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) fused to the B1 domain of streptococcal protein G (GB1)

  • Munirah S. Abd Rahim
  • Lisa A. M. Sommer
  • Anja Wacker
  • Martin Schaad
  • Sonja A. Dames
Article

Abstract

FAT C-terminal (FATC) is a circa 33 residue-long domain. It controls the kinase functionality in phosphatidylinositol-3 kinase-related kinases (PIKKs). Recent NMR- and CD-monitored interaction studies indicated that the FATC domains of all PIKKs can interact with membrane mimetics albeit with different preferences for membrane properties such as surface charge and curvature. Thus they may generally act as membrane anchoring unit. Here, we present the 1H, 15N, and 13C chemical shift assignments of the DPC micelle immersed FATC domains of the human PIKKs ataxia-telangiectasia mutated (ATM, residues 3024–3056) and DNA protein kinase catalytic subunit (DNA-PKcs, residues 4096–4128), both fused to the 56 residue long B1 domain of Streptococcal protein G (GB1). Each fusion protein is 100 amino acids long and contains in the linking region between the GB1 tag and the FATC region a thrombin (LVPRGS) and an enterokinase (DDDDK) protease site. The assignments pave the route for the detailed structural characterization of the membrane mimetic bound states, which will help to better understand the role of the proper cellular localization at membranes for the function and regulation of PIKKs. The chemical shift assignment of the GB1 tag is useful for NMR spectroscopists developing new experiments or using GB1 otherwise for case studies in the field of in-cell NMR spectroscopy or protein folding. Moreover it is often used as purification tag. Earlier we showed already that GB1 does not interact with membrane mimetics and thus does not disturb the NMR monitoring of membrane mimetic interactions of attached proteins.

Keywords

Ataxia telangiectasia mutated (ATM) DNA-dependent kinase catalytic subunit (DNA-PKcs) FATC Phosphatidylinositol-3 kinase-related kinases (PIKKs) B1 domain of Streptococcal protein G (GB1) Chemical shift assignment 

Notes

Acknowledgements

This work was supported by a grant from the German Research Foundation to S.A.D. (DA1183/3-1 and -2). S.A.D acknowledges further financial support from the Technische Universität München diversity and talent management office (Laura Bassi award) and the Helmholtz portfolio theme ‘metabolic dysfunction and common disease’ of the Helmholtz Zentrum München. M.S.A.R. is supported by a Ph. D. fellowship from the German academic exchange service (DAAD). Prof. Dr. Michael Sattler and Prof. Dr. Bernd Reif from the Technische Universität München (TUM) and the Helmholtz Zentrum München we thank very much for hosting our group and for sharing their facilities with us.

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Copyright information

© Springer Science+Business Media B.V., part of Springer Nature 2018

Authors and Affiliations

  1. 1.Department of Chemistry, Biomolecular NMR SpectroscopyTechnische Universität MünchenGarchingGermany
  2. 2.Roche Diagnostics GmbHCentralised and Point of Care SolutionsPenzbergGermany
  3. 3.Division of Radiopharmaceutical ChemistryGerman Cancer Research Center (DKFZ)HeidelbergGermany
  4. 4.Quintiles AGBaselSwitzerland
  5. 5.Institute of Structural BiologyHelmholtz Zentrum MünchenNeuherbergGermany

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