A biophysical and structural analysis of the interaction of BLNK with 14-3-3 proteins

Lorenzo Soini, Seppe Leysen, Jeremy Davis, Christian Ottmann (Corresponding author)

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

B-cell linker protein (BLNK) is an adaptor protein that orchestrates signalling downstream of B-cell receptors. It has been reported to undergo proteasomal degradation upon binding to 14-3-3 proteins. Here, we report the first biophysical and structural study of this protein-protein interaction (PPI). Specifically, we investigated the binding of mono- and di- phosphorylated BLNK peptides to 14-3-3 using fluorescent polarization (FP) and isothermal titration calorimetry assays (ITC). Our results suggest that BLNK interacts with 14-3-3 according to the gatekeeper model, where HPK1 mediated phosphorylation of Thr152 (pT152) allows BLNK anchoring to 14-3-3, and an additional phosphorylation of Ser285 (pS285) by AKT, then further improves the affinity. Finally, we have also solved a crystal structure of the BLNKpT152 peptide bound to 14-3-3σ. These findings could serve as important tool for compound discovery programs aiming to modulate this interaction with 14-3-3.

Original languageEnglish
Article number107662
Number of pages6
JournalJournal of Structural Biology
Volume212
Issue number3
DOIs
Publication statusPublished - 1 Dec 2020

Keywords

  • Adaptor proteins 14-3-3 and BLNK
  • Gatekeeper
  • Isothermal titration calorimetry
  • Phosphorylation
  • X-ray protein crystallography

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