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A Split-Abl Kinase for Direct Activation in Cells.

Juan E Diaz1, Charles W Morgan1, Catherine E Minogue2

  • 1Department of Pharmaceutical Chemistry, University of California San Francisco, San Francisco, CA 94158, USA.

Cell Chemical Biology
|September 19, 2017
PubMed
Summary
This summary is machine-generated.

Researchers engineered a split-cAbl kinase (sKin-Abl) for controlled activation using rapamycin. This novel split-kinase technology enables precise temporal, dose, and spatial control over kinase activity, with broad applications.

Keywords:
ABL kinaseNeuCodephosphoproteomicsprotein kinaseselective kinase activationsplit-protein engineering

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Area of Science:

  • Molecular Biology
  • Biochemistry
  • Cell Biology

Background:

  • Understanding individual kinase functions requires tools for selective activation.
  • Current methods lack precise temporal, dose, and spatial control over kinase activity.

Purpose of the Study:

  • To engineer a split-cAbl kinase (sKin-Abl) for rapamycin-induced activation.
  • To demonstrate temporal, dose, and compartmentalization control of kinase activity.
  • To explore the broader applicability of the split-kinase strategy to other kinases.

Main Methods:

  • Empirical screening in mammalian cells to identify an optimal split site in the N lobe of cAbl.
  • Utilizing small-molecule-induced dimerization for kinase activity restoration.
  • Employing quantitative proteomics (NeuCode SILAC) to quantify phosphotyrosine (pTyr) modification.

Main Results:

  • Successfully engineered sKin-Abl, which is inactive until rapamycin-induced dimerization.
  • Demonstrated that the split site is transferable to Src Tyr kinase and AKT.
  • Identified 432 phosphotyrosine sites, including known Abl substrates and novel targets, using quantitative proteomics.

Conclusions:

  • The developed split-kinase technology provides precise control over kinase activation.
  • This technology is applicable to various kinases, including Tyr and Ser/Thr kinases.
  • Split-kinase systems offer a powerful tool for dissecting cellular kinase functions.