Engineered allosteric activation of kinases in living cells

Andrei V. Karginov, Feng Ding, Pradeep Kota, Nikolay V. Dokholyan, Klaus M. Hahn

Research output: Contribution to journalArticle

118 Citations (Scopus)

Abstract

Studies of cellular and tissue dynamics benefit greatly from tools that can control protein activity with specificity and precise timing in living systems. Here we describe an approach to confer allosteric regulation specifically on the catalytic activity of protein kinases. A highly conserved portion of the kinase catalytic domain is modified with a small protein insert that inactivates catalytic activity but does not affect other protein functions (Fig. 1a). Catalytic activity is restored by addition of rapamycin or non-immunosuppresive rapamycin analogs. Molecular modeling and mutagenesis indicate that the protein insert reduces activity by increasing the flexibility of the catalytic domain. Drug binding restores activity by increasing rigidity. We demonstrate the approach by specifically activating focal adhesion kinase (FAK) within minutes in living cells and show that FAK is involved in the regulation of membrane dynamics. Successful regulation of Src and p38 by insertion of the rapamycin-responsive element at the same conserved site used in FAK suggests that our strategy will be applicable to other kinases.

Original languageEnglish (US)
Pages (from-to)743-747
Number of pages5
JournalNature Biotechnology
Volume28
Issue number7
DOIs
StatePublished - Jul 2010

Fingerprint

Focal Adhesion Protein-Tyrosine Kinases
Phosphotransferases
Sirolimus
Chemical activation
Cells
Proteins
Catalyst activity
Adhesion
Catalytic Domain
Allosteric Regulation
Mutagenesis
Molecular modeling
Rigidity
Protein Kinases
Tissue
Membranes
Pharmaceutical Preparations

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology
  • Molecular Medicine
  • Biomedical Engineering

Cite this

Karginov, Andrei V. ; Ding, Feng ; Kota, Pradeep ; Dokholyan, Nikolay V. ; Hahn, Klaus M. / Engineered allosteric activation of kinases in living cells. In: Nature Biotechnology. 2010 ; Vol. 28, No. 7. pp. 743-747.
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Engineered allosteric activation of kinases in living cells. / Karginov, Andrei V.; Ding, Feng; Kota, Pradeep; Dokholyan, Nikolay V.; Hahn, Klaus M.

In: Nature Biotechnology, Vol. 28, No. 7, 07.2010, p. 743-747.

Research output: Contribution to journalArticle

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