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Angew Chem Int Ed Engl. 2017 Sep 11;56(38):11409-11414. doi: 10.1002/anie.201704654. Epub 2017 Aug 7.

Dynamic Equilibrium of the Aurora A Kinase Activation Loop Revealed by Single-Molecule Spectroscopy.

Author information

1
National Heart & Lung Institute, SAF Building, Imperial College London, London, SW7 2AZ, UK.
2
Cancer Research UK Cancer Therapeutics Unit, The Institute of Cancer Research, 15 Cotswold Road, Sutton, Surrey, SM2 5NG, UK.

Abstract

The conformation of the activation loop (T-loop) of protein kinases underlies enzymatic activity and influences the binding of small-molecule inhibitors. By using single-molecule fluorescence spectroscopy, we have determined that phosphorylated Aurora A kinase is in dynamic equilibrium between a DFG-in-like active T-loop conformation and a DFG-out-like inactive conformation, and have measured the rate constants of interconversion. Addition of the Aurora A activating protein TPX2 shifts the equilibrium towards an active T-loop conformation whereas addition of the inhibitors MLN8054 and CD532 favors an inactive T-loop. We show that Aurora A binds TPX2 and MLN8054 simultaneously and provide a new model for kinase conformational behavior. Our approach will enable conformation-specific effects to be integrated into inhibitor discovery across the kinome, and we outline some immediate consequences for structure-based drug discovery.

KEYWORDS:

Aurora A; activation loop; drug design; kinases; single-molecule studies

PMID:
28700101
PMCID:
PMC5601181
DOI:
10.1002/anie.201704654
[Indexed for MEDLINE]
Free PMC Article

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