Structural mechanism of synergistic activation of Aurora kinase B/C by phosphorylated INCENP

Kamal R. Abdul Azeez,Channing Yu,Sneha Chatterjee,Jonathan M. Elkins,Frank Sobott,Todd R. Golub

doi:10.1038/s41467-019-11085-0

Abstract

Aurora kinases B and C (AURKB/AURKC) are activated by binding to the C-terminal domain of INCENP. Full activation requires phosphorylation of two serine residues of INCENP that are conserved through evolution, although the mechanism of this activation has not been explained. Here we present crystal structures of the fully active complex of AURKC bound to INCENP, consisting of phosphorylated, activated, AURKC and INCENP phosphorylated on its TSS motif, revealing the structural and biochemical mechanism of synergistic activation of AURKC:INCENP. The structures show that TSS motif phosphorylation stabilises the kinase activation loop of AURKC. The TSS motif phosphorylations alter the substrate-binding surface consistent with a mechanism of altered kinase substrate selectivity and stabilisation of the protein complex against unfolding. We also analyse the binding of the most specific available AURKB inhibitor, BRD-7880, and demonstrate that the well-known Aurora kinase inhibitor VX-680 disrupts binding of the phosphorylated INCENP TSS motif.

Highlights

Aurora kinases B and C (AURKB/AURKC) are activated by binding to the C-terminal domain of inner centromere protein (INCENP)
This fits with a mechanism where INCENP pS893/pS894 alters the substrate selectivity of AURKB/AURKC towards itself, enabling autoactivation by phosphorylation in trans
The structure of partially active AURKB:INCENP contains an activation loop domain swap;[15]. This may be a mechanism for phosphorylation of non-consensus sites[18], and it is possible that phosphorylation of the INCENP TSS motif by AURKB proceeds in trans, while INCENP is bound to an interacting kinase domain

Summary

Introduction

Aurora kinases B and C (AURKB/AURKC) are activated by binding to the C-terminal domain of INCENP. Previous crystal structures of human or X. laevis AURKB bound to INCENP revealed an extensive interaction of AURKB with the C-terminal IN-box region[14,15] These previous structures were of partially active AURKB: INCENP complexes lacking a phosphorylated TSS motif and the mechanistic role of this evolutionarily conserved motif has not been explained. To understand better the molecular mechanisms of regulation of AURKB and AURKC by INCENP we determined structures of fully active (phosphorylated) human AURKC bound to the phosphorylated C-terminal IN-box section of human INCENP (residues 835–903) These structures demonstrate the structural changes upon activation and the roles of essential conserved residues. We present crystallographic and biophysical characterisation of a potent and extremely selective AURKB and AURKC inhibitor[16]

Methods

Results

Conclusion