Structure and inhibitor specificity of the PCTAIRE-family kinase CDK16.

Sarah E Dixon-Clarke,Tobias Krojer,Alex N Bullock,Saifeldin N Shehata,Timothy D Sharpe,Kei Sakamoto,Frank Von Delft

doi:10.1042/bcj20160941

Abstract

CDK16 (also known as PCTAIRE1 or PCTK1) is an atypical member of the cyclin-dependent kinase (CDK) family that has emerged as a key regulator of neurite outgrowth, vesicle trafficking and cancer cell proliferation. CDK16 is activated through binding to cyclin Y via a phosphorylation-dependent 14-3-3 interaction and has a unique consensus substrate phosphorylation motif compared with conventional CDKs. To elucidate the structure and inhibitor-binding properties of this atypical CDK, we screened the CDK16 kinase domain against different inhibitor libraries and determined the co-structures of identified hits. We discovered that the ATP-binding pocket of CDK16 can accommodate both type I and type II kinase inhibitors. The most potent CDK16 inhibitors revealed by cell-free and cell-based assays were the multitargeted cancer drugs dabrafenib and rebastinib. An inactive DFG-out binding conformation was confirmed by the first crystal structures of CDK16 in separate complexes with the inhibitors indirubin E804 and rebastinib, respectively. The structures revealed considerable conformational plasticity, suggesting that the isolated CDK16 kinase domain was relatively unstable in the absence of a cyclin partner. The unusual structural features and chemical scaffolds identified here hold promise for the development of more selective CDK16 inhibitors and provide opportunity to better characterise the role of CDK16 and its related CDK family members in various physiological and pathological contexts.

Highlights

cyclin-dependent kinase 16 (CDK16) is a newly recognised member of the CDK protein kinase family [1,2] that is activated upon binding to the membrane-associated protein cyclin Y (CCNY) or its homologue cyclin Y-like 1 (CCNYL1) [3,4,5]
To identify protein constructs suitable for structural studies, human CDK16 was recombinantly expressed in E. coli with a variety of N- and C-terminal truncations
The position of CDK16 (PCTAIRE1) within the CDK family was uncertain: the identity of its cyclin partner remained elusive and its existence was cast in doubt [7]

Summary

Introduction

CDK16 is a newly recognised member of the CDK protein kinase family [1,2] that is activated upon binding to the membrane-associated protein cyclin Y (CCNY) or its homologue cyclin Y-like 1 (CCNYL1) [3,4,5]. These interactions are facilitated by a ‘PCTAIRE’ sequence motif in the kinase αC helix that is conserved in CDK17 and CDK18, but diverged from the classical ‘PSTAIRE’ motif found in CDK2 [6]. CDK16 has been implicated in other diverse processes, including vesicle trafficking [11,12], glucose homeostasis [13,14] and muscle differentiation [15]

Methods

Results

Conclusion