The structure and substrate specificity of human Cdk12/Cyclin K

Nat Commun. 2014 Mar 24;5:3505. doi: 10.1038/ncomms4505.

Abstract

Phosphorylation of the RNA polymerase II C-terminal domain (CTD) by cyclin-dependent kinases is important for productive transcription. Here we determine the crystal structure of Cdk12/CycK and analyse its requirements for substrate recognition. Active Cdk12/CycK is arranged in an open conformation similar to that of Cdk9/CycT but different from those of cell cycle kinases. Cdk12 contains a C-terminal extension that folds onto the N- and C-terminal lobes thereby contacting the ATP ribose. The interaction is mediated by an HE motif followed by a polybasic cluster that is conserved in transcriptional CDKs. Cdk12/CycK showed the highest activity on a CTD substrate prephosphorylated at position Ser7, whereas the common Lys7 substitution was not recognized. Flavopiridol is most potent towards Cdk12 but was still 10-fold more potent towards Cdk9. T-loop phosphorylation of Cdk12 required coexpression with a Cdk-activating kinase. These results suggest the regulation of Pol II elongation by a relay of transcriptionally active CTD kinases.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blotting, Western
  • Crystallization
  • Cyclin-Dependent Kinases / chemistry*
  • Cyclin-Dependent Kinases / metabolism
  • Cyclins / chemistry*
  • Cyclins / metabolism
  • Enzyme-Linked Immunosorbent Assay
  • HeLa Cells
  • Humans
  • Immunoprecipitation
  • Mass Spectrometry
  • Models, Molecular*
  • Multiprotein Complexes / chemistry*
  • Multiprotein Complexes / metabolism
  • Protein Conformation
  • Substrate Specificity

Substances

  • CCNK protein, human
  • Cyclins
  • Multiprotein Complexes
  • CDK12 protein, human
  • Cyclin-Dependent Kinases

Associated data

  • PDB/4NST