A Photoaffinity Displacement Assay and Probes to Study the Cyclin-Dependent Kinase Family

Angew Chem Int Ed Engl. 2019 Nov 25;58(48):17322-17327. doi: 10.1002/anie.201906321. Epub 2019 Oct 31.

Abstract

The CDK family plays a crucial role in the control of the cell cycle. Dysregulation and mutation of the CDKs has been implicated in cancer and the CDKs have been investigated extensively as potential therapeutic targets. Selective inhibition of specific isoforms of the CDKs is crucial to achieve therapeutic effect while minimising toxicity. We present a group of photoaffinity probes designed to bind to the family of CDKs. The site of crosslinking of the optimised probe, as well as its ability to enrich members of the CDK family from cell lysates, was investigated. In a proof of concept study, we subsequently developed a photoaffinity probe-based competition assay to profile CDK inhibitors. We anticipate that this approach will be widely applicable to the study of small molecule binding to protein families of interest.

Keywords: cancer; cyclin-dependent kinases; inhibitors; photoaffinity labeling; proteomics.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Affinity Labels / chemistry*
  • Antineoplastic Agents / chemistry*
  • Binding, Competitive
  • Cross-Linking Reagents / chemistry*
  • Cyclin-Dependent Kinases / antagonists & inhibitors*
  • Drug Screening Assays, Antitumor
  • Mass Spectrometry
  • Molecular Structure
  • Photochemical Processes
  • Protein Isoforms / chemistry*
  • Protein Kinase Inhibitors / chemistry*
  • Roscovitine
  • Structure-Activity Relationship

Substances

  • Affinity Labels
  • Antineoplastic Agents
  • Cross-Linking Reagents
  • Protein Isoforms
  • Protein Kinase Inhibitors
  • Roscovitine
  • Cyclin-Dependent Kinases