CDK-containing macromolecular assemblies

The growth and division of cells is strictly controlled at the molecular level by a number of enzymes that include the cyclin dependent protein kinases (CDKs). CDKs 1, 2, 4 and 6 are switched on and off in an orderly sequence, to ensure that cell division starts and stops at the required time. Other members of the CDK family (CDKs 7, 8 and 9) are also important for the control of transcription, the process by which genes are transcribed into messenger RNA that in turn serves as the template for protein synthesis. Regulation of transcription ensures the timely expression of proteins required for cell growth and differentiation. Errors in either the control of cell cycle progression or transcription can lead to uncontrolled cell growth and proliferation. Although CDK family members are closely related in sequence, biological studies have revealed that each has unique properties. Their activities can be regulated both by association with different families of regulatory proteins and by enzymatic modification of the protein sequence by addition of phosphoryl or acetyl residues to specific amino acid side chains. Our work, primarily using the technique of X-ray crystallography, allows us to see the structures of CDKs and the complexes they form at atomic resolution and so to learn how they differ from each other. The aim of this proposal is to build upon our research into the structural and functional properties of CDK/cyclin pairs, to establish how structural and biochemical properties are altered when they are located in multi-protein complexes. To this end we propose to identify and reconstitute selected CDK-containing complexes using heterologous expression systems. We will characterise these complexes by structural, biochemical, and biophysical methods. Aberrant CDK activity has been linked to cancer, neurological diseases, and rheumatoid arthritis In recent years, understanding a particular defect that leads to disease has led to exciting new medicines directed towards a particular target (e.g. the drugs Gleevec, Iressa and Herceptin for cancer treatment). A number of CDK-selective inhibitors are in clinical trials for the treatment of cancer. These agents all act by binding to the CDK active site to block CDK activity. Compounds that block other interactions made by CDK/cyclin complexes represent an alternative target for CDK-directed therapies. The work described in this project will aid the further development of such compounds and may also reveal additional targets for inhibitor development.

Cyclin-dependent protein kinases (CDKs) play an orchestrating role in cellular processes that include transcriptional regulation and the control of cell-cycle progression. Aberrant CDK activity is implicated in diverse clinical conditions including cancer, stroke, neurodegeneration and HIV infection. Whereas CDKs isolated ex vivo are found in large, multi-protein complexes, the prevailing understanding of the CDK structure-function relationship has largely been deduced from the study of variously phosphorylated monomeric or cyclin-bound CDKs. We aim to extend available understanding of the composition, structure and functional properties of the multiprotein complexes that characterise the different CDKs. In particular we will develop a structural view of the complexes that localise and regulate CDK7 and CDK9 activity in transcriptional control, selected complexes that contain CDK4 and CDK6 and coordinate cell-cycle progression with checkpoint and apoptotic pathways, and complexes of CDKs 1 and 2 that coordinate their activity with the ubiquitin-proteolysis system.