Abstract | BACKGROUND: Chemical inhibitors of cyclin-dependent kinases (CDKs) have great therapeutic potential against various proliferative and neurodegenerative disorders. Olomoucine, a 2,6,9-trisubstituted purine, has been optimized for activity against CDK1/ cyclin B by combinatorial and medicinal chemistry efforts to yield the purvalanol inhibitors. Although many studies support the action of purvalanols against CDKs, the actual intracellular targets of 2,6, 9-trisubstituted purines remain unverified. RESULTS: To address this issue, purvalanol B (95. ) and an N6-methylated, CDK-inactive derivative (95M. ) were immobilized on an agarose matrix. Extracts from a diverse collection of cell types and organisms were screened for proteins binding purvalanol B. In addition to validating CDKs as intracellular targets, a variety of unexpected protein kinases were recovered from the 95. matrix. Casein kinase 1 (CK1) was identified as a principal 95. matrix binding protein in Plasmodium falciparum, Leishmania mexicana, Toxoplasma gondii and Trypanosoma cruzi. Purvalanol compounds also inhibit the proliferation of these parasites, suggesting that CK1 is a valuable target for further screening with 2,6,9-trisubstituted purine libraries. CONCLUSIONS: That a simple batchwise affinity chromatography approach using two purine derivatives facilitated isolation of a small set of highly purified kinases suggests that this could be a general method for identifying intracellular targets relevant to a particular class of ligands. This method allows a close correlation to be established between the pattern of proteins bound to a small family of related compounds and the pattern of cellular responses to these compounds.
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Authors | M Knockaert, N Gray, E Damiens, Y T Chang, P Grellier, K Grant, D Fergusson, J Mottram, M Soete, J F Dubremetz, K Le Roch, C Doerig, P Schultz, L Meijer |
Journal | Chemistry & biology
(Chem Biol)
Vol. 7
Issue 6
Pg. 411-22
(Jun 2000)
ISSN: 1074-5521 [Print] United States |
PMID | 10873834
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
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Chemical References |
- Enzyme Inhibitors
- Cyclin-Dependent Kinases
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Topics |
- Amino Acid Sequence
- Animals
- Chromatography, Affinity
(methods)
- Cyclin-Dependent Kinases
(antagonists & inhibitors)
- Enzyme Inhibitors
(pharmacology)
- Eukaryota
(enzymology)
- Molecular Sequence Data
- Oocytes
(drug effects, enzymology)
- Rats
- Starfish
(cytology)
- Substrate Specificity
- Swine
- Xenopus laevis
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