Intracellular targets of cyclin-dependent kinase inhibitors: identification by affinity chromatography using immobilised inhibitors.
Chem Biol
; 7(6): 411-22, 2000 Jun.
Article
em En
| MEDLINE
| ID: mdl-10873834
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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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Cromatografia de Afinidade
/
Quinases Ciclina-Dependentes
/
Inibidores Enzimáticos
Tipo de estudo:
Diagnostic_studies
Limite:
Animals
Idioma:
En
Ano de publicação:
2000
Tipo de documento:
Article