Identification of binding domains on the p21Cip1 cyclin-dependent kinase inhibitor.
Members of the recently discovered family of cyclin-dependent kinases inhibitors (CKIs) appear to play an essential regulatory role in the control of cell proliferation. To investigate the molecular basis of the interaction between these proteins and the cyclin-dependent kinases (CDKs), we performed a systematic mutagenesis of the CKI family member p21Cip1 using the alanine-scanning strategy. We have examined the interaction between in vitro translated human cdk2, cyclins A and D1, purified proliferating cell nuclear antigen (PCNA) and a set of human p21Cip1 mutants fused to glutathione S-transferase. Independent domains that are required for the interaction with cdk2 and with PCNA have been identified. The cdk2 binding domain is located in the N-terminal part of the protein, between residues 45 and 60, a region that is fully conserved in the p27Kip1 inhibitor. A PCNA binding region was localised to the C-terminus of the protein, between residues 142 and 163. These findings define protein motifs that are highly conserved between members of the CKI family and that are likely to play an essential function in the regulation of the G1/S transition.
Goubin
F
F
Laboratoire de Pharmacologie et de Toxicologie Fondamentales, CNRS, Université Paul Sabatier, Toulouse III, France.
Ducommun
B
B
D000595:Amino Acid Sequence; D001665:Binding Sites; D042846:CDC2-CDC28 Kinases; D018797:Cell Cycle Proteins; D051357:Cyclin-Dependent Kinase 2; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050760:Cyclin-Dependent Kinase Inhibitor p27; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D016213:Cyclins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017354:Point Mutation; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D025521:Tumor Suppressor Proteins
C493956:CDKN1A protein, human; D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D008869:Microtubule-Associated Proteins; D018809:Proliferating Cell Nuclear Antigen; D025521:Tumor Suppressor Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases