Kinetics of dimerization and interactions of p13suc1 with cyclin-dependent kinases.
The impact of p13suc1 on the conformation and regulation of cyclin-dependent kinases (cdks) and cyclins was investigated by spectroscopic and rapid kinetic approaches. In the absence of phosphorylation on cdks, p13suc1 formed stable complexes, mainly stabilized by hydrophobic interactions, specifically with cdk2 and cdc2. The presence of cyclin A, associated with cdk2 or cdc2, increased the stability of the interaction between cdk2 and p13suc1 by a factor of 2. However, cyclin A did not modify the association rate of p13suc1 to cdk2, but the dissociation rate, which was decreased 3-fold. Moreover, binding of p13suc1 to cdk2 resulted in a 2-fold decrease in the release of nucleotide from cdk2, indicating that p13suc1 induces a marked change in the structure of the nucleotide binding site of cdks. On the basis of the structure of cdk2/CksHs1 complex and on our kinetic results, we propose that the binding of Cks proteins to C-lobe of cdk2 is stabilized by the presence of cyclin A and that it may modify the orientation of the loop carrying residues 14 and 15 and their consequent access for dephosphorylation by cdc25 phosphatases. Finally, we have shown that dimerization of p13suc1 in the presence of zinc abolishes its interaction with cdks, which suggests that the binding of p13suc1 to cdk2 or cdk2/cyclin A may be regulated by dimerization of p13suc1 in vivo.
Morris
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MC
Centre de Recherches de Biochimie Macromoleculaire, CNRS, Montpellier, France.
Heitz
F
F
Divita
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G
D000595:Amino Acid Sequence; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism; D002942:Circular Dichroism; D018844:Cyclin-Dependent Kinases / Q000737:chemistry* / Q000378:metabolism*; D016213:Cyclins / Q000378:metabolism; D019281:Dimerization; D005454:Fluorescence Polarization; D005656:Fungal Proteins / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae; D029702:Schizosaccharomyces pombe Proteins; D013050:Spectrometry, Fluorescence; D015032:Zinc / Q000494:pharmacology
D018797:Cell Cycle Proteins; D016213:Cyclins; D005656:Fungal Proteins; D046911:Macromolecular Substances; D029702:Schizosaccharomyces pombe Proteins; C051655:Suc1 protein, S pombe; D018844:Cyclin-Dependent Kinases; D015032:Zinc