The Characterization of Human cdc2 Kinase and CDK2

Abstract

p34^cdc2 kinase plays a key role in the initiation of mitosis. The activity of this kinase requires the binding of a protein, named cyclin, to it. The kinase forms a heterodimer with cyclin. Cyclin A or cyclin B is the counterpart of this complex. The differences in the activity between cyclin A/cdc2 kinase and cyclin B/cdc2 kinase have not been cleared. In recent years, the other cdc2-like kinases were identified. One of them was CDK2 (cyclin dependent kinase 2). CDK2 could rescue the defect of the budding yeast CDC28 mutation, which arrested the cells at a point named START, in G₁ phase. Then, CDK2 was thought to be worked at G₁ through S phase in a cell cycle, but the details on the role of this kinase has not been cleared so far. In this study, we separated the human cyclin A/cdc2 kinase, cyclin B/cdc2 kinase and CDK2, each other by use of column chromatography, and characterized the each kinase. These kinases had the same substrate specificities when the synthesized peptides were tested. They phosphorylated the threonine residue in the sequence -Thr-Pro-Lys-Lys-Ala- but hardly phosphorylated threonine residue the sequence -Thr-Pro-Lys-Ala-Lys-. They had some differences in the substrate-proference when the native proteins were tested. In a cell cycle of human cells, the activity of cdc2 kinase increased at G₂/M phase and the activity of CDK2 was high from S through M phase. These data suggested that cdc2 kinase works at the transition from G₂ to M phase and that CDK2 works from G₁ through G₂/M phase. They could phosphorylate different protein-substrates having the common phosphorylated sequence -Thr-Pro-X-Lys-.