INTERACTION STUDIES BETWEEN THE P21CIP1 WAF1 CYCLIN-DEPENDENT KINASE INHIBITOR AND PROLIFERATING CELL NUCLEAR ANTIGEN (PCNA) BY SURFACE-PLASMON RESONANCE/

The cyclin-dependent kinase (CDK) inhibitor p21Cip1 consists of two do mains that interact with CDKs and proliferating cell nuclear antigen ( PCNA), respectively, We have investigated the interaction between p21C ip1 and PCNA using surface plasmon resonance (SPR) technology and comp ared the results with those obtained from other sources such as the ye ast two-hybrid system, Whilst other methods are only semi-quantitative , the SPR technique allowed us to determine the kinetic parameters of the interaction, The apparent equilibrium constant K-D calculated for these kinetic parameters was 3.2 x 10(-7) M, We further demonstrate th e use of SPR to study the interaction between mutant proteins and to d etermine their actual K-D, The interaction between p21Cip1/PCNA is sho wn to be dependent upon the trimeric conformation of PCNA since a poin t mutant that abolishes PCNA-PCNA interaction also abolishes PCNA's in teraction with p21Cip1, Finally, we demonstrate that SPR can be used t o characterise the interaction of p21Cip1 and PCNA in the presence of short competitive peptides.