Scanning force microscopy of the complexes of p53 core domain with supercoiled DNA

We used scanning force microscopy to analyse the interaction of the core do main of the tumor suppressor protein p53 (p53CD, amino acid residues 94 to 312), with supercoiled DNA (scDNA) molecules. The com Flexes were attached to a mica substrate by the divalent cation spreading technique. p53CD bound to supercoiled plasmid pPGM1 bearing the consensus sequence 5'-AGACATGCCTA GACTGCCT-3' (p53CON) was imaged as a globular complex. Only one such comple x was observed with each scDNA molecule. In contrast, binding to supercoile d pBlue-script II SK- DNA (lacking the consensus sequence) resulted in the appearance of multiple, variable size complexes of various sizes on single DNA molecules. Addition of p53CD to scDNA containing a cruciform-forming (A T),, insert resulted in the binding of the protein exclusively at the cruci form. The data presented here suggest that p53CD can form stable specific a nd non-specific complexes with supercoiled DNA molecules, albeit of variabl e multimeric organization. (C) 2000 Academic Press.