INTERACTIONS OF USF AND KU ANTIGEN WITH A HUMAN DNA REGION CONTAININGA REPLICATION ORIGIN

By means of a combination of ion-exchange and sequence-specific affini ty chromatography techniques, we have purified to homogeneity two prot ein complexes binding in a human DNA region (B48) previously recognize d to contain a DNA replication origin. The DNA sequence used for the p rotein purification (B48 binding site) contains a binding site for bas ic-helix-loop-helix DNA binding proteins. The first complex is compose d of two polypeptides of 42- and 44-kDa; its size, heat stability, and target DNA sequence suggest that it corresponds to transcription fact or USF; furthermore, the 42-kDa polypeptide is recognized by antibodie s raised against 43-kDa-USF. The second complex is represented by equi molar amounts of two proteins of 72 and 87 kDa; microsequencing of the two species indicated that they correspond to the human Ku antigen. I n analogy with Ku, they produce a regular pattern of footprints withou t an apparent sequence-specificity, and their binding can be competed by unspecific DNA provided that it contains free ends. The potential r ole of B48 binding site and of these cognate proteins in origin activa tion is discussed.