Synthetic peptides to selected sequences in human DNA polymerase delta (pol delta) were used to identify the region involved in the interaction of pol delta to proliferating cell nuclear antigen. Peptides corresponding to sequences in five regions in the amino terminus of human pol delta and three in the carboxyl terminus, which are conserved with the yeast homologs of pol delta, were tested. These studies showed that the peptide corresponding to the N2 region (residues 129-149) selectively and specifically inhibited the PCNA stimulation of pol delta. This inhibition was relieved by titration with excess PCNA. The identification of the N-2 region as being involved in PCNA binding was supported by studies that demonstrated that the N2 peptide could bind PCNA. Deletion mutants of pol delta expressed in Sf9 cells provided evidence that the binding region for PCNA was located in the first 182 residues of the amino terminus. These studies provide reasonable evidence that residues within the region 129-149 of pol delta are involved in the binding site for PCNA.