Human DNA polymerase delta is a pentameric holoenzyme with dimeric p12 subunit: Implications in enzyme architecture and PCNA interaction

Abstract

AbstractHuman DNA polymerase delta (Polδ), a holoenzyme consisting of p125, p50, p68 and p12 subunits, plays an essential role in all the three DNA transaction processes. Herein, using multiple physicochemical and cellular approaches we found that the p12 protein forms a dimer in solution. In vitro reconstitution and pull-down of cellular Polδ by tagged p12 authenticates pentameric nature of this critical holoenzyme. Further, a consensus PIP motif at the extreme carboxyl terminal tail and a homodimerization domain at the amino-terminus of the p12 subunit were identified. Our mutational analyses of p12 subunit suggest that 3RKR5 motif is critical for dimerization that facilitates p12 binding to IDCL of PCNA via its PIP motif 98QCSLWHLY105. Additionally, we observed that oligomerization of the smallest subunit of Polδs is evolutionarily conserved as Cdm1 of S. pombe also dimerzes. Thus, we suggest that human Polδ is a pentameric complex with a dimeric p12 subunit; and discuss implications of p12 dimerization in regulating enzyme architecture and PCNA interaction during DNA replication.