Purification, cDNA cloning, and gene mapping of the small subunit of human DNA polymerase ε

Abstract

HeLa DNA polymerase s (pol ε), possibly involved in both DNA replication and DNA repair, consists of a catalytic subunit of 261 kDa and a tightly bound peptide with a relative molecular mass of 55 kDa. The cDNA of the 261-kDa polypeptide has been independently cloned, sequenced, and then overexpressed in insect cells to give a soluble, but catalytically unstable protein, suggesting that the small subunit of HeLa pol ε might be important for stability. HeLa pal ε has been isolated by immunoaffinity purification to obtain sequence information which enabled the cloning of a full-length human cDNA encoding the small subunit. The clone encoded nine proteolytic peptides obtained from the subunit. The 59,434-Da predicated polypeptide has 26% identity and 44% homology to the yeast pal ε 80-kDa subunit, DPB2. Using fluorescence in situ hybridization, the human poI ε p59 locus (DPE2) was assigned to chromosome 14q13-q21.