Heat-Shock Proteins Associated with Base Excision Repair Enzymes in HeLa Cells

Mendez, F., Sandigursky, M., Franklin, W.A., Kenny, M.K., Kureekattil, R. and Bases, R. Heat-Shock Proteins Associated with Base Excision Repair Enzymes in HeLa Cells. Two enzymes of base excision repair (BER), uracil DNA glycosylase (UDG) and DNA polymerase β (β pol), from HeLa cells co-eluted from Superose 12 FPLC columns. The UDG was completely displaced from 150–180-kDa fractions to 30–70-kDa fractions by brief treatment with 0.5 N NaCl, pH 3.0, as expected when protein–protein associations are disrupted, but β pol was not displaced by this treatment. UDG was not essential to the presence of β pol in the 150–180-kDa enzyme complex. β pol and UDG apparently reside in separate but co-eluting structures. Immunoaffinity chromatography showed that the association of UDG and β pol was accounted for by attachment in common to DNA and that the association was abolished by eliminating DNA. Evidence for base excision repairosomes containing UDG and β pol in protein–protein assemblies was not found. However, UDG and human AP endonuclease (HAP1) were associated with HSP70 and HSP27, which are present in 150–180-kDa and 30–70-kDa proteins of cell sonicates. The association of HSPs with BER enzymes was confirmed by hydroxyl radical protein–protein footprinting and immunoaffinity tests. The association of HSPs and BER enzymes is a novel finding. HSP binding may account for the presence of BER enzymes in the two large size class fractions and HSPs may have functional roles in BER.