The HAP1 protein stimulates the turnover of human mismatch-specific thymine-DNA-glycosylase to process 3,N(4)-ethenocytosine residues.

When present in DNA, 3,N(4)-ethenocytosine (epsilon C) residues are potentially mutagenic and carcinogenic in vivo. The enzymatic activity responsible for the repair of the epsilon C residues in human cells is the hTDG protein, the human thymine-DNA-glycosylase that removes thymine in a T/G base pair [Proc. Natl. Acad. Sci., U.S.A., 95 (1998) 8508]. One of the distinctive properties of the hTDG protein is that it remains tightly bound to the AP-site resulting from its glycosylase activity. In this paper we report that the human AP endonuclease, the HAP1 (Ape1, APEX Ref-1) protein, stimulates the processing of epsilon C residues by the hTDG protein in vitro, in a dose-dependent manner. This property of HAP1 protein is specific since E.coli Fpg, Nfo and Nth proteins, all endowed with an AP nicking activity, do not show similar features. The results suggest that the HAP1 protein displaces the hTDG protein bound to the AP-site and therefore increases the turnover of the hTDG protein. However, using a variety of techniques including gel retardation assay, surface plasmon resonance and two-hybrid system, it was not possible to detect evidence for a complex including the substrate, the hTDG and HAP1 proteins.