Human and Pyrococcus furiosus FEN-1 (five prime exonuclease-1 or flap endonuclease-1) activities as a function of temperature. (A) The 5' branched DNA flap substrate used in the FEN-1 endonuclease assays. FEN-1 loads onto the free 5' end of the substrate strand, slides to the junction of ssDNA and dsDNA formed at the intersection of the three DNA strands, and cleaves at the junction as indicated by the arrows. The asterisk at the 5' end of the substrate strand indicates 32P radiolabel. (B) hFEN-1 and pFEN-1 were incubated with this flap substrate at either 37 degrees C or 55 degrees C as indicated. The 19-nucleotide and 21-nucleotide reaction products result from FEN-1 catalyzed cleavage one nucleotide 3' and one nucleotide 5' of the junction point. While the human enzyme was active only at the lower temperature (lane 1), the pyrococcal enzyme showed substantially higher activity at 55 degrees C (lane 4). Lane C is the control (no enzyme).
From: Hosfield et al. (1998) Structure of the DNA repair and replication endonuclease and exonuclease FEN-1: Coupling DNA and PCNA binding to FEN-1 activity. Cell 95: 135-146.