Figure 7.
WRN helicase activity and FEN-1 nuclease activity are responsible for HJ cleavage. (A) WRN ATPase/helicase but not exonuclease activity is required for stimulation of FEN-1 HJ resolution. Reactions containing 2.5 fmol of HJ(X12-1), wild-type or mutant WRN (12 nM) and FEN-1 (29 nM) were conducted under standard conditions. Phosphorimage of a typical native gel is shown. WT, wild-type WRN; K, WRN-K577M; X, WRN-E84A. (B) FEN-1 nuclease activity is responsible for HJ resolution. Reactions were conducted under standard conditions using either wild-type FEN-1N-HIS, wild-type FEN-1C-HIS, or nuclease defective FEN-1C-HIS-D181A proteins as indicated. Phosphorimage of a typical native gel is shown. (▴), heat-denatured substrate control.