Sirt6 stimulates Myh and APE1 activities. a mSirt6 enhances mMyh glycosylase activity. DNA substrates used are shown with arrows indicating the cleavage sites. Lane 1 5′-FAM-labeled A/Go-containing DNA. Lane 2 5 nM A/Go-DNA was incubated with 0.5 nM mMyh. Lanes 3–8 are similar to lane 2 but with added 1, 2, 4, 8, 16, and 32 nM mSirt6, respectively. Lane 9 A/Go-DNA was incubated with 32 nM mSirt6 without mMyh. Arrows mark the intact DNA substrate (S) and the cleavage product (P). Percentage (%) of product generated is shown below each lane. c mSirt6 enhances the AP endonuclease activity of hAPE1. Lane 1 3′-FAM-labeled tetrahydrofuran (THF)/G-DNA. Lane 2 20 nM THF/G-DNA was incubated with 0.002 nM hAPE1. Lanes 3–6 are similar to lane 2 but with added 1, 2, 4, and 8 nM mSirt6, respectively. Lane 7 THF/G-DNA was incubated with 8 nM mSirt6 without APE1. e mSirt6 enhances the phosphodiesterase activity of hAPE1 that cleaves the 3′FAM from a U/G-containing DNA. Lane 1 3′-FAM-labeled U/G-DNA. Lane 2 20 nM U/G-DNA was incubated with 0.01 nM hAPE1. Lanes 3–7 are similar to lane 2 but with added 1, 2, 4, 8, and 16 nM mSirt6, respectively. Lane 8 U/G-DNA was incubated with 16 nM mSirt6 without APE1. The product is a free FAM. b, d, and f, Quantitative analyses of the fold of stimulation from results as in a, c, and e, respectively. Error bars indicate SD; n = 3.