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Figure 6. PCNA enhances the processivity of Exo1 in resection. (A). Resection of a 3′ 32P-labeled 6 kb dsDNA fragment by recombinant, His-tagged Exo1(WT), Exo1(ΔPIP) or Exo1(D173A) in the absence or in the presence of PCNA. PCNA: 300 nM (monomer); All Exo1 proteins: 15 nM. (B). In the in vitro resection reactions containing purified Exo1 and PCNA or containing Exo1 alone, circular ssDNA derived from pBluescript SK(−), which binds and sequesters Exo1, was added at 0 or 2 min. PCNA exhibited stimulatory effects on DNA resection by Exo1(WT), but not Exo1(ΔPIP), when the sequestering ssDNA was added at 2 min, but not at 0 min, presumably because Exo1 (WT) was tethered to the DNA substrate by PCNA during the first 2 min incubation. PCNA: 300 nM (monomer); All Exo1 proteins: 15 nM; pBluescript SK(−) ssDNA: 3 nM. Image published in: Chen X et al. (2013) © The Author(s) 2013. Creative Commons Attribution license Larger Image Printer Friendly View

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