Fig. 1

Identification of APEX2 and P0 homologues in C. intestinalis and purification results. a and b The amino acid sequences of C. intestinalis APEX2 and P0 are aligned with H. sapiens and S. cerevisiae homologues for APEX2 and H. sapiens and D. melanogaster homologues for P0. Amino acid residues are highlighted in black (identical) or grey (similar). a Amino acid sequence alignment among APEX2 homologues. Conserved endonuclease/exonuclease/phosphatase domain are enclosed by a black box. b Amino acid sequence alignment among P0 homologues. c-e Purification of GST-CiAPEX2 (c), GST-CiP0 (d), tag-free CiP0 (d) and His-CiAPEX2 (e). Each protein was purified from bacteria lysate using column chromatography. Purified proteins were electrophoresed on a 10% SDS-PAGE gel, and stained with Coomassie brilliant blue. The arrows indicate each of the purified proteins. Especially in (d), upper arrow indicates GST-CiP0 and lower arrow indicates tag-free CiP0. c and (d GST-CiAPEX2 (c), GST-CiP0 (d) and tag-free CiP0 (d) purification. The fractions are as follows: SUP, supernatant; GST/FT, flow-through fraction from Glutathione-Sepharose 4B column; GST/Wash, washed fraction by buffer A; GST/Elu, fraction eluted by 40 mM glutathione; tag-free CiP0, the product of GST-CiP0 thrombin cleavage; Cleaved GST, the byproduct of GST-CiP0 thrombin cleavage. e His-CiAPEX2 purification. The fractions are as follows: SUP, supernatant; His/FT, flow-through fraction from Chelating Sepharose Fast Flow; His/ Wash, washed fraction by buffer D containing 10 mM imidazole; His/Elu, fraction eluted by 0.5 M imidazole; Q/ FT, flow-through fraction from HiTrap-Q; Q/0.235-0.32 M, fractions collected in 0.235-0.32 M NaCl