FAAP20 is a novel component of the FA nuclear core complex. (A) Silver-stained gel showing polypeptide bands isolated by 2-step purification, first purification using anti-FLAG (α-FLAG) and second purification using talon beads (Co⁺⁺), from nuclear extracts of HeLaS3:vector or HeLaS3:_His6FANCA/_FLAG-FAAP100 cells. Polypeptides identified by MS analysis are indicated, including the novel 20 kDa polypeptide, FAAP20 (asterisk). (B) Silver-stained gel showing the polypeptide bands purified from nuclear extracts of HeLaS3:vector or HeLaS3:_His6-FLAGFAAP20 using 2-step purification as described in panel A. Polypeptides identified by MS analysis are indicated. (C) Immunoblot of immunoprecipitated HeLa cell extract showing core-complex proteins present in IPs with anti-FAAP20 (lane 3), but absent in IPs with pre-immune serum (lane 4). FT is the flow-thru fraction. (D) Immunoblot showing FAAP20 coprecipitated in FLAG-IPs from HeLaS3:_His6-FLAGFANCG, HeLaS3: _His6-FLAGFANCL, and HeLaS3:_His6-FLAGFAAP100 and not in HeLaS3:vector. (E) Immunoblot showing identical gel filtration profiles on cofractionation of FANCA, FANCG, and FAAP20 using a superose 6 gel filtration column.