Characterization of PARP-1 Constructs. (a) Schematic of PARP-1 indicating the domain structure and constructs used in these experiments. Domain D corresponds to the PARP1 BRCT domain. Functional domains are indicated: ZF1, first zinc finger domain; ZF2, second zinc finger domain; ZF3, third zinc finger domain; WGR, WGR domain. Domains previously demonstrated to interact with XRCC1 are identified with a red asterisk. Molecular masses of constructs, including purification tags are indicated. (b) Schematic of XRCC1 domain structure and constructs used in these experiments. Domain previously identified to interact with PARP-1 is marked with a red asterisk. Molecular masses of constructs, including purification tags are indicated. (c) Gel-filtration chromatogram of domain constructs of XRCC1 and PARP-1. Plots are colored as in (a) (green, blue, red, or light purple) and (b) (black or gray). (d) Gel-filtration chromatogram of mixtures of X1BRCTa and the various PARP-1 domain constructs from panel (a). Chromatograms of the potential complexes are colored based on the scheme of the PARP-1 constructs identified in panel (a). Note that each of the curves in this panel exhibits two maxima that approximately correspond to the maximum positions in panel c and the maximum of the X1BRCTa in panel c (black curve). Molecular mass standards used are represented as colored circles (conalbumin (76.6 kDa), pink; ovalbumin (44.3 kDa), brown; carbonic anhydrase (29.0 kDa), orange; cytochrome C (14.3 kDa), gold) in panels (c) and (d).