Extended Data Fig. 5: Interaction of HPF1 with the nucleosome stabilizes the PARP2–HPF1–nucleosome complex.

a, Native gel showing the PARP2–HPF1–nucleosome complex assembly with equimolar amounts of free DNA and nucleosomes. Nucleosomal and free DNA are labelled with Alexa 488. PARP2–HPF1 binds nucleosomes with higher affinity than free DNA. b, EMSA analysis of the assembly of PARP2–nucleosome and PARP2–HPF1–nucleosome complexes. HPF1 contributes to stability of the complex. Native gel is stained with SYBR Gold. c, SDS–PAGE showing quality of wild-type and mutant HPF1 proteins. d, EMSA analysis of PARP2–HPF1–nucleosome complex assembly with wild-type and mutant HPF1. Mutations in loops that interact with nucleosomal DNA destabilize the complex. Native gel is stained with SYBR Gold. e, One PARP2–HPF1 in the map with two PARP2–HPF1 complexes shows flexibility in the N-terminal region of HPF1. Note an additional density spanning from HPF1 to the double-strand DNA break site. This density could be generated by missing HPF1 helices, HPF1 and the PARP2 N-terminal tail or the H3 tail. f, Superposition of the PARP2 catalytic domains from the PARP2–HPF1 crystal structure (grey; PDB 6TX3) and the PARP2–HPF1–nucleosome cryo-EM model (violet and magenta). HPF1 is slightly rearranged in the cryo-EM structure as compared to the X-ray structure. g, Superposition of the PARP2 catalytic domains from the PARP2–HPF1 crystal structure (grey; PDB 6TX3) and the PARP2–HPF1–nucleosome cryo-EM model. The model is coloured by r.m.s.d. One representative experiment of at least 3 independent experiments is shown for all biochemical data. For gel source data, see Supplementary Fig. 1.