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
f lexibility 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 6T X3) 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 6T X3) 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.