Extended Data Fig. 3: DNA engagement by S. cerevisiae CMG following translocation onto dsDNA.

a, Comparison of conformations I and II demonstrating the similarity in the overall complex architecture. For clarity, conformation I is rendered as a surface, whilst conformation II is shown as a cartoon. b, Overview of MCM C-tier domains bound to dsDNA, highlighting ATPase site occupancy in conformations I and II. c, Cryo-EM density (grey) at the MCM C-tier Mcm3-Mcm5 interface for each conformation. Mcm3, cyan; Mcm5, blue; AMP-PNP, red. d, Changes in ATPase site occupancy between conformations I and II correspond to movement of Mcm3/5/7 AAA+ domains and their DNA-binding loops (helix-2-insertion, H2I; presensor-1, PS1). Arrows indicate the relative movement of Mcm5 and Mcm3. The outward movement of Mcm3/Mcm7 in conformation II - associated with opening of the Mcm3/5 interface and loss of nucleotide at this ATPase site - leads to loss of the canonical contacts (described in panel g) formed between the Mcm3 H2I/PS1 loops and the leading-strand template DNA phosphate backbone. As such conformation II may reflect a partially disengaged state. e, Comparison of MCM:Dia2^LRR interface between conformations I and II, demonstrating lack of conformational changes in this region. f, Model of DNA in cryo-EM density (mesh) for conformation I demonstrating distortion of the B-form DNA duplex within the MCM N-tier; similar DNA density is observed within the MCM N-tier for conformation II. Approximate trajectories of DNA strands within the MCM N-tier are shown as dotted paths. g, Engagement of the leading-strand template DNA phosphate backbone by Mcm H2I/PS1 loops in complexes that have translocated onto dsDNA is comparable to that previously observed for CMG bound to ssDNA¹³. Contacts shown for the representative Mcm6 subunit (conformation I). Specific contacts with the DNA phosphate moieties indicated by dashed yellow lines.