(A) Representative reference-free 2D class averages of DnaB (RF) compared to forward reprojections of the 3D structure obtained after multi-reference refinement (FRP).
(B) The 3D reconstruction of DnaB shows a closed ring with a broad inner channel.
(C) Representative reference-free 2D class averages of the DnaBC complex (RF) compared to forward reprojections of the 3D structure obtained after multi-reference refinement (FRP).
(D) The 3D reconstruction for DnaBC shows a 3-tiered, cracked-ring particle.
See .

(A) Closed-ring B. stearothermophilus DnaB crystal structure (PDB ID 2R6A, ()) fitted in the E.coli DnaB EM reconstruction.
(B) Closed-ring B. stearothermophilus DnaB crystal structure (PDB ID 2R6A) filtered at 25 Å.
(C) DnaBC forms a three-tiered, right-handed, cracked-ring structure. The upper tier is colored in blue (1), middle in purple (2) and lower in orange (3).
(D) DnaBC view showing the discontinuity in the cracked ring. The orientation is orthogonal to that of panel C.
(E) The DnaBC lower tier exhibits 6₁ symmetry.
(F) The DnaBC upper tier exhibits 3₁ symmetry.
(G) Longitudinal DnaBC section, showing the central channel that runs along the three tiers. The orientation is equivalent to that of panel D
(H) DnaBC displayed at lower threshold (5.7 σ contour) to highlight the discontinuity of the ring. This view is equivalent to that of panel D.

(A) Primary structure of E.coli DnaB and E.coli DnaC.
(B) Structure of a typical DnaB N-terminal domain homodimer (PDB ID 2R6A, ()).
(C) Docking of DnaB N-terminal domain homodimers into tier 1 of the DnaBC model. Subunits are alternatingly colored as per panel B.
(D) Docking of DnaB RecA folds into tier 2 of the DnaBC model. Subunits are alternatingly colored purple and pink. The density corresponding to the third tier has been removed for clarity.
(E) Side view of both domains of DnaB fitted into the DnaBC complex. The density corresponding to the third tier is not shown.

(A) Detail view of DnaC_AAA+ crystal structure (PDB ID 3ECC ()). The DnaC ATPase domains appear to productively associate with nucleotide binding in this complex.
(B) DnaC_AAA+ hexamer (reconstituted by imposing the crystallographically-observed 61 packing arrangement on PDB ID 3ECC), fitted into the lower tier of the DnaBC complex.
(C) Composite model for DnaBC. The six columns of unaccounted density connecting the DnaC_AAA+ and DnaB RecA domains likely correspond to the DnaC N-terminal regions, which bind DnaB. See .
(D) SAXS analysis of DnaBC. Experimental scattering data (1 mM AMPPNP, pink) fits well to a theoretical curve calculated from the DnaBC EM reconstruction (blue), but not to that of a closed-ring DnaB hexamer (purple). Improved fitting is obtained from a multiple ensemble search containing a mixture of DnaBC model and free DnaB (green). See .
(E) Clamp-loader AAA+ domains oligomerize into a right-handed spiral to open a processivity clamp (PDB ID 3U5Z ()).

(A) The N-terminal homodimers of DnaB in the absence of nucleotide form a wide, closed triangular collar (PDB ID 2R6A, ()). DnaB RecA domains are displayed as surfaces, whereas the N-terminal domains and linker helices are shown as light blue/orange cylinders.
(B) The N-terminal domains of DnaB within the DnaBC complex undergo a marked positional shift from the closed ring state, forming new packing arrangements between dimers. The DnaB RecA domains form a cracked spiral (boxed inset); DnaC is omitted for clarity. A schematic of the arrangement for the N-terminal domain dimers is shown in the upper left corner of each panel. Arrows in panel A indicate the movement of the DnaB NTDs to the state shown in panel B.
See and and .

(A) Removal of the N-terminal domain of the DnaB-family G40P protein allows its associated RecA ATPase to assemble into a helical filament (PDB ID 3BH0, ()).
(B) The fitted DnaB RecA domains within DnaBC structure adopt a spiral pitch similar to that seen for N-terminally truncated G40P.
(C) The DnaC NTD can promote the DnaB-dependent unwinding of a topologically-closed DNA substrate that requires helicase ring opening and loading. (Top) Gel showing unwinding of a 3’ tailed oligonucleotide annealed to circular M13mp18 ssDNA in the presence of DnaB with or without DnaC or the DnaC NTD. Lanes N and D indicate the native and boiled substrate, respectively. (Bottom) Quantification of product seen by gel. Concentrations of DnaB hexamers in the reactions are shown; DnaC, when present, was included at a 2-fold molar excess of DnaB monomers. Columns and error bars represent the average and standard deviations, respectively, of at least five measurements.
(D) DnaC and the DnaC NTD can promote the DnaB-dependent unwinding of a topologically-accessible forked DNA substrate. Plot shows unwinding of a fluorophore/quench-labeled DNA by DnaB in the presence or absence of DnaC or the DnaC NTD. Data points and error bars represent the average and standard deviation, respectively, from at least six measurements. See .

(A) DnaC opens and remodels DnaB to facilitate DNA loading and unwinding. Closedring DnaB cannot engage a topologically-closed DNA substrate. DnaC associates with the helicase, remodeling the N-terminal collar and triggering helicase opening. In the presence of ATP, DnaC AAA+ domains further assemble into a helical conformation that stabilizes the open-ring complex and assists with DNA binding. ATP hydrolysis by DnaC and/or DnaG help disengage the loader (; ) leaving an active helicase encircled around DNA. See also .
(B) Hypothetical model showing how DnaC is free to associate with a DnaA filament even when bound to DnaB (as proposed in ()). The model was generated by aligning a DnaA filament bound to single-stranded DNA (PDB ID 3R8F ()) – which bears an exposed arginine-finger at the 5’ end of the complex – with the solvent accessible nucleotide-binding face of the terminal DnaC protomer in DnaBC in a manner consistent with typical AAA+/AAA+ interactions. The superposition co-aligns the pores of the all three proteins, positioning DNA bound by the central channel of DnaA to enter into the helicase/loader complex.