Structural Studies of a Bacterial Condensin Complex Reveal ATP-Dependent Disruption of Intersubunit Interactions

Abstract

Condensins are key mediators of chromosome condensation across organisms. Like other condensins, the bacterial MukBEF condensin complex consists of an SMC family protein dimer containing two ATPase head domains, MukB, and two interacting subunits, MukE and MukF. We report complete structural views of the intersubunit interactions of this condensin along with ensuing studies that reveal a role for the ATPase activity of MukB. MukE and MukF together form an elongated dimeric frame, and MukF's C-terminal winged-helix domains (C-WHDs) bind MukB heads to constitute closed ring-like structures. Surprisingly, one of the two bound C-WHDs is forced to detach upon ATP-mediated engagement of MukB heads. This detachment reaction depends on the linker segment preceding the C-WHD, and mutations on the linker restrict cell growth. Thus ATP-dependent transient disruption of the MukB-MukF interaction, which creates openings in condensin ring structures, is likely to be a critical feature of the functional mechanism of condensins.

MukB-mediated Catenation of DNA Is ATP and MukEF Independent

“…MukF stimulation of the MukB ATPase was maximal at the substoichiometric ratio of MukF to MukB of 1:4. This observation seems somewhat at odds with structural studies showing that the MukF dimer binds at the bottom (compared with the orientation of the coiled-coil regions) of the MukB head domain dimer and that ATP hydrolysis causes the head domain dimer to come apart, with one MukF monomer becoming largely dissociated from the complex, leaving only a flexible linker bound to one of the monomers of the MukB head domain (23).…”

“…Our observations are consistent with these previous observations; however, we observe distinct inhibition of the MukB-mediated catenation reaction by the addition of ATP, with less inhibition observed with AMP-PNP, and no inhibition observed with ADP. The binding of AMP-PNP is known to bring the head domains together (23). Thus, the observation that AMP-PNP does not inhibit the catenation reaction to the same extent as ATP suggests that it is not the closing of the MukB head domains per se that is inhibitory, but the cycling between the closed and open conformations supported by the presence of ATP.…”

“…Biochemical and structural studies have demonstrated that the binding of two ATPs is shared by the two head domains in the dimeric protein, bringing them together in a closed conformation (23). ATPase activity is stimulated by binding of the kleisin, MukF, and its associated protein, MukE (24), suggesting that the cycle of ATP binding and hydrolysis could regulate the opening and closing of the tripartite ring of the SMC complexes.…”

“…This seems quite unlikely. The presumptive DNAbinding site on the head domain dimer is only about 80 Å across at its widest point (23). Instead, we argue that MukB is playing an active role by binding two separate DNA rings and bringing them together in such a way that the helices either become intertwined or are brought close enough together that when Topo III passes strands catenation occurs (Fig.…”

Escherichia coli condensin MukB stimulates topoisomerase IV activity by a direct physical interaction

“…Similarly, the MukBEF complex remains stably associated with the E. coli chromosome following cell lysis, suggesting that this complex also serves a scaffolding role that organizes the bacterial chromosome into a higher-order structure (71). Electron microscopy studies have revealed that MukBEF complexes can aggregate to form rosettes (72), and high-resolution structural studies suggest that this process may occur through MukF/E mediated bridging of MukB dimers (73).…”