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Abstract

Division of one mother cell into two daughter cells resides at the very core of living organisms. To ensure that the cell’s genetic material is equally segregated into both daughter cells, cells undergo a sophisticated succession of highly controlled events. One of these events is the packaging of chromatin fibers into mitotic chromosomes that can be transported by spindle microtubules. The key factor for this chromosome condensation process is the five-subunit condensin complex. Condensin is thought to shape chromosomes by actively extruding large chromatin loops, yet how condensin can create such loops has remained largely unknown. To shed light onto the mechanism of condensin-mediated chromatin condensation, insights into the structure of the complex will be indispensable. While X-ray crystallography proved efficient in providing said information for individual parts of the complex, the architecture of the entire complex remained unknown. The aim of the work described in this PhD thesis was to close this crucial gap in knowledge by elucidating the condensin holo complex structure. I employed cryogenic electron microscopy to first solve the structure of the Saccharomyces cerevisiae condensin holo complex in its apo and nucleotide-bound states. In the absence of nucleotide, condensin adopts a rod-like conformation. The HEAT-repeat subunit Ycs4 stably interacts with the ATPase head domains of closely aligned Smc2 and Smc4 subunits, while the Ycg1 HEAT-repeat subunit is flexibly tethered to the rest of the complex through the Brn1 subunit. Instead of forming a fully stretched rod, the Smc2–Smc4 coiled-coil arm segment contains a kink that results in the hinge folding back onto the coiled coils. In a second apo state, the Smc2 and Smc4 heads are bridged by Ycs4, which splits apart the Smc2–Smc4 coiled coils from the head to the joint regions. Addition of ATP induces a drastic structural rearrangement. The ATPase heads engage, which results in an increase in flexibility and opening of the coiled coils. Furthermore, Ycg1 and Ycs4 swap positions as ATP releases Ycs4 from the Smc4 ATPase head, which in turn provides access for Ycg1 to directly bind the Smc2 ATPase head domain. These data provide a structural framework for the condensin ATPase cycle and suggest that an ATP-driven exchange of the Ycs4 and Ycg1 subunits interconverts DNA binding sites that might form the core of the condensin DNA loop-extruding activity.