CTP and parS control ParB partition complex dynamics and ParA-ATPase activation for ParABS-mediated DNA partitioning

ParABS partition systems, comprising the centromere-like DNA sequence parS, the parS-binding ParB-CTPase and the nucleoid-binding ParA-ATPase, ensure faithful segregation of bacterial chromosomes and low-copy-number plasmids. F-plasmid partition complexes containing ParBF and parSF move by generating and following a local concentration gradient of nucleoid-bound ParAF. However, the process of ParBF activation of ParAF-ATPase has not been defined. We studied CTP- and parSF-stimulated ParAF--ParBF complex assembly leading to ParAF-ATPase activation. Activation of DNA-bound ParAF-ATP dimers for ATP hydrolysis requires binding of two ParBF N-terminal domains. CTP or parSF enhances the ATPase rate without increasing ParAF--ParBF assembly kinetics. Together, parSF and CTP accelerate ParAF--ParBF assembly without further increasing the ATPase rate. Magnetic-tweezers experiments showed that CTP promotes multiple ParBF loading onto parSF-containing DNA, generating condensed partition complex-like assemblies. We propose that ParBF in the partition complex adopts a conformation that enhances ParBF--ParBF and ParAF--ParBF interactions promoting efficient partitioning.