Bacterial actin: Architecture of the ParMRC plasmid DNA partitioning complex

During eukaryotic cell division, the DNA molecules are partitioned to opposite poles of the cell, ensuring accurate distribution of chromosomes between the dividing cells. In prokaryotes, the mechanism for accurate segregation is unknown. Previous studies have focused on partitioning of a plasmid, Rl, which encodes a stability operon, par, that is required for proper segregation of this low-copy-number plasmid. The par operon of Rl encodes three components: ParM, an actin-like ATPase, ParR, a DNA-bind- ing protein, and parC, a centromere-like DNA sequence element to which ParR binds. In the presence of ATP, ParM polymerizes and then disassembles. Disassembly is prevented, however, by the binding of ParR to parC, which stabilizes the ParM filaments. In the current model for DNA segregation, the ATP-dependent polymerization of ParM is believed to move newly replicated plasmids to the opposite poles. This study therefore determined how the ParM filaments are formed and stabilized and ascertained the sites involved to effect both mechanisms. Mutational studies, pull-down and gel- shift assays for point and deletion mutants of Rl ParR, and electron microscopy at varying concentrations of labeled parC established that the ParR- parC complex forms the clamp that binds at both the C-terminal ends of ParM, forming ring structures that bind ATP and stabilize the ParM filaments. ParR binds through its N-terminus to parC, forming a rigid scaffold of protein (ParR) wrapped by a DNA helix (parC). This binding does not include the Rl promoter region within the parC domain, because the promoter region extends out as a loop from the ParR binding region. Additionally, studies with mutants of ParM also identified the binding sites of the ParR-parC complex on ParM. A model for ParM polymerization consistent with electron micrograph images was thus proposed whereby ATP binding polymerizes ParM. Binding of the ParR-parC complex clamps the C-terminal ParM-ATP complex stabilizing filament formation. Hydrolysis of ATP to ADP dislocates ParR-parC and translocates ParR to one side to allow binding of a new ParM-ATP monomer. ParR translocates back, rocking the clamp, and the cycle of hydrolysis, translocation, and monomer addition is repeated until elongation is completed (Fig. 1). © 2008 Data Trace Publishing Company.