Invisible Binding: DnaA, the Initiator of Chromosome Replication in Bacteria, Associates with the Escherichia Coli Inner Membrane In Vivo

DnaA initiates chromosome replication in most known bacteria and its activity is controlled to execute only once every cell division cycle. ATP in the active ATP-DnaA is hydrolyzed after initiation and ADP is replaced back to ATP on the verge of next initiation. Thus DnaA acts as a molecular switch, in which the nucleotide recycling couples key processes in the cell. Two putative recycling mechanisms presume binding of DnaA either to the membrane or to specific chromosomal sites, promoting nucleotide dissociation. While there is no doubt that DnaA interacts with artificial membranes in vitro, it is still controversial as to whether it binds the cytoplasmic membrane in vivo. We sought after DnaA-membrane interaction in E. coli cells employing fluorescent microscopy and cell fractionation with both native and fluorescent DnaA hybrids. A small (5-10%) but reliable portion of DnaA is indeed membrane associated, though invisible in fluorescent cell images.