Interplay Between CedA, rpoB and Double Stranded DNA: A Step Towards Understanding CedA Mediated Cell Division in E. coli

Sharma P, Tomar AK, and Kundu B, 107(Pt B):2026-2033, Int J Biol Macromol., 2018

In DnaAcos mutant E. coli cells, cell division is inhibited due to excessive initiation of chromosomal replication. Previous studies suggest, Cell division activator (CedA), a double stranded-DNA binding protein, acts as a regulatory protein in DnaAcos mutants to begin cell division and regular colony formation. CedA is known to interact with various subunits of RNA polymerase complex, including rpoB. Reported herein are the mechanistic aspects of interactions between CedA, rpoB, and double stranded DNA (ds-DNA) by using a combination of biophysical and insilico experiments. The kinetics of the binding interaction of ds-DNA with CedA, rpoB, and CedA-rpoB complex was studied using Bio-Layer Interferometry (BLI). A Pall ForteBio Octet RED96 system equipped with Streptavidin (SA) Biosensor probes was used to perform all BLI assays. Biotinylated-DNA was immobilized on SA sensor tips. Association and dissociation kinetics of CedA, rpoB, and CedA-rpoB complex were studied. The binding data were globally fitted using 2:1 binding model. Kinetic parameters (kon and koff) and the KD values were determined for all binding interactions. Overall results of this study suggest that chromosome over-replication in E. coli signals CedA to recruit rpoB to specific DNA site(s), thereby initiating cell division.

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