Differences in Unfolding Energetics of CcdB Toxins From V. fischeri and E. coli

Abstract

Ccd system is a toxin-antitoxin module (operon) located on plasmids and chromosomes of bacteria. CcdB(F) encoded by ccd operon located on Escherichia coli plasmid F and CcdB(Vfi) encoded by ccd operon located on Vibrio fischeri chromosome are members of the CcdB family of toxins. Native CcdBs are dimers that bind to gyrase-DNA complexes and inhibit DNA transcription and replication. While thermodynamic stability and unfolding characteristics of the plasmidic CcdB(F) in denaturant solutions are reported in detail, the corresponding information on the chromosomal CcdB(Vfi) is rather scarce. Therefore, we studied urea-induced unfolding of CcdB(Vfi) at various temperatures and protein concentrations by circular dichroism spectroscopy. Global model analysis of spectroscopic data suggests that CcdB(Vfi) dimer unfolds to the corresponding monomeric components in a reversible two-state manner. Results reveal that at physiological temperatures CcdB(Vfi) exhibits lower thermodynamic stability compared to CcdB(F). At high urea concentrations CcdB(Vfi), similarly to CcdB(F), retains a significant amount of secondary structure. Differences in thermodynamic parameters of CcdB(Vfi) and CcdB(F) unfolding can reasonably be explained by the differences in their structural features.