Hydrodynamic characterization of the FtsZ protein from Escherichia coli demonstrates the presence of linear and lateral trimers.

FtsZ is a bacterial protein that plays a crucial role in cytokinesis by forming the Z-ring. This ring acts as a scaffold to recruit other division proteins and guide the synthesis of septal peptidoglycan, which leads to cell constriction. In its native state, the FtsZ protein from Escherichia coli (EcFtsZ) is a multi-oligomer comprising dimers, trimers, tetramers, and hexamers in a dynamic self-association equilibrium depending on its concentration. This study employed classical methods of analytical biochemistry that included native polyacrylamide gel electrophoresis, size-exclusion chromatography, sedimentation through sucrose gradients, and chemical cross-linking with formaldehyde to characterize the EcFtsZ. The dimers, trimers, and tetramers are the most prevalent oligomers of the EcFtsZ protein; however, the trimer has been understudied compared to the dimer. In this study, we characterized uncross-linked trimers by exclusion chromatography and crosslinked trimers by sedimentation. The results of size-exclusion chromatography demonstrated that the uncross-linked trimer of EcFtsZ has a mass of 128.8 kDa and a frictional ratio f/f_o of 1.96, which coincides with the theoretical frictional ratio of 1.80 for a linear trimer. The EcFtsZ protein treated with formaldehyde resulted in a polypeptide band of 128 kDa recognized by anti-FtsZ antibodies and a frictional ratio S_max/S_20,w equal to 1.95, which agrees with the theoretical calculation of the frictional ratio of a lateral trimer. The protein-protein interaction prediction program (PEPPI) identified a contact site between subunits in the C-terminal linker region of the EcFtsZ protein, which has the potential to interfere with the recognition of the C-terminal linker by the ClpX(P) protease.