Bacterial cell division relies on a complex protein network, and since most rod-shaped bacteria divide at midcell, correct placement of the division apparatus is crucial. One of the most studied division-site selection systems is the Min system, best understood in Escherichia coli, where defects lead to aberrant cell division and thus minicells. The active component MinC is localized through MinD, a Walker-type ATPase that dimerizes and interacts with the membrane upon ATP binding. The third protein MinE stimulates MinD ATPase activity, breaking the dimer and releasing it from the membrane, leading to a pole-to-pole oscillation. In contrast, Bacillus subtilis localizes MinD through MinJ and DivIVA to regions of negative curvature without oscillation.
Previously, we described a role of the Min system in B. subtilis in divisome disassembly (1) and high MinD dynamics to allow translocation to new septa (2). Here, we aimed to investigate how MinD membrane interaction is controlled and how ATPase activity is triggered in B. subtilis.
We employed in-vitro and in-vivo methods, including biochemical analysis of purified MinD and ATPase mutants as well as single-particle tracking (SPT) of MinD fusions.
Our findings suggest that MinD ATPase activity is solely stimulated by lipid-membrane interaction, while a membrane-binding mutant (I260E) is inactive. Bio-layer interferometry (BLI) with immobilized liposomes confirmed binding of monomeric MinD mutants (G12V, K16A) independent of ATP or dimerization, while a trapped-dimer mutant (D40A) showed reduced dissociation. SPT affirmed these findings, indicating similar shifts in mobility and dwelling times.
In summary, these results support the hypothesis that frequent MinD membrane association and rapid dissociation upon dimerization generates a sharp protein gradient in B. subtilis, without the requirement for a functional MinE homolog. Quick cycling of MinD in dense areas aided by local MinJ recruitment establishes a dynamic gradient, ensuring correct Min component localization throughout the cell cycle.
van Baarle S, Bramkamp M. 2010. PLoS One 5:e9850.Feddersen H, Wurthner L, Frey E, Bramkamp M. 2021. mBio 12.