FtsK with a unique N-terminal extension is involved in coordinating the final steps of chromosome segregation with asymmetric division in mycobacterial cells.

In this study, we identify and functionally characterize a previously unrecognized, conserved N-terminal extension of mycobacterial FtsK (nFtsK) that contributes to the coordination of chromosome segregation with asymmetric cell division. Although FtsK is broadly conserved in bacteria as a late-stage DNA translocase and divisome component, mycobacterial FtsK uniquely contains a long, positively charged, intrinsically disordered N-terminal region. We show that this extension is dispensable for core FtsK functions-including septal localization, DNA translocation, and completion of cell division-but instead modulates the spatial confinement, temporal regulation, and interaction specificity of these processes. Mechanistically, nFtsK mediates specific protein interactions, binds the anionic phospholipids cardiolipin and phosphatidic acid, and exhibits intrinsic affinity for negatively curved membranes. Together, our findings uncover a lineage-specific adaptation that fine-tunes a conserved molecular machine, enhancing the robustness of asymmetric cell division in mycobacteria.IMPORTANCEFaithful cell division requires precise coordination between chromosome segregation and septum formation. Mycobacteria lack the canonical spatial regulators found in many bacteria, raising the question of how this coordination is achieved in asymmetrically dividing cells. Here, we identify a lineage-specific N-terminal extension of the DNA translocase FtsK that contributes to spatial control during mycobacterial division. This intrinsically disordered region associates with anionic lipids and curved membrane regions at the septum, coupling chromosome translocation to septal maturation. Our findings reveal how mycobacteria adapt conserved DNA translocation machinery to their unconventional division mode.