Mechanism for kinetic polarity revealed by FtsZ from a cell wall-less bacteriumSpiroplasma

Abstract

AbstractFtsZ, the tubulin homolog essential for bacterial cell division, assembles the Z-ring at the division site, and directs peptidoglycan synthesis by its treadmilling action. A key unanswered question is how FtsZ achieves its kinetic polarity that drives treadmilling. To obtain insights into fundamental features of FtsZ assembly dynamics independent of peptidoglycan synthesis, we report the characterization of FtsZ from the cell wall-less bacteriumSpiroplasma melliferum(SmFtsZ). SmFtsZ was a slow GTPase and required higher critical concentration (CC) for polymerization as compared toEscherichia coliFtsZ (EcFtsZ). In FtsZs, a conformational switch from R- to T-state favors polymerization. In the crystal structures of SmFtsZ, conformation of the GTP-bound N-terminal domain (NTD) was in the T-state, while the relative orientation of the NTD and the C-terminal domain (CTD) matched the R-state. Hence, the crystal structures represent a conformational state that facilitates preferential binding of the NTD of the monomeric GTP-bound SmFtsZ to the CTD-exposed end of SmFtsZ filament. CTD of the nucleotide-bound monomer cannot bind to the NTD-exposed end of the filament unless relative rotation of the domains leads to cleft opening. The mutation F224M in SmFtsZ cleft resulted in higher GTPase activity and lower CC, whereas the corresponding M225F in EcFtsZ resulted in cell division defects inE. coli. These results suggest that relative rotation of the domains leading to cleft opening is a rate-limiting step of polymerization. This slows the addition of monomers to the NTD-exposed end of the filament in comparison to the CTD end, thus explaining kinetic polarity.