Synthetic developmental regulator MciZ targets FtsZ across Bacillus species and inhibits bacterial division

Summary Cell division in most bacteria is directed by FtsZ, a conserved tubulin‐like GTPase that assembles forming the cytokinetic Z‐ring and constitutes a target for the discovery of new antibiotics. The developmental regulator MciZ, a 40‐amino acid peptide endogenously produced during Bacillus subtilis sporulation, halts cytokinesis in the mother cell by inhibiting FtsZ. The crystal structure of a FtsZ:MciZ complex revealed that bound MciZ extends the C‐terminal β‐sheet of FtsZ blocking its assembly interface. Here we demonstrate that exogenously added MciZ specifically inhibits B. subtilis cell division, sporulation and germination, and provide insight into MciZ molecular recognition by FtsZ from different bacteria. MciZ and FtsZ form a complex with sub‐micromolar affinity, analyzed by analytical ultracentrifugation, laser biolayer interferometry and isothermal titration calorimetry. Synthetic MciZ analogs, carrying single amino acid substitutions impairing MciZ β‐strand formation or hydrogen bonding to FtsZ, show a gradual reduction in affinity that resembles their impaired activity in bacteria. Gene sequences encoding MciZ spread across genus Bacillus and synthetic MciZ slows down cell division in Bacillus species, including pathogenic Bacillus cereus and Bacillus anthracis. Moreover, B. subtilis MciZ is recognized by the homologous FtsZ from Staphylococcus aureus and inhibits division when it is expressed into S. aureus cells. Synthetic peptide MciZ is a specific inhibitor of B. subtilis division, sporulation and germination that binds with high affinity to key cell division protein FtsZ, blocking its assembly and the formation of the cytokinetic Z‐ring. MciZ activity extends to other Bacillus, including pathogenic B. cereus – anthracis. MciZ is also recognized by FtsZ from S. aureus and intracellularly expressed MiciZ inhibits S. aureus division. Our results suggest the possibility of designing MciZ‐based peptide antibacterials.