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Publication Abstract from PubMed

The essential bacterial division protein FtsZ uses GTP binding and hydrolysis to assemble into dynamic filaments that treadmill around the Z-ring, guiding septal wall synthesis and cell division. FtsZ is a structural homolog of tubulin and a target for discovering new antibiotics. Here, using FtsZ from the pathogen S aureus (SaFtsZ), we reveal that, prior to assembly, FtsZ monomers require nucleotide binding for folding; this is possibly relevant to other mesophilic FtsZs. Apo-SaFtsZ is essentially unfolded, as assessed by nuclear magnetic resonance and circular dichroism. Binding of GTP (>/= 1mM) dramatically shifts the equilibrium towards the active folded protein. Supportingly, SaFtsZ refolded with GDP crystallizes in a native structure. Apo-SaFtsZ also folds with 3.4 M glycerol, enabling high-affinity GTP binding (KD 20 nM determined by isothermal titration calorimetry) similar to thermophilic stable FtsZ. Other stabilizing agents that enhance nucleotide binding include ethylene glycol, trimethylamine N-oxide (TMAO), and several bacterial osmolytes. High salt stabilizes SaFtsZ without bound nucleotide in an inactive twisted conformation. We identified a cavity behind the SaFtsZ-GDP nucleotide-binding pocket that harbors different small compounds, which is available for extended nucleotide-replacing inhibitors. Furthermore, we devised a competition assay to detect any inhibitors that overlap the nucleotide site of SaFtsZ, or Escherichia coli FtsZ, employing osmolyte-stabilized apo-FtsZs and the specific fluorescence anisotropy change of mant-GTP upon dissociation from the protein. This robust assay provides a basis to screening for high affinity GTP-replacing ligands, which combined with structural studies and phenotypic profiling should facilitate development of a next generation of FtsZ-targeting antibacterial inhibitors.

Nucleotide-induced folding of cell division protein FtsZ from Staphylococcus aureus.,Huecas S, Canosa-Valls AJ, Araujo-Bazan L, Ruiz FM, Laurents DV, Fernandez-Tornero C, Andreu JM FEBS J. 2020 Jan 29. doi: 10.1111/febs.15235. PMID:31997533^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.