2 research outputs found
Interactions of Bacterial Cell Division Protein FtsZ with C8-Substituted Guanine Nucleotide Inhibitors. A Combined NMR, Biochemical and Molecular Modeling Perspective
FtsZ
is the key protein of bacterial cell-division and target for
new antibiotics. Selective inhibition of FtsZ polymerization without
impairing the assembly of the eukaryotic homologue tubulin was demonstrated
with C8-substituted guanine nucleotides. By combining NMR techniques
with biochemical and molecular modeling procedures, we have investigated
the molecular recognition of C8-substituted-nucleotides by FtsZ from <i>Methanococcus jannaschii</i> (Mj-FtsZ) and <i>Bacillus
subtilis</i> (Bs-FtsZ). STD epitope mapping and trNOESY bioactive
conformation analysis of each nucleotide were employed to deduce differences
in their recognition mode by each FtsZ species. GMP binds in the same
anti conformation as GTP, whereas 8-pyrrolidino-GMP binds in the syn
conformation. However, the anti conformation of 8-morpholino-GMP is
selected by Bs-FtsZ, while Mj-FtsZ binds both anti- and syn-geometries.
The inhibitory potencies of the C8-modified-nucleotides on the assembly
of Bs-FtsZ, but not of Mj-FtsZ, correlate with their binding affinities.
Thus, MorphGTP behaves as a nonhydrolyzable analog whose binding induces
formation of Mj-FtsZ curved filaments, resembling polymers formed
by the inactive forms of this protein. NMR data, combined with molecular
modeling protocols, permit explanation of the mechanism of FtsZ assembly
impairment by C8-substituted GTP analogs. The presence of the C8-substituent
induces electrostatic remodeling and small structural displacements
at the association interface between FtsZ monomers to form filaments,
leading to complete assembly inhibition or to formation of abnormal
FtsZ polymers. The inhibition of bacterial Bs-FtsZ assembly may be
simply explained by steric clashes of the C8-GTP-analogs with the
incoming FtsZ monomer. This information may facilitate the design
of antibacterial FtsZ inhibitors replacing GTP
Synthetic Inhibitors of Bacterial Cell Division Targeting the GTP-Binding Site of FtsZ
Cell division protein FtsZ is the
organizer of the cytokinetic
Z-ring in most bacteria and a target for new antibiotics. FtsZ assembles
with GTP into filaments that hydrolyze the nucleotide at the association
interface between monomers and then disassemble. We have replaced
FtsZ’s GTP with non-nucleotide synthetic inhibitors of bacterial
division. We searched for these small molecules among compounds from
the literature, from virtual screening (VS), and from our in-house
synthetic library (UCM), employing a fluorescence anisotropy primary
assay. From these screens we have identified the polyhydroxy aromatic
compound UCM05 and its simplified analogue UCM44 that specifically
bind to <i>Bacillus subtilis</i> FtsZ monomers with micromolar
affinities and perturb normal assembly, as examined with light scattering,
polymer sedimentation, and negative stain electron microscopy. On
the other hand, these ligands induce the cooperative assembly of nucleotide-devoid
archaeal FtsZ into distinct well-ordered polymers, different from
GTP-induced filaments. These FtsZ inhibitors impair localization of
FtsZ into the Z-ring and inhibit bacterial cell division. The chlorinated
analogue UCM53 inhibits the growth of clinical isolates of antibiotic-resistant <i>Staphylococcus aureus</i> and <i>Enterococcus faecalis</i>. We suggest that these interfacial inhibitors recapitulate binding
and some assembly-inducing effects of GTP but impair the correct structural
dynamics of FtsZ filaments and thus inhibit bacterial division, possibly
by binding to a small fraction of the FtsZ molecules in a bacterial
cell, which opens a new approach to FtsZ-based antibacterial drug
discovery