ABSTRACT
Inhibition of FtsZ assembly has been found to stall bacterial cell division. Here, we report the identification of a potent carbocyclic curcumin analogue (2d) that inhibits Bacillus subtilis 168 cell proliferation by targeting the assembly of FtsZ. 2d also showed potent inhibitory activity (minimum inhibitory concentrations of 2-4 mg/L) against several clinically important species of Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus. In addition, 2d displayed a significantly reduced inhibitory effect on human cervical cancer cells in comparison to its effect on bacterial cells. Using live cell imaging of GFP-FtsZ by confocal microscopy, 2d was found to rapidly perturb the cytokinetic FtsZ rings in Bacillus subtilis cells. The immunofluorescence imaging of FtsZ also showed that 2d destroyed the Z-ring in bacteria within 5 min. Prolonged treatment with 2d produced filamentous bacteria, but 2d had no detectable effect either on the nucleoids or on the membrane potential of bacteria. 2d inhibited FtsZ assembly in vitro, whereas it had minimal effects on tubulin assembly. Interestingly, 2d strongly enhanced the GTPase activity of FtsZ and reduced the GTPase activity of tubulin. Furthermore, 2d bound to purified FtsZ with a dissociation constant of 4.0 ± 1.1 µM, and the binding of 2d altered the secondary structures of FtsZ. The results together suggested that the non-natural curcumin analogue 2d possesses powerful antibacterial activity against important pathogenic bacteria, and the evidence indicates that 2d inhibits bacterial proliferation by targeting FtsZ.
Subject(s)
Anti-Bacterial Agents/pharmacology , Bacillus subtilis/drug effects , Bacterial Proteins/antagonists & inhibitors , Curcumin/pharmacology , Cytoskeletal Proteins/antagonists & inhibitors , Methicillin-Resistant Staphylococcus aureus/drug effects , Animals , Anti-Bacterial Agents/chemical synthesis , Bacillus subtilis/genetics , Bacillus subtilis/growth & development , Bacillus subtilis/ultrastructure , Bacterial Proteins/biosynthesis , Bacterial Proteins/genetics , Binding Sites , Cloning, Molecular , Curcumin/analogs & derivatives , Curcumin/chemical synthesis , Cyclization , Cytoskeletal Proteins/biosynthesis , Cytoskeletal Proteins/genetics , Escherichia coli/genetics , Escherichia coli/metabolism , Gene Expression , Genes, Reporter , Goats , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , HeLa Cells , Humans , Methicillin-Resistant Staphylococcus aureus/genetics , Methicillin-Resistant Staphylococcus aureus/growth & development , Methicillin-Resistant Staphylococcus aureus/ultrastructure , Microbial Sensitivity Tests , Molecular Imaging , Protein Binding , Protein Structure, Secondary , Recombinant Proteins/biosynthesis , Recombinant Proteins/genetics , Species Specificity , Tubulin/genetics , Tubulin/metabolismABSTRACT
Filamenting temperature-sensitive mutant Z (FtsZ), an essential cell division protein in bacteria, has recently emerged as an important and exploitable antibacterial target. Cytokinesis in bacteria is regulated by the assembly dynamics of this protein, which is ubiquitously present in eubacteria. The perturbation of FtsZ assembly has been found to have a deleterious effect on the cytokinetic machinery and, in turn, upon cell survival. FtsZ is highly conserved among prokaryotes, offering the possibility of broad-spectrum antibacterial agents, while its limited sequence homology with tubulin (an essential protein in eukaryotic mitosis) offers the possibility of selective toxicity. This review aims to summarize current knowledge regarding the mechanism of action of FtsZ, and to highlight existing attempts toward the development of clinically useful inhibitors.
Subject(s)
Anti-Bacterial Agents/pharmacology , Bacteria/drug effects , Bacterial Proteins/antagonists & inhibitors , Bacterial Proteins/metabolism , Cytoskeletal Proteins/antagonists & inhibitors , Cytoskeletal Proteins/metabolism , Animals , Bacteria/cytology , Bacteria/metabolism , Bacterial Infections/drug therapy , Bacterial Infections/microbiology , Bacterial Proteins/chemistry , Cytokinesis/drug effects , Cytoskeletal Proteins/chemistry , Drug Discovery , Humans , MiceABSTRACT
Synthesis and biological evaluation of some 5-ethoxycarbonyl-6-isopropylamino-4-(substitutedphenyl)aminopyrimidines have been achieved by cyclization of N-[2-ethoxycarbonyl-2-cyano-1-(isopropylamino)vinyl] formamidine in presence of dry HCl in dioxane followed by nucleophilic substitution of 4-chloro group with substituted aromatic amine or phenoxide. Target compounds were evaluated for their analgesic and anti-inflammatory potential by known experimental models. Some of the compounds emerged out as more potent than standard drugs. Very low ulcer index was observed for the potent compounds.
