Chrysophaentins are competitive inhibitors of FtsZ and inhibit Z-ring formation in live bacteria.
The bacterial cell division protein FtsZ polymerizes in a GTP-dependent manner to form a Z-ring that marks the plane of division. As a validated antimicrobial target, considerable efforts have been devoted to identify small molecule FtsZ inhibitors. We recently discovered the chrysophaentins, a novel suite of marine natural products that inhibit FtsZ activity in vitro. These natural products along with a synthetic hemi-chrysophaentin exhibit strong antimicrobial activity toward a broad spectrum of Gram-positive pathogens. To define their mechanisms of FtsZ inhibition and determine their in vivo effects in live bacteria, we used GTPase assays and fluorescence anisotropy to show that hemi-chrysophaentin competitively inhibits FtsZ activity. Furthermore, we developed a model system using a permeable Escherichia coli strain, envA1, together with an inducible FtsZ-yellow fluorescent protein construct to show by fluorescence microscopy that both chrysophaentin A and hemi-chrysophaentin disrupt Z-rings in live bacteria. We tested the E. coli system further by reproducing phenotypes observed for zantrins Z1 and Z3, and demonstrate that the alkaloid berberine, a reported FtsZ inhibitor, exhibits auto-fluorescence, making it incompatible with systems that employ GFP or YFP tagged FtsZ. These studies describe unique examples of nonnucleotide, competitive FtsZ inhibitors that disrupt FtsZ in vivo, together with a model system that should be useful for in vivo testing of FtsZ inhibitor leads that have been identified through in vitro screens but are unable to penetrate the Gram-negative outer membrane.
Laboratory of Bioorganic Chemistry, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, United States.
Jessica L
JL
Keffer
Sonia
S
Huecas
Jared T
JT
Hammill
Peter
P
Wipf
Jose M
JM
Andreu
Carole A
CA
Bewley
D000581:Amidohydrolases / Q000235:genetics / Q000378:metabolism; D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D001593:Benzyl Compounds / Q000138:chemical synthesis / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D001599:Berberine / Q000737:chemistry / Q000494:pharmacology; D003598:Cytoskeletal Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D004926:Escherichia coli / Q000187:drug effects / Q000378:metabolism; D004988:Ethers, Cyclic / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology; D020558:GTP Phosphohydrolases / Q000378:metabolism; D006094:Gram-Positive Bacteria / Q000187:drug effects; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D001593:Benzyl Compounds; D003598:Cytoskeletal Proteins; D004988:Ethers, Cyclic; C071525:FtsZ protein, Bacteria; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; C587119:chrysophaentin A; C587121:hemi-chrysophaetin; C054562:yellow fluorescent protein, Bacteria; D001599:Berberine; D000581:Amidohydrolases; C097307:UDP-3-O-acyl-N-acetylglucosamine deacetylase; D020558:GTP Phosphohydrolases
Antibiotics; Bacterial cytoskeleton; Drug resistant bacteria; Natural products