Investigations into the Antibacterial Mechanism of Action of Viridicatumtoxins
- 4 June 2020
- journal article
- research article
- Published by American Chemical Society (ACS) in ACS Infectious Diseases
- Vol. 6 (7), 1759-1769
- https://doi.org/10.1021/acsinfecdis.0c00031
Abstract
Viridicatumtoxins are a rare class of tetracycline-like antibiotics that strongly inhibit drug-resistant Gram-positive bacteria. Although reported to exhibit in vitro inhibition activity to undecaprenyl pyrophosphate synthase (UPPS), an essential enzyme in bacterial cell wall synthesis, the biological targets and mechanism of action of viridicatumtoxins especially the drug-target interactions remain largely unknown. In this study, the structure of Enterococcus faecalis UPPS (EfaUPPS) was first determined, uncovering that EfaUPPS can not only form a typical functional dimer but also an unexpected atypical dimer. We then observed that viridicatumtoxins A (VirA) and B (VirB) are able to bind to UPPSs of E. faecalis, S. aureus, and E. coli in a direct and high-affinity manner as evidenced by in vitro enzyme inhibition assay, surface plasmon resonance (SPR) binding analysis, and in vivo growth inhibition assay, demonstrating that viridicatumtoxins exert antibacterial effects through UPPS binding. The key amino acid residues involved in the interactions with VirA and VirB in EfaUPPS binding pocket were revealed by molecular docking studies, and further validated by site-directed mutagenesis. A single mutation of EfaUPPS at D29A, N31A and R42A can obviously increase their affinities to VirA, while a single mutation at W228A conferred significant resistance to VirA. Moreover, translation inhibition assay showed that VirA and VirB can weakly inhibit E. coli 70S ribosome. The weak inhibition of ribosome was proposed to be attributed to steric hindrance between viridicatumtoxin ring F and 70S ribosome helix 34 by molecular docking study. Our structural, biochemical and computational investigations on the interactions of viridicatumtoxins with UPPS and 70S ribosome not only disclosed the potential biological targets of viridicatumtoxins, but also provided a theoretical basis for structural optimization to make new viridicatumtoxin derivatives with improved antimicrobial activities.Keywords
Funding Information
- State Key Laboratory of Natural Medicines (SKLNMKF202004)
- Natural Science Foundation of Jiangsu Province (BK20190798)
- National Natural Science Foundation of China (81903526, 81991523)
- U.S. Department of Health and Human Services (P20 GM103641)
- National Science Foundation (OIA 1655740)
- Jiangsu Specially-Appointed Professor Talent Program
- Fok Ying Tung Education Foundation (171033)
This publication has 30 references indexed in Scilit:
- Total Synthesis and Structural Revision of Viridicatumtoxin BAngewandte Chemie, 2013
- Target- and Resistance-Based Mechanistic Studies with TP-434, a Novel Fluorocycline AntibioticAntimicrobial Agents and Chemotherapy, 2012
- Discovery of Antagonists of PqsR, a Key Player in 2-Alkyl-4-quinolone-Dependent Quorum Sensing in Pseudomonas aeruginosaCell Chemical Biology, 2012
- Features and development of CootActa crystallographica. Section D, Structural biology, 2010
- PHENIX: a comprehensive Python-based system for macromolecular structure solutionActa crystallographica. Section D, Structural biology, 2010
- AutoDock4 and AutoDockTools4: Automated docking with selective receptor flexibilityJournal of Computational Chemistry, 2009
- Functional, Biophysical, and Structural Bases for Antibacterial Activity of TigecyclineAntimicrobial Agents and Chemotherapy, 2006
- [20] Processing of X-ray diffraction data collected in oscillation modeMethods in Enzymology, 1997
- The CCP4 suite: programs for protein crystallographyActa crystallographica. Section D, Structural biology, 1994
- Viridicatumtoxin, a new mycotoxin from Penicillium viridicatum westlingToxicology and Applied Pharmacology, 1973