The bicyclomycin sensitivities of 38 bicyclomycin-resistant mutants of transcription termination protein rho and the location of their mutations support a structural model of rho based on the F1 ATPase

Moyse, Keith A; Knight, Jason S.; Richardson, John P.

doi:10.1006/jmbi.2000.4090

Cited by 14 publications

(13 citation statements)

References 55 publications

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“…Confirmation of these collective findings now comes from recently solved X-ray crystallographic structures of the bicyclomycin-rho complex [118]. These images validated our working homology structure model [74], and documented the 1 binding site [74,127,141] and the structural units in rho necessary for activity. Significantly, the X-ray structural data [118] have revealed an additional factor that likely contributes to 1's mechanism of action.…”

Section: Insights Into the Mechanism Of Action Of Bicyclomycin From Ssupporting

confidence: 71%

“…Adjacent to the cleft, and within a few angstroms of it, is the G helix of the neighboring subunit (residues 332-336). A similar projection of the rho 1 binding pocket has been presented by Richardson and coworkers [141].…”

Section: Derivation Of a Working Model For Rho And Projections On Thementioning

confidence: 62%

“…Three 1-resistant mutation sites, S266 and M219 and K181, whose residue was modified by 54 [132] and 55 [137], were located in the central portion of the structure near the ATP hydrolysis domain. Close by are the 1-resistant mutants D210G and L208R reported by [8,141]. However, K336, the residue modified by affinity label 56 [80], and G337, the residue when mutated to serine leads to 1 resistance [34], both resided on the same subunit but on the opposite face of rho from the ATP hydrolysis pocket.…”

Section: Derivation Of a Working Model For Rho And Projections On Thementioning

confidence: 92%

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The Molecular Basis for the Mode of Action of Bicyclomycin

Kohn

Widger

2005

CDTID

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Bicyclomycin (1) is a clinically useful antibiotic exhibiting activity against a broad spectrum of Gram-negative bacteria and against the Gram-positive bacterium, Micrococcus luteus. Bicyclomycin has been used to treat diarrhea in humans and bacterial diarrhea in calves and pigs and is marketed by Fujisawa (Osaka, Japan) under the trade name Bicozamycin. The structure of 1 is unique among antibiotics, and our studies document that its mechanism of action is novel. Early mechanistic proposals suggested that 1 reacted with nucleophiles (e.g., a protein sulfhydryl group) necessary for the remodeling the peptidoglycan assembly within the bacterial cell wall. We, however, showed that 1 targeted the rho transcription termination factor in Escherichia coli. The rho protein is integral to the expression of many gene products in E. coli and other Gram-negative bacteria, and without rho the cell losses viability. Rho is a member of the RecA-type ATPase class of enzymes that use nucleotide contacts to couple oligonucleotide translocation to ATP hydrolysis. Bicyclomycin is the only known selective inhibitor of rho. In this article, we integrate the evidence obtained from bicyclomycin structure-activity studies, site-directed mutagenesis investigations, bicyclomycin affinity labels, and biochemical and biophysical measurements with recent X-ray crystallographic images of the bicyclomycin-rho complex to define the rho antibiotic binding site and to document the pathway for rho inhibition by 1. Together, the structural and functional studies demonstrate how 1, a modest rho inhibitor, can disrupt the rho molecular machinery thereby leading to a catastrophic effect caused by the untimely overproduction of proteins not normally expressed constitutively, thus leading to a toxic effect on the cells.

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Section: Insights Into the Mechanism Of Action Of Bicyclomycin From Ssupporting

confidence: 71%

Section: Derivation Of a Working Model For Rho And Projections On Thementioning

confidence: 62%

Section: Derivation Of a Working Model For Rho And Projections On Thementioning

confidence: 92%

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The Molecular Basis for the Mode of Action of Bicyclomycin

Kohn

Widger

2005

CDTID

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“…13 Bicyclomycin was bacteriostatic with the parental strain (KD3505; Figure 1b, open circles) unless tetracycline was also present to elicit lethal synergy (Figure 1b Lethal synergy due to bicyclomycin plus inhibitors of gene expression remained unchanged once the MIC threshold was exceeded ( Figure 1a). Even increasing the bicyclomycin concentration to 10×MIC had no effect on the rate of killing, observed as survival decreasing gradually over a 3 h period ( Figure 1c).…”

Section: Lethal Synergy Involving Bicyclomycin and Inhibitors Of Genementioning

confidence: 99%

Lethal synergy involving bicyclomycin: an approach for reviving old antibiotics

Malik

Zhao³

et al. 2014

Journal of Antimicrobial Chemotherapy

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Background: One way to address the growing problem of antimicrobial resistance is to revive old compounds that may have intrinsic lethal activity that is obscured by protective factors. Bicyclomycin is an old inhibitor of the Rho transcription terminator that by itself shows little rapid lethal activity. However, bicyclomycin participates in bacteriostatic synergy, which raises the possibility that conditions for lethal synergy may exist, perhaps through a suppression of protective factors.Methods: Bicyclomycin was combined with bacteriostatic inhibitors of gene expression, and bactericidal activity was measured with several cultured Gram-negative pathogens.Results: When used alone, bicyclomycin failed to rapidly kill growing cultures of Escherichia coli; however, the additional presence of bacteriostatic concentrations of tetracycline, chloramphenicol or rifampicin led to rapid killing. Four other pathogen species, Acinetobacter baumannii, Klebsiella pneumoniae, Salmonella enterica serotype Typhimurium and Shigella dysenteriae, also exhibited enhanced killing when bicyclomycin was combined with tetracycline or rifampicin. This lethal synergy was achieved at low concentrations (slightly above the MIC) for all agents tested in combinations. Follow-up work with E. coli indicated that lethal synergy arose from a blockage of transcription elongation. Moreover, lethal synergy was reduced when bicyclomycin was added 60 min before tetracycline, suggesting that bicyclomycin induces a protective factor. Conclusions:The action of bicyclomycin illustrates the potential present in a largely abandoned antibacterial agent; it exhibits lethal synergy when coadministered with known, bacteriostatic inhibitors of gene expression. The identification of protective factors, which are currently uncharacterized, may reveal new ways to promote the lethal action of some old antibiotics.

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“…Bicyclomycin is the only known antibiotic to target Rho, and it does not affect RNA or ATP binding to Rho but does inhibit Rho-dependent transcription termination and ATP hydrolysis (167)(168)(169). Substitutions in E. coli Rho residues L208, M219, S266, and G337 conferred resistance to bicyclomycin, suggesting that Rho was the binding target of bicyclomycin (170,171).…”

Section: Termination Factor Rhomentioning

confidence: 99%

Bacterial Transcription as a Target for Antibacterial Drug Development

Yang

Lewis

2016

Microbiol Mol Biol Rev

113

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SUMMARYTranscription, the first step of gene expression, is carried out by the enzyme RNA polymerase (RNAP) and is regulated through interaction with a series of protein transcription factors. RNAP and its associated transcription factors are highly conserved across the bacterial domain and represent excellent targets for broad-spectrum antibacterial agent discovery. Despite the numerous antibiotics on the market, there are only two series currently approved that target transcription. The determination of the three-dimensional structures of RNAP and transcription complexes at high resolution over the last 15 years has led to renewed interest in targeting this essential process for antibiotic development by utilizing rational structure-based approaches. In this review, we describe the inhibition of the bacterial transcription process with respect to structural studies of RNAP, highlight recent progress toward the discovery of novel transcription inhibitors, and suggest additional potential antibacterial targets for rational drug design.

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The bicyclomycin sensitivities of 38 bicyclomycin-resistant mutants of transcription termination protein rho and the location of their mutations support a structural model of rho based on the F1 ATPase

Cited by 14 publications

References 55 publications

The Molecular Basis for the Mode of Action of Bicyclomycin

The Molecular Basis for the Mode of Action of Bicyclomycin

Lethal synergy involving bicyclomycin: an approach for reviving old antibiotics

Bacterial Transcription as a Target for Antibacterial Drug Development

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