Dataset Information

Covalent DNA Binding Is Essential for Gram-Negative Antibacterial Activity of Broad Spectrum Pyrrolobenzodiazepines.

ABSTRACT: It is urgent to find new antibiotic classes against multidrug-resistant bacteria as the rate of discovery of new classes of antibiotics has been very slow in the last 50 years. Recently, pyrrolobenzodiazepines (PBDs) with a C8-linked aliphatic-heterocycle have been identified as a new broad-spectrum antibiotic class with activity against Gram-negative bacteria. The active imine moiety of the reported lead pyrrolobenzodiazepine compounds was replaced with amide to obtain the non-DNA binding and noncytotoxic dilactam analogues to understand the structure-activity relationship further and improve the safety potential of this class. The synthesised compounds were tested against panels of multidrug-resistant Gram-positive and Gram-negative bacteria, including WHO priority pathogens. Minimum inhibitory concentrations for the dilactam analogues ranged from 4 to 32 mg/L for MDR Gram-positive bacteria, compared to 0.03 to 2 mg/L for the corresponding imine analogues. At the same time, they were found to be inactive against MDR Gram-negative bacteria, with a MIC > 32 mg/L, compared to a MIC of 0.5 to 32 mg/L for imine analogues. A molecular modelling study suggests that the lack of imine functionality also affects the interaction of PBDs with DNA gyrase. This study suggests that the presence of N10-C11 imine moiety is crucial for the broad-spectrum activity of pyrrolobenzodiazepines.

SUBMITTER: Picconi P

PROVIDER: S-EPMC9774941 | biostudies-literature | 2022 Dec

REPOSITORIES: biostudies-literature

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Covalent DNA Binding Is Essential for Gram-Negative Antibacterial Activity of Broad Spectrum Pyrrolobenzodiazepines.

Picconi Pietro P Hind Charlotte K CK Sutton J Mark JM Rahman Khondaker Miraz KM

Antibiotics (Basel, Switzerland) 20221207 12

It is urgent to find new antibiotic classes against multidrug-resistant bacteria as the rate of discovery of new classes of antibiotics has been very slow in the last 50 years. Recently, pyrrolobenzodiazepines (PBDs) with a C8-linked aliphatic-heterocycle have been identified as a new broad-spectrum antibiotic class with activity against Gram-negative bacteria. The active imine moiety of the reported lead pyrrolobenzodiazepine compounds was replaced with amide to obtain the non-DNA binding and n ...[more]

PMID: 36551427

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Project description:The increasing prevalence of drug-resistant bacteria has significantly diminished the effectiveness of antibiotics in clinical settings, leading to the emergence of untreatable bacterial infections. To address this public health challenge, the gut microbiome represents a promising source of novel antimicrobial therapeutics. In this study, we screened mouse intestinal isolates for growth inhibitory activity against the human enteric pathogen Vibrio cholerae and identified a strain of spore-forming Bacillus velezensis, named BVM7, that produced a potent antibiotic with activity against V. cholerae and a broad spectrum of enteric and opportunistic pathogens. Characterization of the antimicrobial compounds produced by BVM7 revealed that they were primarily secreted antimicrobial peptides (AMPs) produced during stationary-phase growth. Furthermore, our results showed that introducing either BVM7 vegetative cells or spores into mice precolonized with V. cholerae or Enterococcus faecalis significantly reduced the burden of infection. Interestingly, we also observed that BVM7 was sensitive to a group of Lactobacillus probiotic strains and that inoculation of Lactobacilli could eliminate BVM7 and potentially restore the native gut microbiome. These findings highlight the potential of bacteria from the gut microbiome as a source for novel antimicrobial compounds and a tool for managing bacterial infections by in situ bio-delivery of multiple AMPs. IMPORTANCE The rise of antibiotic-resistant pathogens poses a challenge to public health. The gut microbiome presents a promising source of new antimicrobials and treatments. By screening murine gut commensals, we found a spore-forming Bacillus velezensis strain, BVM7, that exhibited antimicrobial activity toward a wide array of enteric and opportunistic bacterial pathogens. In addition to showing that this killing effect occurred through the action of secreted antimicrobial peptides (AMPs), we demonstrate that BVM7 vegetative cells and spores can be used to treat infections of both Gram-positive and Gram-negative pathogens in vivo. By expanding our knowledge of the antimicrobial properties of bacteria in the gut microbiome, we hope to contribute insights for developing novel drugs and therapeutic interventions.

Project description:The development of new antibiotics is imperative to fight increasing mortality rates connected to infections caused by multidrug-resistant (MDR) bacteria. In this context, Gram-negative pathogens listed in the WHO priority list are particularly problematic. Darobactin is a ribosomally produced and post-translationally modified bicyclic heptapeptide antibiotic selectively killing Gram-negative bacteria by targeting the outer membrane protein BamA. The native darobactin A producer Photorhabdus khanii HGB1456 shows very limited production under laboratory cultivation conditions. Herein, we present the design and heterologous expression of a synthetically engineered darobactin biosynthetic gene cluster (BGC) in Escherichia coli to reach an average darobactin A production titre of 13.4 mg L-1. Rational design of darA variants, encoding the darobactin precursor peptide with altered core sequences, resulted in the production of 13 new 'non-natural' darobactin derivatives and 4 previously hypothetical natural darobactins. One of the non-natural compounds, darobactin 9, was more potent than darobactin A, and showed significantly improved activity especially against Pseudomonas aeruginosa (0.125 μg mL-1) and Acinetobacter baumannii (1-2 μg mL-1). Importantly, it also displayed superior activity against MDR clinical isolates of E. coli (1-2 μg mL-1) and Klebsiella pneumoniae (1-4 μg mL-1). Independent deletions of genes from the darobactin BGC showed that only darA and darE, encoding a radical forming S-adenosyl-l-methionine-dependent enzyme, are required for darobactin formation. Co-expression of two additional genes associated with the BGCs in hypothetical producer strains identified a proteolytic detoxification mechanism as a potential self-resistance strategy in native producers. Taken together, we describe a versatile heterologous darobactin platform allowing the production of unprecedented active derivatives in good yields, and we provide first experimental evidence for darobactin biosynthesis processes.

Dataset Information

Covalent DNA Binding Is Essential for Gram-Negative Antibacterial Activity of Broad Spectrum Pyrrolobenzodiazepines.

Publications

Covalent DNA Binding Is Essential for Gram-Negative Antibacterial Activity of Broad Spectrum Pyrrolobenzodiazepines.

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