Dataset Information

Xanthocillin X Effectively Kills A.Baumannii via Dysregulation of Heme Biosynthesis

ABSTRACT: Isonitrile natural products exhibit promising antibacterial activities, however, their mode of action (MoA) remains largely unknown. Based on the nanomolar potency of xanthocillin X (Xan) against diverse difficult-to-treat Gram-negative bacteria, including the critical priority pathogen Acinetobacter baumannii, we performed in-depth studies to decipher its MoA. While neither metal binding nor cellular protein targets were relevant for Xan´s antibiotic effects, sequencing of resistant strains revealed a conserved mutation in the heme biosynthesis enzyme porphobilinogen synthase (PbgS). This mutation caused impaired enzymatic efficiency indicative of reduced heme production. This discovery led to the validation of an untapped mechanism by which direct heme sequestration of Xan prevents its binding into cognate enzyme pockets resulting in uncontrolled cofactor biosynthesis, accumulation of porphyrins and corresponding stress with deleterious effects for bacterial viability. Thus, Xan represents a promising antibiotic displaying activity even against multidrug resistant strains while exhibiting low toxicity to human cells.

INSTRUMENT(S): Orbitrap Fusion, Q Exactive HF

ORGANISM(S): Escherichia Coli Acinetobacter Baumannii Atcc 19606 = Cip 70.34 = Jcm 6841

SUBMITTER: Ines Hübner

LAB HEAD: Stephan A. Sieber

PROVIDER: PXD021400 | Pride | 2021-09-09

REPOSITORIES: Pride

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Publications

Broad Spectrum Antibiotic Xanthocillin X Effectively Kills Acinetobacter baumannii via Dysregulation of Heme Biosynthesis.

Hübner Ines I Shapiro Justin A JA Hoßmann Jörn J Drechsel Jonas J Hacker Stephan M SM Rather Philip N PN Pieper Dietmar H DH Wuest William M WM Sieber Stephan A SA

ACS central science 20210120 3

Isonitrile natural products exhibit promising antibacterial activities. However, their mechanism of action (MoA) remains largely unknown. Based on the nanomolar potency of xanthocillin X (Xan) against diverse difficult-to-treat Gram-negative bacteria, including the critical priority pathogen Acinetobacter baumannii, we performed in-depth studies to decipher its MoA. While neither metal binding nor cellular protein targets were detected as relevant for Xan's antibiotic effect ...[more]

PMID: 33791430

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Project description:The response regulator HrrA belonging to the HrrSA two-component system (previously named CgtSR11) is known to be repressed by the global iron-dependent regulator DtxR in Corynebacterium glutamicum. Sequence analysis indicated an involvement of the HrrSA system in heme-dependent gene expression. Growth experiments revealed that the non-pathogenic soil bacterium C. glutamicum is able to use hemin or hemoglobin as sole iron source. In DNA microarray analyses a putative operon encoding the hemin-binding protein HtaA and the putative hemin ABC transporter HmuTUV showed a strong upregulation in heme-grown cells. Deletion of the hmu operon clearly affects heme utilization, but does not completely abolish growth on heme or hemoglobin. As a central part of this study, we investigated the regulon of the response regulator HrrA via comparative transcriptome analysis of a hrrA deletion mutant and C. glutamicum wild type in combination with DNA-protein interaction studies with purified HrrA protein. Our data provide evidence for a heme-dependent transcriptional activation of heme oxygenase (hmuO), an enzyme involved in the utilization of heme as iron source. Besides hmuO, HrrA was shown to activate the expression of heme-containing components of the respiratory chain, namely ctaD and the ctaE-qcrCAB operon encoding subunits I and III of cytochrome aa3 oxidase and three subunits of the cytochrome bc1 complex. Furthermore, HrrA represses almost all genes involved in heme biosynthesis, including glutamyl-tRNA reductase (hemA), uroporphyrinogen decarboxylase (hemE), and ferrochelatase (hemH). Thus, our data clearly emphasize a central role of the HrrSA system in the control of heme homeostasis in C. glutamicum. Three biological replicates of each experiment were performed. Experiment 1: Transcriptome comparison of wild type grown und FeSO4 or heme as iron source; Exp. 2: WT vs. hrrA deletion mutant grown on FeSO4; Exp. 3: WT vs. hrrA mutant grown on heme. For analysis via DNA microarraysose RNA was isolated from exponentially growing cells cultivated in CgXII medium containing glucose as carbon source and either 2.5 uM FeSO4 or 2.5 uM heme as iron source.

Dataset Information

Xanthocillin X Effectively Kills A.Baumannii via Dysregulation of Heme Biosynthesis

Publications

Broad Spectrum Antibiotic Xanthocillin X Effectively Kills <i>Acinetobacter baumannii</i> <i>via</i> Dysregulation of Heme Biosynthesis.

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