Different Biosynthetic Pathways to Fosfomycin in Pseudomonas syringae and Streptomyces Species

Institute of Genomic Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
Antimicrobial Agents and Chemotherapy (Impact Factor: 4.45). 05/2012; 56(8):4175-83. DOI: 10.1128/AAC.06478-11
Source: PubMed

ABSTRACT Fosfomycin is a wide-spectrum antibiotic that is used clinically to treat acute cystitis in the United States. The compound is produced by several strains of streptomycetes and pseudomonads. We sequenced the biosynthetic gene cluster responsible for fosfomycin production in Pseudomonas syringae PB-5123. Surprisingly, the biosynthetic pathway in this organism is very different from that in Streptomyces fradiae and Streptomyces wedmorensis. The pathways share the first and last steps, involving conversion of phosphoenolpyruvate to phosphonopyruvate (PnPy) and 2-hydroxypropylphosphonate (2-HPP) to fosfomycin, respectively, but the enzymes converting PnPy to 2-HPP are different. The genome of P. syringae PB-5123 lacks a gene encoding the PnPy decarboxylase found in the Streptomyces strains. Instead, it contains a gene coding for a citrate synthase-like enzyme, Psf2, homologous to the proteins that add an acetyl group to PnPy in the biosynthesis of FR-900098 and phosphinothricin. Heterologous expression and purification of Psf2 followed by activity assays confirmed the proposed activity of Psf2. Furthermore, heterologous production of fosfomycin in Pseudomonas aeruginosa from a fosmid encoding the fosfomycin biosynthetic cluster from P. syringae PB-5123 confirmed that the gene cluster is functional. Therefore, two different pathways have evolved to produce this highly potent antimicrobial agent.

Download full-text


Available from: Bradley S Evans, Mar 18, 2014
  • [Show abstract] [Hide abstract]
    ABSTRACT: Peptidoglycan is the main component of the bacterial cell wall. It is a complex, three-dimensional mesh that surrounds the entire cell and is composed of strands of alternating glycan units cross-linked by short peptides. Its biosynthetic machinery has been, for the past five decades, a preferred target for the discovery of antibacterials. Synthesis of the peptidoglycan occurs sequentially within three cellular compartments (cytoplasm, membrane, and periplasm), and inhibitors of proteins that catalyze each stage have been identified, although not all are applicable for clinical use. A number of these antimicrobials, however, have been rendered inactive by resistance mechanisms. The employment of structural biology techniques has been instrumental in the understanding of such processes, as well as the development of strategies to overcome them. This review aims at providing an overview of resistance mechanisms developed towards antibiotics that target bacterial cell wall precursors and its biosynthetic machinery. Strategies towards the development of novel inhibitors that could overcome resistance are also discussed.
    Protein Science 03/2014; 23(3). DOI:10.1002/pro.2414 · 2.86 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Bacterial infections caused by antibiotic resistant isolates have become a major health problem in recent years since they are very difficult to treat, leading to an increase in morbidity and mortality. Fosfomycin is a broad spectrum bactericidal antibiotic that inhibits cell wall biosynthesis in both Gram negative and Gram positive bacteria. This antibiotic has a unique mechanism of action and inhibits the initial step in peptidoglycan biosynthesis by blocking the enzyme MurA. Fosfomycin has been used successfully for the treatment of urinary tract infections over a long time, but the increased emergence of antibiotic resistance has made fosfomycin a suitable candidate for the treatment of infections caused by multidrug-resistant pathogens, especially in combination with other therapeutic partners. The acquisition of fosfomycin resistance could threaten the reintroduction of this antibiotic for the treatment of bacterial infection. Here we analyse the mechanism of action and molecular mechanisms for the development of fosfomycin resistance, including the modification of the antibiotic target, reduced antibiotic uptake and antibiotic inactivation. In addition, we describe the role of each pathway in clinical isolates.
    06/2013; 2(1). DOI:10.3390/antibiotics20x000x
  • [Show abstract] [Hide abstract]
    ABSTRACT: ExpA (GacA) is a global response regulator that controls the expression of major virulence genes such as those encoding plant cell wall degrading enzymes (PCWDEs) in the model soft rot phytopathogen Pectobacterium wasabiae SCC3193. Several studies in pectobacteria as well as in related phytopathogenic γ-proteobacteria such as Dickeya and Pseudomonas suggest that the control of virulence by ExpA and its homologues is executed partly by modulating the activity of RsmA, an RNA-binding post-transcriptional regulator. To elucidate the extent of the overlap between the ExpA and RsmA regulons in P. wasabiae we characterized both regulons by microarray analysis. To do this, we compared the transcriptomes of the wild-type strain, an expA mutant, an rsmA mutant, and an expA rsmA double mutant. The microarray data of selected virulence related genes were confirmed through quantitative RT-PCR (qPCR). Subsequently, assays were performed to link the observed transcriptome differences to changes in bacterial phenotypes such as growth, motility, PCWDE production, and virulence in planta. An extensive overlap between the ExpA and RsmA regulons was observed, suggesting that a substantial portion of ExpA regulation appears to be mediated through RsmA. However, a number of genes involved in the electron transport chain and oligogalacturonide metabolism, among other processes, were identified as regulated by ExpA independently of RsmA. These results suggest that ExpA may only partially impact fitness and virulence via RsmA.
    Applied and Environmental Microbiology 01/2014; DOI:10.1128/AEM.03829-13 · 3.95 Impact Factor