Involvement of Enterobactin Synthesis Pathway in Production of Microcin H47

Abstract

Microcin H47 (MccH47) is a gene-encoded peptide antibiotic produced by an Escherichia coli clinical isolate which is active on strains of gram-negative bacteria. Its uptake by E. coli K-12-susceptible cells depends on the presence of any of the outer membrane proteins Cir, Fiu, and FepA, the three catechol receptors of this organism. The nucleotide sequence of a portion of the MccH47 genetic system that had not yet been studied was elucidated. Five open reading frames were identified, three of which corresponded to genes encoding functions related to catechol-type siderophores: mchA and mchS1 are iroB and iroD homologues, respectively, and mchS4 was found to promote the production of the catecholate siderophore enterobactin. The possible relationship between enterobactin synthesis and MccH47 production was studied. Enterobactin-deficient strains failed to produce MccH47 when transformed with the antibiotic genetic determinants and upon introduction of the ent genetic cluster, the production of both the siderophore and MccH47 was restored. Further studies demonstrated that at least the enterobactin nonribosomal peptide synthase EntF is necessary for MccH47 synthesis. The relationship found between MccH47 and catecholate siderophore production is discussed, and a model outlining MccH47 synthesis is proposed.

Siderophore-Based Iron Acquisition and Pathogen Control

“…After hydrolysis from the siderophore carrier, the serylthioribosyl pyrimidine moiety blocks protein biosynthesis by inhibiting seryl-tRNA synthetases (307). Danomycins and salmycins produced by actinomycetes contain the trishydroxamate danoxamine as a siderophore unit that is connected with an aminoglycoside moiety (147,326) (13,316). In addition to enterobactin synthesis, the modification requires genes of the iroA cluster for salmochelin synthesis, essentially iroB and iroD for glucosylation and backbone linearization, respectively.…”

Comparative Analysis of Chromosome-Encoded Microcins

“…Thus, the antibiotic is taken up by E. coli K-12 cells through the catechol pathway, involving the receptor Cir, Fiu, or FepA, and the TonB complex (24). This mode of synthesis, which endows the peptide antibiotic with the ability to enter gram-negative cells through the catechol uptake pathway, has been called the "catechol strategy" (2).…”

“…Salmochelins are mono-or diglucosylated derivatives of the catecholate siderophore enterobactin (Ent). Thus, the cell must provide enterobactin, which is converted into salmochelins, and these are employed for MccH47 synthesis (2,7). After being synthesized, MccH47 is secreted through a type I apparatus that is formed by the products of the mchE and mchF genes and by the outer membrane protein TolC.…”

“…MccH47 is then composed of two moieties, peptide and salmochelin. The peptide possesses the deleterious activity, and the salmochelin confers the uptake properties to the molecule (2,20). Thus, the antibiotic is taken up by E. coli K-12 cells through the catechol pathway, involving the receptor Cir, Fiu, or FepA, and the TonB complex (24).…”

Use of mchI Encoding Immunity to the Antimicrobial Peptide Microcin H47 as a Plasmid Selection Marker in Attenuated Bacterial Live Vectors

“…The mch47 peptide is synthesized as a 75-residue precursor that is processed during secretion to a 60-residue mature extracellular peptide of 4.9 kDa (36). The mch47 operon also encodes an ATP binding cassette (ABC) export system specific for H47 (2), a catecholate siderophore production system that is proposed to enhance MccH47 uptake by target bacteria (1,33), and an immunity protein that is required for bacterial self immunity (10,37). MccH47 targets the proton channel of the F 0 portion of ATP synthase, which abolishes the regulated entry of protons, leading to the depolarization of target cell membranes (38,48).…”