Fosfonochlorin

A set of α-quaternary 3-chloro-1-hydroxyalkylphosphonates, analogues of fosfomycin and fosfonochlorin, some of which are new compounds, was synthesized. The compounds were screened for bioactivity against several clinical and standard microbial isolates. Some were found to have moderate activity. The activity was higher with phenyl protection of the phosphoryl ester groups and α-phenyl substitution. Compound 11 was as effective or more potent than fosfomycin or chloramphenicol against several Gram-negative bacteria as well as against some Gram-positive ones.

A new antibiotic, fosfonochlorin, was found in the culture filtrate of four strains of fungi freshly isolated from soil samples. These strains were identified as Fusarium avenaceum, Fusarium oxysporum, Fusarium tricinctum and Talaromyces flavus. Fosfonochlorin was a low molecular weight antibiotic (MW 158), soluble in water and methanol, but insoluble in acetone, ethyl acetate and chloroform. It was named after its possession of phosphorus and chlorine atoms, each one molar in its structure. The structure was determined as chloroacetylphosphonic acid mainly by the 1H NMR and mass spectrometric analyses. It was moderately active against some species of Gram-negative bacteria and its synergistic effect with glucose-6-phosphate was observed on Staphylococcus aureus and Escherichia coli. Spheroplast formation of the susceptible organisms with this antibiotic suggested that it might inhibit their cell wall synthesis.

The fungal metabolite Fosfonochlorin features a chloroacetyl moiety that is unusual within known phosphonate natural product biochemistry. Putative biosynthetic genes encoding Fosfonochlorin in Fusarium and Talaromyces spp. were investigated through reactions of encoded enzymes with synthetic substrates and isotope labelling studies. We show that the early biosynthetic steps for Fosfonochlorin involve the reduction of phosphonoacetaldehyde to form 2-hydroxyethylphosphonic acid, followed by oxidative intramolecular cyclization of the resulting alcohol to form (S)-epoxyethylphosphonic acid. The latter reaction is catalyzed by FfnD, a rare example of a non-heme iron/2-(oxo)glutarate dependent oxacyclase. In contrast, FfnD behaves as a more typical oxygenase with ethylphosphonic acid, producing (S)-1-hydroxyethylphosphonic acid. FfnD thus represents a new example of a ferryl generating enzyme that can suppress the typical oxygen rebound reaction that follows abstraction of a substrate hydrogen by a ferryl oxygen, thereby directing the substrate radical towards a fate other than hydroxylation.