Blocks in the pseudouridimycin pathway unlock hidden metabolites in the Streptomyces producer strain

Abstract

We report a metabolomic analysis of Streptomyces sp. ID38640, a soil isolate that produces the bacterial RNA polymerase inhibitor pseudouridimycin. The analysis was performed on the wild type, on three newly constructed and seven previously reported mutant strains disabled in different genes required for pseudouridimycin biosynthesis. The results indicate that Streptomyces sp. ID38640 is able to produce, in addition to lydicamycins and deferroxiamines, as previously reported, also the lassopeptide ulleungdin, the non-ribosomal peptide antipain and the osmoprotectant ectoine. The corresponding biosynthetic gene clusters were readily identified in the strain genome. We also detected the known compound pyridindolol, for which we propose a previously unreported biosynthetic gene cluster, as well as three families of unknown metabolites. Remarkably, the levels of most metabolites varied strongly in the different mutant strains, an observation that enabled detection of metabolites unnoticed in the wild type. Systematic investigation of the accumulated metabolites in the ten different pum mutants identified shed further light on pseudouridimycin biosynthesis. We also show that several Streptomyces strains, able to produce pseudouridimycin, have distinct genetic relationship and metabolic profile with ID38640.

Reference-Grade Genome and Large Linear Plasmid of Streptomyces rimosus: Pushing the Limits of Nanopore Sequencing

“…From previous studies, S. rimosus ATCC 10970 was known to produce oxytetracycline, rimocidin ( 1 ), rimosamide ( 26 ), and tyrobetaine ( 40 ), and we confirmed their biosynthesis in our experiment. Interestingly, pseudouridimycin was also detected in our extracts, although 4 pum genes from the homologous pum BGC ( 28 ) were absent. We also detected the rimocidin analogs CE108 and amide CE108, previously characterized only in strain Streptomyces diastaticus var.…”

“…PumB is a putative exporter, and PumC is a predicted DeoR transcriptional regulator. PumD is a putative hydrolase of the HAD family with the predicted function in the initial steps of biosynthesis ( 27 , 28 ). Despite the absence of pumA , pumB , pumC , and pumD homolog genes in BGC no.…”

Biosynthesis of C-nucleoside antibiotics in actinobacteria: recent advances and future developments

“…The knock-out mutant strains were correspondingly able to accumulate the specialized intermediates, including pseudouridine (PU), 5′-amino pseudouridine (APU), Gln-APU, N -hydroxy-Gln-APU guanidinoacetic acid (GAA) [ 9 ]. Recently, the minimal PUM BGC was generated and expressed in the heterologous host Streptomyces coelicolor M1146 under control of strong promotors including pumB and pumD–N [ 71 , 72 ]. Besides that, all core genes involved in the formation PUM could be identified in Streptomyces albus DSM 40763 ( S. albus ), a previously reported strain capable of producing the C -nucleoside strepturidin, which leads to the structural revision of the antibiotic as PUM.…”

“…Based on in silico analysis, genetic, and feeding experiments of precursors or intermediates, the complete PUM biosynthetic pathway was tentatively proposed as indicated in Fig. 7 b [ 71 ].

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“…After that, the intermediate 31 is N -hydroxylated by the oxidoreductase PumE to generate 32 . In a parallel way, another intermediate GAA is produced from glycine and arginine by the amidinotransferase PumN, and the GAA molecule is added to compound 32 by PumM leading to the final compound, PUM, which is transported out of the cell by PumL (transporter) [ 71 ]. More recently, PumF has been identified as a positive regulator by controlling the transcription of the pumK–N unit [ 71 ].…”