Abstract
We announce the sequencing of Streptomyces chartreusis NRRL 12338 and NRRL 3882 and Streptomyces lysosuperificus ATCC 31396. These are producers of tunicamycins, chartreusins, cephalosporins, holomycins, and calcimycin. The announced genomes, together with the published Streptomyces clavuligerus genome, will facilitate data mining of these secondary metabolites.
GENOME ANNOUNCEMENT
The tunicamycins are structurally unique compounds that inhibit N-acetylhexosamine-1-phosphate-prenol phosphate transferases, blocking formation of N-acetylmuramylpentapeptide-undecanol pyrophosphate in bacteria or N-acetylglucosamine-dolichol pyrophosphate in eukaryotes. These are essential intermediates in the biosynthesis of eukaryotic N-glycoproteins and of bacterial peptidoglycan. Tunicamycins were discovered by Tamura from Streptomyces lysosuperificus ATCC 31396 (10). They are nucleotide-type antibiotics that contain two unusual features: an 11-carbon dialdose sugar called tunicamine and an anomeric-to-anomeric α,β-1,11-glycosidic linkage between the tunicamine and N-acetylglucosamine. Tunicamycins are also produced by two strains of Streptomyces chartreusis (NRRL 3882 and NRRL 12338), Streptomyces clavuligerus NRRL 3585, Clavibacter toxicus NCPPB 3552, and Clavibacter michiganensis subsp. michiganensis (11). Similar compounds are from Streptomyces bobiliae (mycospocidins), Streptomyces griseoflavus subsp. thuringiensis (streptovirudins), and antibiotic 24010 (4). Twelve tunicamycin biosynthetic genes (tunA to tunL) have been identified in S. chartreusis NRRL 3882 and S. clavuligerus NRRL 3585 and are also present on the sequenced genome of Actinosynnema mirum DSM 44827, although with a truncated tunL (3, 13). S. clavuligerus NRRL 3585 also produces clavulinic acid (a lactamase inhibitor), cephalosporin-type antibiotics, and holomycin, and S. chartreusis strains also produce chartreusin and calcimycin (calcium ionophore A23187). Gene clusters that code for biosynthesis of these natural products have also been identified (9, 6, 12, 14).
Genomic sequences were generated using paired-end sequencing with a 75-bp run on an Illumina Genome Analyzer II× at the Wisconsin Biotechnology Center. Libraries prepared using the Nextera DNA sample prep kit were tagged and pooled. The NRRL 12338 library was covered by 2,417,521 paired reads, NRRL 3882 by 618,150 paired reads, and ATCC 31396 by 571,352 paired reads. Assembly was performed using a custom pipeline that combined Velvet version 1.0.15 (15), SOAPdenovo version 1.04 (7), and EULER-SR version 1.1.2 (2) assemblies in gsAssembler version 2.5.3 (8) using 400,000 paired-end reads reformatted for gsAssembler as additional input, followed by final scaffolding with SSPACE version 1.1 (1). This resulted in a total of 8,859,518 bp, 9,028,970 bp, and 8,505,115 bp assembled into 112, 3,703, and 2,823 scaffolds, 351, 5,817, and 5,885 contigs, and N50 scaffold values of 228,498 bp, 5,008 bp, and 7,704 bp for NRRL 12338, NRRL 3882, and ATCC 31396, respectively. Prodigal (5) was used for open reading frame (ORF) prediction, resulting in 8,016 (NRRL 12338), 11,795 (NRRL 3882), and 10,616 (ATCC 31396) ORFs. G+C contents for each strain were 71.3%, NRRL 12338; 70.8%, NRRL 3882; and 72.0%, ATCC 31396.
Here, we announce the genomic sequencing of three tunicamycin-producing streptomyces, S. chartreusis NRRL 12338 and NRRL 3882 and Tamura's original S. lysosuperificus isolate. Taken together with the previously completed genomes of S. clavuligerus NRRL 3585 and Actinosynnema mirum DSM 44827, these will aid our understanding of tunicamycin genetics and biosynthesis and of several other commercially important natural products.
Nucleotide sequence accession numbers.
Nucleotide sequences are available in GenBank: project number 72669, Streptomyces lysosuperificus ATCC 31396, accession number AGDC00000000; project number 72671, Streptomyces chartreusis NRRL 3882, accession number AGDD00000000; and project number 72673, Streptomyces chartreusis NRRL 12338, accession number AGDE00000000.
Acknowledgments
This work was supported by the National Institutes of Health (GM PO1 GM077596). J.R.D. was supported by a postdoctoral fellowship from the Institute for Genomic Biology at the University of Illinois at Urbana-Champaign.
The contents of this work are solely the responsibility of the authors and do not necessarily represent the official views of the NIGMS or NIH. The mention of firm names or trade products does not imply that they are endorsed or recommended by the U.S. Department of Agriculture over other firms or similar products not mentioned.
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