Nonomuraea sp. ATCC 55076 harbours the largest actinomycete chromosome to date and the kistamicin biosynthetic gene cluster.
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ABSTRACT:
SUBMITTER: Nazari B
PROVIDER: S-EPMC5463735 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Glycopeptides and ?-lactams inhibit bacterial peptidoglycan synthesis in Gram-positive bacteria; resistance to these antibiotics is studied intensively in enterococci and staphylococci because of their relevance to infectious disease. Much less is known about antibiotic resistance in glycopeptide-producing actinomycetes that are likely to represent the evolutionary source of resistance determinants found in bacterial pathogens. Nonomuraea sp. ATCC 39727, the producer of A40926 (the precursor for the semisynthetic dalbavancin), does not harbor the canonical vanHAX genes. Consequently, we investigated the role of the ?-lactam-sensitive D,D-peptidase/D,D-carboxypeptidase encoded by vanYn, the only van-like gene found in the A40926 biosynthetic gene cluster, in conferring immunity to the antibiotic in Nonomuraea sp. ATCC 39727. Taking advantage of the tools developed recently to genetically manipulate this uncommon actinomycete, we varied vanYn gene dosage and expressed vanHatAatXat from the teicoplanin producer Actinoplanes teichomyceticus in Nonomuraea sp. ATCC 39727. Knocking out vanYn, complementing a vanYn mutant, or duplicating vanYn had no effect on growth but influenced antibiotic resistance and, in the cases of complementation and duplication, antibiotic production. Nonomuraea sp. ATCC 39727 was found to be resistant to penicillins, but its glycopeptide resistance was diminished in the presence of penicillin G, which inhibits VanYn activity. The heterologous expression of vanHatAatXat increased A40926 resistance in Nonomuraea sp. ATCC 39727 but did not increase antibiotic production, indicating that the level of antibiotic production is not directly determined by the level of resistance. The vanYn-based self-resistance in Nonomuraea sp. ATCC 39727 resembles the glycopeptide resistance mechanism described recently in mutants of Enterococcus faecium selected in vitro for high-level resistance to glycopeptides and penicillins.
| S-EPMC4135873 | biostudies-literature
Project description:A straight-chain, spore-forming actinobacterium, strain YIM 120770(T), was isolated from soil. Phylogenetic analysis on the basis of 16S rRNA gene sequence comparisons revealed that the isolate represents a distinct cluster within the clade comprising the genus Nonomuraea and is related most closely to Nonomuraea rhizophila YIM 67092(T) (96.5% similarity). Cells of strain YIM 120770(T) grew in the presence of 0-3% (w/v) NaCl, at 15-37 °C and at pH 7.0-8.0. The diagnostic amino acid was meso-diaminopimelic acid, cell hydrolysates contained madurose, glucose, mannose, ribose and galactose, the predominant cellular fatty acids were 10-methyl C(17:0) and iso-C(16:0), and the DNA G+C content was 66.4 mol%, data consistent with affiliation of strain YIM 120770(T) to the genus Nonomuraea. Strain YIM 120770(T) shared low levels of 16S rRNA gene sequence similarity (<97%) with the type strains of recognized species of the genus Nonomuraea and could be differentiated from its closest phylogenetic relative based on phenotypic characteristics. These results suggested that strain YIM 120770(T) represents a novel species of the genus Nonomuraea, for which the name Nonomuraea soli sp. nov. is proposed. The type strain is YIM 120770(T) (=DSM 45533(T)=JCM 17347(T)).
| S-EPMC3541754 | biostudies-literature
Project description:A novel actinomycete, designated strain NEAU-Z6(T), was isolated from eggplant (Solanum melongena L.) root. Phylogenetic analysis based on the 16S rRNA gene sequence showed that strain NEAU-Z6(T) belonged to the genus Nonomuraea, with highest sequence similarity to Nonomuraea monospora PT 708(T) (98.83 %), Nonomuraea rosea GW 12687(T) (98.55 %) and Nonomuraea rhizophila YIM 67092(T) (98.02 %). Sequence similarities between strain NEAU-Z6(T) and other species of the genus Nonomuraea ranged from 97.94 % (Nonomuraea candida HMC10(T)) to 96.30 % (Nonomuraea wenchangensis 210417(T)). Key morphological, physiological and chemotaxonomic characteristics of strain NEAU-Z6(T) were congruent with the description of the genus Nonomuraea. The G+C content of the genomic DNA was 64.51 mol%. DNA-DNA relatedness and comparative analysis of physiological, biochemical and chemotaxonomic data allowed genotypic and phenotypic differentiation of strain NEAU-Z6(T) from closely related species. Thus, strain NEAU-Z6(T) represents a novel species of the genus Nonomuraea, for which the name Nonomuraea solani sp. nov. is proposed. The type strain is NEAU-Z6(T) ( = CGMCC 4.7037(T) = DSM 45729(T)).
| S-EPMC3749519 | biostudies-literature
Project description:In glycopeptide-resistant enterococci and staphylococci, high-level resistance is achieved by replacing the C-terminal d-alanyl-d-alanine of lipid II with d-alanyl-d-lactate, thus reducing glycopeptide affinity for cell wall targets. Reorganization of the cell wall in these organisms is directed by the vanHAX gene cluster. Similar self-resistance mechanisms have been reported for glycopeptide-producing actinomycetes. We investigated glycopeptide resistance in Nonomuraea sp. ATCC 39727, the producer of the glycopeptide A40926, which is the precursor of the semisynthetic antibiotic dalbavancin, which is currently in phase III clinical trials. The MIC of Nonomuraea sp. ATCC 39727 toward A40926 during vegetative growth was 4 microg/ml, but this increased to ca. 20 microg/ml during A40926 production. vanHAX gene clusters were not detected in Nonomuraea sp. ATCC 39727 by Southern hybridization or by PCR with degenerate primers. However, the dbv gene cluster for A40926 production contains a gene, vanY (ORF7), potentially encoding an enzyme capable of removing the terminal d-Ala residue of pentapeptide peptidoglycan precursors. Analysis of UDP-linked peptidoglycan precursors in Nonomuraea sp. ATCC 39727 revealed the predominant presence of the tetrapeptide UDP-MurNAc-l-Ala-d-Glu-meso-Dap-d-Ala and only traces of the pentapeptide UDP-MurNAc-l-Ala-d-Glu-meso-Dap-d-Ala-d-Ala. This suggested a novel mechanism of glycopeptide resistance in Nonomuraea sp. ATCC 39727 that was based on the d,d-carboxypeptidase activity of vanY. Consistent with this, a vanY-null mutant of Nonomuraea sp. ATCC 39727 demonstrated a reduced level of glycopeptide resistance, without affecting A40926 productivity. Heterologous expression of vanY in a sensitive Streptomyces species, Streptomyces venezuelae, resulted in higher levels of glycopeptide resistance.
| S-EPMC2876364 | biostudies-literature
Project description:UnlabelledThe actinomycete Nonomuraea sp. strain ATCC 39727 produces the glycopeptide A40926, the precursor of dalbavancin. Biosynthesis of A40926 is encoded by the dbv gene cluster, which contains 37 protein-coding sequences that participate in antibiotic biosynthesis, regulation, immunity, and export. In addition to the positive regulatory protein Dbv4, the A40926-biosynthetic gene cluster encodes two additional putative regulators, Dbv3 and Dbv6. Independent mutations in these genes, combined with bioassays and liquid chromatography-mass spectrometry (LC-MS) analyses, demonstrated that Dbv3 and Dbv4 are both required for antibiotic production, while inactivation of dbv6 had no effect. In addition, overexpression of dbv3 led to higher levels of A40926 production. Transcriptional and quantitative reverse transcription (RT)-PCR analyses showed that Dbv4 is essential for the transcription of two operons, dbv14-dbv8 and dbv30-dbv35, while Dbv3 positively controls the expression of four monocistronic transcription units (dbv4, dbv29, dbv36, and dbv37) and of six operons (dbv2-dbv1, dbv14-dbv8, dbv17-dbv15, dbv21-dbv20, dbv24-dbv28, and dbv30-dbv35). We propose a complex and coordinated model of regulation in which Dbv3 directly or indirectly activates transcription of dbv4 and controls biosynthesis of 4-hydroxyphenylglycine and the heptapeptide backbone, A40926 export, and some tailoring reactions (mannosylation and hexose oxidation), while Dbv4 directly regulates biosynthesis of 3,5-dihydroxyphenylglycine and other tailoring reactions, including the four cross-links, halogenation, glycosylation, and acylation.ImportanceThis report expands knowledge of the regulatory mechanisms used to control the biosynthesis of the glycopeptide antibiotic A40926 in the actinomycete Nonomuraea sp. strain ATCC 39727. A40926 is the precursor of dalbavancin, approved for treatment of skin infections by Gram-positive bacteria. Therefore, understanding the regulation of its biosynthesis is also of industrial importance. So far, the regulatory mechanisms used to control two other similar glycopeptides (balhimycin and teicoplanin) have been elucidated, and beyond a common step, different clusters seem to have devised different strategies to control glycopeptide production. Thus, our work provides one more example of the pitfalls of deducing regulatory roles from bioinformatic analyses only, even when analyzing gene clusters directing the synthesis of structurally related compounds.
| S-EPMC4518825 | biostudies-literature
Project description:Rare actinomycete genera are accepted as a promising source of novel metabolites having pharmaceutical importance. One such genus of rare actinomycete is Nonomuraea. The present study was aimed at characterizing the antibiotic producing Nonomuraea strain JAJ18 which was previously isolated from coastal solar saltern. Strain JAJ18 was recognized as a member of genus Nonomuraea based on its almost complete 16S rRNA gene sequence and phenotypic characteristics. The strain JAJ18 was found to be closely related to Nonomuraea maheshkhaliensis 16-5-14(T) (98.90%), Nonomuraea candida HMC10(T) (98.58%), and Nonomuraea jabiensis A4036(T) (98.43%). From cell-free culture broth of strain JAJ18, an antibiotic was extracted and purified by silica column chromatography. The obtained antibiotic was found to be active against a range of Gram-positive and Gram-negative bacteria including drug-resistant Staphylococcus, with minimal inhibitory concentration (MIC) ranging from 0.5 to 16.0 µg mL(-1). The structural characteristics of antibiotic were determined by FTIR and NMR spectroscopy. The antibiotic was identified to be an aliphatic rich compound with significant dissimilarity to known antibiotics reported from members of the genus, Nonomuraea. As the trends to discover novel metabolites from Nonomuraea are vibrant, further studies are needed to understand the structural and biotechnological significance of antibiotic compound produced by Nonomuraea sp. JAJ18.
| S-EPMC4281464 | biostudies-literature
Project description:There is accumulating evidence that the ability of actinomycetes to produce antibiotics and other bioactive secondary metabolites has been underestimated due to the presence of cryptic gene clusters. The activation of dormant genes is therefore one of the most important areas of experimental research for the discovery of drugs in these organisms. The recent observation that several actinomycetes possess two RNA polymerase beta-chain genes (rpoB) has opened up the possibility, explored in this study, of developing a new strategy to activate dormant gene expression in bacteria. Two rpoB paralogs, rpoB(S) and rpoB(R), provide Nonomuraea sp. strain ATCC 39727 with two functionally distinct and developmentally regulated RNA polymerases. The product of rpoB(R), the expression of which increases after transition to stationary phase, is characterized by five amino acid substitutions located within or close to the so-called rifampin resistance clusters that play a key role in fundamental activities of RNA polymerase. Here, we report that rpoB(R) markedly activated antibiotic biosynthesis in the wild-type Streptomyces lividans strain 1326 and also in strain KO-421, a relaxed (rel) mutant unable to produce ppGpp. Site-directed mutagenesis demonstrated that the rpoB(R)-specific missense H426N mutation was essential for the activation of secondary metabolism. Our observations also indicated that mutant-type or duplicated, rpoB often exists in nature among rare actinomycetes and will thus provide a basis for further basic and applied research.
| S-EPMC2632078 | biostudies-literature
Project description:Aspergillus terreus is a filamentous ascomycota, which is prominent for its production of lovastatin, an antihypercholesterolemic drug. The commercial importance of lovastatin with annual sales of billions of dollars made us to focus on lovastatin biosynthetic cluster proteins. The analysis of these lovastatin biosynthetic cluster proteins with different perspectives such as physicochemical property, structure based analysis and functional studies were done to find out the role and function of every protein involved in the lovastatin biosynthesis pathway. Several computational tools are used to predict the physicochemical properties, secondary structural features, topology, patterns, domains and cellular location. There are 8 unidentified proteins in lovastatin biosynthetic cluster, in which 6 proteins have homologous partners, and annotation transfer is done based on the closely related homologous genes, and their structures are also modeled. The two other proteins that do not have homologous partners are predicted as PQ loop repeat protein that may be involved in glycosylation machinery and as thiolase-acyl activity by the integrated functional analysis approach.
| S-EPMC3124688 | biostudies-literature
Project description:The 53-kb biosynthetic gene cluster for the novel anticholesterol natural product herboxidiene was identified in Streptomyces chromofuscus ATCC 49982 by genome sequencing and gene inactivation. In addition to herboxidiene, a biosynthetic intermediate, 18-deoxy-herboxidiene, was also isolated from the fermentation broth of S. chromofuscus ATCC 49982 as a minor metabolite.
| S-EPMC3298124 | biostudies-literature
Project description:Actinomycete bacteria isolated from freshwater environments are an unexplored source of natural products. Here we report the complete genome of the Great Lakes-derived Micromonospora sp. strain B006, revealing its potential for natural product biosynthesis. The 7-megabase pair chromosome of strain B006 was sequenced using Illumina and Oxford Nanopore technologies followed by Sanger sequencing to close remaining gaps. All identified biosynthetic gene clusters (BGCs) were manually curated. Five known BGCs were identified encoding desferrioxamine, alkyl- O-dihydrogeranylmethoxyhydroquinone, a spore pigment, sioxanthin, and diazepinomicin, which is currently in phase II clinical trials to treat Phelan-McDermid syndrome and co-morbid epilepsy. We report here that strain B006 is indeed a producer of diazepinomicin and at yields higher than previously reported. Moreover, 11 of the 16 identified BGCs are orphan, eight of which were transcriptionally active under the culture condition tested. Orphan BGCs include an enediyne polyketide synthase and an uncharacteristically large, 36-module polyketide synthase-nonribosomal peptide synthetase BGC. We developed a genetics system for Micromonospora sp. B006 that will contribute to deorphaning BGCs in the future. This study is one of the few attempts to report the biosynthetic capacity of a freshwater-derived actinomycete and highlights this resource as a potential reservoir for new natural products.
| S-EPMC6174880 | biostudies-literature