Genome-Led Discovery of the Antibacterial Cyclic Lipopeptide Kutzneridine A and Its Silent Biosynthetic Gene Cluster from Species.

Genome analysis of sp. CA-103260 revealed a putative lipopeptide-encoding biosynthetic gene cluster (BGC) that was cloned into a bacterial artificial chromosome (BAC) and heterologously expressed in M1152. As a result, a novel cyclic lipo-tetrapeptide containing two diaminopropionic acid residues and an exotic ,-acetonide ring, kutzneridine A (), was isolated and structurally characterized.

Maramycin, a Cytotoxic Isoquinolinequinone Terpenoid Produced through Heterologous Expression of a Bifunctional Indole Prenyltransferase/Tryptophan Indole-Lyase in .

Isoquinolinequinones represent an important family of natural alkaloids with profound biological activities. Heterologous expression of a rare bifunctional indole prenyltransferase/tryptophan indole-lyase enzyme from P8-A2 in J1074 led to the activation of a putative isoquinolinequinone biosynthetic gene cluster and production of a novel isoquinolinequinone alkaloid, named maramycin (). The structure of maramycin was determined by analysis of spectroscopic (1D/2D NMR) and MS spectrometric data.

Biosynthesis of the Azoxy Compound Azodyrecin from P8-A2.

Azoxy compounds are a distinctive group of bioactive secondary metabolites characterized by a unique RN═N(O)R moiety. The azoxy moiety is present in various classes of metabolites that exhibit various biological activities. The enzymatic mechanisms underlying azoxy bond formation remain enigmatic.

Systems Analysis of Highly Multiplexed CRISPR-Base Editing in Streptomycetes.

CRISPR tools, especially Cas9n-sgRNA guided cytidine deaminase base editors such as CRISPR-BEST, have dramatically simplified genetic manipulation of streptomycetes. One major advantage of CRISPR base editing technology is the possibility to multiplex experiments in genomically instable species. Here, we demonstrate scaled up Csy4 based multiplexed genome editing using CRISPR-mcBEST in .

Properties of Multidrug-Resistant Mutants Derived from Heterologous Expression Chassis Strain J1074.

J1074 is a popular platform to discover novel natural products via the expression of heterologous biosynthetic gene clusters (BGCs). There is keen interest in improving the ability of this platform to overexpress BGCs and, consequently, enable the purification of specialized metabolites. Mutations within gene for the β-subunit of RNA polymerase are known to increase rifampicin resistance and augment the metabolic capabilities of streptomycetes.

Identification and heterologous expression of the globomycin biosynthetic gene cluster.

Globomycin is a cyclic lipodepsipeptide originally isolated from several species which displays strong and selective antibacterial activity against Gram-negative pathogens. Its mode of action is based on the competitive inhibition of the lipoprotein signal peptidase II (LspA), which is absent in eukaryotes and considered an attractive target for the development of new antibiotics. Despite its interesting biological properties, the gene cluster encoding its biosynthesis has not yet been identified.

Identification of the Biosynthetic Gene Cluster for Pyracrimycin A, an Antibiotic Produced by sp.

Actinomycetes make a wealth of complex, structurally diverse natural products, and a key challenge is to link them to their biosynthetic gene clusters and delineate the reactions catalyzed by each of the enzymes. Here, we report the biosynthetic gene cluster for pyracrimycin A, a set of nine genes that includes a core nonribosomal peptide synthase () that utilizes serine and proline as precursors and a monooxygenase () that catalyzes Baeyer-Villiger oxidation. The cluster is similar to the one for brabantamide A; however, pyracrimycin A biosynthesis differs in that feeding experiments with isotope-labeled serine and proline suggest that a ring opening reaction takes place and a carbon is lost from serine downstream of the oxidation reaction.

Activation and Identification of a Griseusin Cluster in sp. CA-256286 by Employing Transcriptional Regulators and Multi-Omics Methods.

are well-known producers of a range of different secondary metabolites, including antibiotics and other bioactive compounds. Recently, it has been demonstrated that "silent" biosynthetic gene clusters (BGCs) can be activated by heterologously expressing transcriptional regulators from other BGCs. Here, we have activated a silent BGC in sp.

Complete Genome Sequence of sp. CA-230715, Encoding a 35-Module Type I Polyketide Synthase.

We report the sequencing, assembly, and annotation of the genome of sp. CA-230715, a potentially interesting producer of natural products. The genome of CA-230715 was sequenced using PacBio, Illumina, and Nanopore technologies.

Characterization and engineering of Streptomyces griseofuscus DSM 40191 as a potential host for heterologous expression of biosynthetic gene clusters.

Streptomyces griseofuscus DSM 40191 is a fast growing Streptomyces strain that remains largely underexplored as a heterologous host. Here, we report the genome mining of S. griseofuscus, followed by the detailed exploration of its phenotype, including the production of native secondary metabolites and ability to utilise carbon, nitrogen, sulphur and phosphorus sources.

Complete Genome Sequence of the Rare Actinobacterium sp. Strain CA-103260.

Here, we report the sequencing, assembly, and annotation of the genome of the rare actinobacterium sp. strain CA-103260. The genome of CA-103260 was sequenced using PacBio and Illumina technologies and it consists of a circular 11,609,901-bp chromosome.

Discovery and Characterization of Epemicins A and B, New 30-Membered Macrolides from sp. CA-103260.

Actinobacteria have been a rich source of novel, structurally complex natural products for many decades. Although the largest genus is , from which the majority of antibiotics in current and past clinical use were originally isolated, other less common genera also have the potential to produce a wealth of novel secondary metabolites. One example is the genus, which currently contains only five reported species.

Complete Genome Sequence of sp. Strain CA-256286.

Here, we report the sequencing, assembly, and annotation of the genome of sp. strain CA-256286. The genome consists of a linear 7,726,360-nucleotide chromosome and a linear 466,817-nucleotide putative plasmid.

Distribution of ε-Poly-l-Lysine Synthetases in Coryneform Bacteria Isolated from Cheese and Human Skin.

ε-Poly-l-lysine is a potent antimicrobial produced through fermentation of and used in many Asian countries as a food preservative. It is synthesized and excreted by a special nonribosomal peptide synthetase (NRPS)-like enzyme called Pls. In this study, we discovered a gene from cheese bacterium that showed high similarity to the Pls from in terms of domain architecture and gene context.

High-Quality Sequencing, Assembly, and Annotation of the Streptomyces griseofuscus DSM 40191 Genome.

Here, we report the sequencing, assembly, and annotation of the genome of DSM 40191. The genome of was sequenced using PacBio and Illumina technologies. It consists of a linear 8,721,740-bp chromosome and three plasmids, pSGRIFU1 (220 kb), pSGRIFU2 (88 kb), and pSGRIFU3 (86 kb).

Highly efficient DSB-free base editing for streptomycetes with CRISPR-BEST.

Streptomycetes serve as major producers of various pharmacologically and industrially important natural products. Although CRISPR-Cas9 systems have been developed for more robust genetic manipulations, concerns of genome instability caused by the DNA double-strand breaks (DSBs) and the toxicity of Cas9 remain. To overcome these limitations, here we report development of the DSB-free, single-nucleotide-resolution genome editing system CRISPR-BEST (CRISPR-Base Editing SysTem), which comprises a cytidine (CRISPR-cBEST) and an adenosine (CRISPR-aBEST) deaminase-based base editor.

Heterologous AdpA transcription factors enhance landomycin production in Streptomyces cyanogenus S136 under a broad range of growth conditions.

Streptomyces cyanogenus S136 is the only known producer of landomycin A (LaA), one of the largest glycosylated angucycline antibiotics possessing strong antiproliferative properties. There is rising interest in elucidation of mechanisms of action of landomycins, which, in turn, requires access to large quantities of the pure compounds. Overproduction of LaA has been achieved in the past through manipulation of cluster-situated regulatory genes.

Analysis of Streptomyces coelicolor M145 genes SCO4164 and SCO5854 encoding putative rhodaneses.

Streptomyces coelicolor genome carries two apparently paralogous genes, SCO4164 and SCO5854, that encode putative thiosulfate sulfurtransferases (rhodaneses). These genes (and their presumed translation products) are highly conserved and widely distributed across actinobacterial genomes. The SCO4164 knockout strain was unable to grow on minimal media with either sulfate or sulfite as the sole sulfur source.

The MalR type regulator AcrC is a transcriptional repressor of acarbose biosynthetic genes in Actinoplanes sp. SE50/110.

Background: Acarbose is used in the treatment of diabetes mellitus type II and is produced by Actinoplanes sp. SE50/110. Although the biosynthesis of acarbose has been intensively studied, profound knowledge about transcription factors involved in acarbose biosynthesis and their binding sites has been missing until now.

Targeted genome editing in the rare actinomycete Actinoplanes sp. SE50/110 by using the CRISPR/Cas9 System.

The application of genome editing technologies, like CRISPR/Cas9 for industrially relevant microorganisms, is becoming increasingly important. Compared to other methods of genetic engineering the decisive factor is that CRISPR/Cas9 is relatively easy to apply and thus time and effort can be significantly reduced in organisms, which are otherwise genetically difficult to access. Because of its many advantages and opportunities, we adopted the CRISPR/Cas9 technology for Actinoplanes sp.

Influence of transition metals on Streptomyces coelicolor and S. sioyaensis and generation of chromate-reducing mutants.

Bacteria-assisted bioremediation is widely recognized as a low-cost method to minimize the consequences of soil pollution with toxic metals originating from industrial sites. Strains used in bioremediation have to deal with high metal load via biosorption, reduction, bioprecipitation, metal sequestration, and/or chelation. Actinobacteria, and streptomycetes in particular, are considered a perspective group for bioremediation as natural soil inhabitants with extensive secondary metabolism.

Cultivable actinomycetes from rhizosphere of birch (Betula pendula) growing on a coal mine dump in Silets, Ukraine.

Five actinomycete strains were isolated from the rhizosphere of birch, one of a few native tree forms capable of thriving on the upper level of a coal mine dump near the village of Silets (Lvivska region, Ukraine). No such strains were isolated from surrounding gangue, or from nearby grass Calamagrostis epigeios. Using 16S rDNA sequencing and analysis of cell wall aminoacids, four of these strains were shown to belong to genus Streptomyces and one to be Amycolatopsis.

[Design of Streptomyces nogalater LV65 strains with higher synthesis of nogalamicin using regulatory genes].

Influence of cloned regulatory genes on biosynthesis of nogalamicin by Streptomyces nogalater LV65 strains has been studied. Gene snorA from the S. nogalater genome was cloned in multicopy replicative plasmid pSOKA and integrative plasmid pR3A.