Heterogeneous rRNA molecules encoded by Streptomyces coelicolor M145 genome are all expressed and assembled into ribosomes

The Streptomyces coelicolor M145 genome harbors six copies of divergent rRNA operons that differ at ~0.2% and ~0.6% of the nucleotide positions in small subunit (SSU) and large subunit (LSU) rRNA genes, respectively. When these rRNA genes are expressed, a single cell may harbor three different kinds of SSU rRNA and five kinds of LSU rRNA. Primer extension analyses revealed that all of the heterogeneous rRNA molecules are expressed and assembled into ribosomes. This finding and the maintenance of the intragenomic variability of rRNA operons imply the existence of functional divergence of rRNA species in this developmentally complex microorganism.     

Discovery of a new peptide natural product by Streptomyces coelicolor genome mining

Analyses of microbial genome sequences reveal numerous examples of gene clusters encoding proteins typically involved in complex natural product biosynthesis but not associated with the production of known natural products. In Streptomyces coelicolor M145 there are several gene clusters encoding new nonribosomal peptide synthetase (NRPS) systems not associated with known metabolites. Application of structure-based models for substrate recognition by NRPS adenylation domains predicts the amino acids incorporated into the putative peptide products of these systems, but the accuracy of these predictions is untested. Here we report the isolation and structure determination of the new tris-hydroxamate tetrapeptide iron chelator coelichelin from S. coelicolor using a genome mining approach guided by substrate predictions for the trimodular NRPS CchH, and we show that this enzyme, which lacks a C-terminal thioesterase domain, together with a homolog of enterobactin esterase (CchJ), are required for coelichelin biosynthesis. These results demonstrate that accurate prediction of adenylation domain substrate selectivity is possible and raise intriguing mechanistic questions regarding the assembly of a tetrapeptide by a trimodular NRPS.     

Sequential deletion of all the polyketide synthase and nonribosomal peptide synthetase biosynthetic gene clusters and a 900-kb subtelomeric sequence of the linear chromosome of Streptomyces coelicolor

Streptomyces coelicolor, with its 8 667 507-bp linear chromosome, is the genetically most studied Streptomyces species and is an excellent model for studying antibiotic production and cell differentiation. Here, we report construction of S. coelicolor derivatives containing sequential deletions of all the 10 polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS) biosynthetic gene clusters and a 900-kb subtelomeric sequence (total c. 1.22 Mb, 14% of the genome). No obvious differences in growth rates and sporulation of the strains were found. An artificially circularized S. coelicolor genome with deletions of total c. 1.6 Mb segments (840-kb for the left and 761-kb for the right arm of the linear chromosome) was obtained. The actinorhodin biosynthetic gene cluster could be overexpressed in some of the constructed strains.     

Estimate of the Genome Size by Renaturation Studies in Streptomyces

The genome sizes of Streptomyces coelicolor and Streptomyces rimosus as calculated by deoxyribonucleic acid reassociation kinetics are approximately 10.5 X 10(6) nucelotide pairs.     

Impact of the first <Emphasis Type="Italic">Streptomyces</Emphasis> genome sequence on the discovery and production of bioactive substances

An important addition to the field of bacterial genomics is the recent publication of the complete genome sequence of Streptomyces coelicolor. This strain has been for some decades the model organism for streptomycetes and other filamentous actinomycetes, Gram-positive bacteria highly valuable for their ability to produce thousands of bioactive metabolites, many of which have found important applications in medicine and agriculture. We discuss here the impacts that the S. coelicolor genome sequence is likely to have on the production of bioactive metabolites by current industrial strains, on the possible development of future superhost(s) for the production of valuable drugs, and on the search for new bioactive substances from microbial sources.     

Molecular cloning, heterologous expression, and functional characterisation of a malate synthase gene from Streptomyces coelicolor A3(2)

With the rapid generation of genetic information from the Streptomyces coelicolor genome project, deciphering the relevant gene products is critical for understanding the genetics of this model streptomycete. A putative malate synthase gene (aceB) from S. coelicolor A3(2) was identified by homology-based analysis, cloned by polymerase chain reaction, and fully sequenced on both strands. The putative malate synthase from S. coelicolor has an amino acid identity of 77% with the malate synthase of S. clavuligerus, and possesses an open reading frame which codes for a protein of 540 amino acids. In order to establish the identity of this gene, the putative aceB clones were subcloned into the expression vector pET24a, and heterologously expressed in Escherichia coli BL21(DE3). Soluble cell-free extracts containing the recombinant putative malate synthase exhibited a specific activity of 1623 (nmol.mg-1.min-1), which is an increment of 92-fold compared to the non-recombinant control. Thus, the gene product was confirmed to be a malate synthase. Interestingly, the specific activity of S. coelicolor malate synthase was found to be almost 8-fold higher than the specific activity of S. clavuligerus malate synthase under similar expression conditions. Furthermore, the genomic organisation of the three Streptomyces aceB genes cloned thus far is different from that of other bacterial malate synthases, and warrants further investigation.     

Complete genome sequence of the erythromycin-producing bacterium Saccharopolyspora erythraea NRRL23338

Saccharopolyspora erythraea is used for the industrial-scale production of the antibiotic erythromycin A, derivatives of which play a vital role in medicine. The sequenced chromosome of this soil bacterium comprises 8,212,805 base pairs, predicted to encode 7,264 genes. It is circular, like those of the pathogenic actinomycetes Mycobacterium tuberculosis and Corynebacterium diphtheriae, but unlike the linear chromosomes of the model actinomycete Streptomyces coelicolor A3(2) and the closely related Streptomyces avermitilis. The S. erythraea genome contains at least 25 gene clusters for production of known or predicted secondary metabolites, at least 72 genes predicted to confer resistance to a range of common antibiotic classes and many sets of duplicated genes to support its saprophytic lifestyle. The availability of the genome sequence of S. erythraea will improve insight into its biology and facilitate rational development of strains to generate high-titer producers of clinically important antibiotics.     

Coelichelin, a new peptide siderophore encoded by the Streptomyces coelicolor genome: structure prediction from the sequence of its non-ribosomal peptide synthetase

A gene cluster for the non-ribosomal synthesis of a peptide of unknown structure has been identified in the partial genome sequence of Streptomyces coelicolor. Using molecular and computational analyses, the total structure of a tripeptide siderophore synthesized by the non-ribosomal peptide synthetase within the cluster has been deduced from the translated sequence of its encoding gene. This represents a novel method for the structural assignment of natural products from genome sequence data.     

A genetic and bioinformatic analysis of Streptomyces coelicolor genes containing TTA codons, possible targets for regulation by a developmentally significant tRNA

The rarest codon in the high G+C genome of Streptomyces coelicolor is TTA, corresponding in mRNA to the UUA codon that is recognized by a developmentally important tRNA encoded by the bldA gene. There are 145 TTA-containing genes in the chromosome of S. coelicolor. Only 42 of these are represented in the genome of Streptomyces avermitilis, among which only 12 have a TTA codon in both species. The TTA codon is less represented in housekeeping genes and orthologous genes, and is more represented in functional-unknown, extrachromosomal or weakly expressed genes. Twenty one TTA-containing chromosomal genes in S. coelicolor were disrupted, including 12 of the 42 genes that are common to both S. avermitillis and S. coelicolor. None of the mutant strains showed any obvious phenotypic differences from the wild-type strain under tested conditions. Possible reasons for this, and the role and evolution of the observed distribution of TTA codons among Streptomyces genes were discussed.     

Biosynthesis of polyketides in heterologous hosts.

Polyketide natural products show great promise as medicinal agents. Typically the products of microbial secondary biosynthesis, polyketides are synthesized by an evolutionarily related but architecturally diverse family of multifunctional enzymes called polyketide synthases. A principal limitation for fundamental biochemical studies of these modular megasynthases, as well as for their applications in biotechnology, is the challenge associated with manipulating the natural microorganism that produces a polyketide of interest. To ameliorate this limitation, over the past decade several genetically amenable microbes have been developed as heterologous hosts for polyketide biosynthesis. Here we review the state of the art as well as the difficulties associated with heterologous polyketide production. In particular, we focus on two model hosts, Streptomyces coelicolor and Escherichia coli. Future directions for this relatively new but growing technological opportunity are also discussed.     

Biosynthesis of Polyketides in Heterologous Hosts

Polyketide natural products show great promise as medicinal agents. Typically the products of microbial secondary biosynthesis, polyketides are synthesized by an evolutionarily related but architecturally diverse family of multifunctional enzymes called polyketide synthases. A principal limitation for fundamental biochemical studies of these modular megasynthases, as well as for their applications in biotechnology, is the challenge associated with manipulating the natural microorganism that produces a polyketide of interest. To ameliorate this limitation, over the past decade several genetically amenable microbes have been developed as heterologous hosts for polyketide biosynthesis. Here we review the state of the art as well as the difficulties associated with heterologous polyketide production. In particular, we focus on two model hosts, Streptomyces coelicolor and Escherichia coli. Future directions for this relatively new but growing technological opportunity are also discussed.     

Mapping of genes involved in macromolecular synthesis on the chromosome of Streptomyces coelicolor A3(2).

The genes for the beta, beta', and seven sigma factor subunits of RNA polymerase, for elongation factors EF-Tu1 and EF-Tu3, and for six rRNA operons were mapped on the combined genetic and physical map of the Streptomyces coelicolor chromosome. Like the previously mapped tRNA genes, the RNA polymerase and rRNA genes map to scattered positions. The lack of rRNA operons in the immediate vicinity of the origin of replication (oriC) and the absence of tRNA genes in any of the rRNA operons are novel features of the Streptomyces chromosome.     

天蓝色链霉菌Streptomyces coelicolor中几丁质酶C(Chi C)的生物信息学分析

利用生物信息学的方法,分析天蓝色链霉菌Streptomyces coelicolor中几丁质酶C（Chi C）的一些基本性质,并针对链霉菌属菌种的几个几丁质酶基因做了进化树,进而验证了天蓝色链霉菌中至少8种几丁质酶的分类;同时对天蓝色链霉菌Streptomyces coelicolor中几丁质酶C（Chi C）蛋白的高级结构作出了预测,得到其编码的属于18家族的蛋白质高级结构图谱。