Genomic Analyses Lead to Novel Secondary Metabolites

Abstract

Genomic analyses of Amycolatopsis orientalis ATCC 43491 strain, deposited as a vancomycin producer, revealed the presence of genetic loci for the production of at least 10 secondary metabolites other than vancomycin. One of these gene clusters, which contained a type I polyketide synthase, was predicted to direct the synthesis of novel class of compound, a glycosidic polyketide ECO-0501 (1). Screening of culture extracts for a compound with the predicted physicochemical properties of the product from this locus, led to the isolation of the 13-Oglucuronide of 13-hydroxy-2,12,14,16,22-pentamethyl-28-(N-methyl-guanidino)-octacosa-2,4,6,8,10,14,20,24-octaenoic acid (2-hydroxy-5-oxo-cyclopent-1-enyl)-amide (ECO-0501, 1). The structure, confirmed by spectral analyses including MS, and 1D and 2D NMR experiments, were in accord with that predicted by genomic analyses. ECO-0501 possessed strong antibacterial activity against a series of Gram-positive pathogens including several strains of methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococci (VRE). ECO-0501 was chemically modified by esterification (1aϳ1c), Nacetylation (1d) and hydrogenation (1e) in order to explore structure activity relationships (SAR). Keywords Amycolatopsis orientalis, ECO-0501, antibacterial, PKS I IntroductionDrug-resistant bacterial infections are a growing health concern. Resistance has been developed to every major class of antibiotics on the market, and an increasing number of pathogenic bacteria are becoming resistant to multiple classes of antibiotics, thereby limiting treatment options. Hence, there is a renewed urgency for the discovery of new classes of antibiotics for the treatment of drug resistant bacterial infections. To accelerate the discovery of such potential antibacterial candidates from natural resources a new, fast and efficient technology is needed. The genomics of secondary metabolite biosynthesis recently evolved to the point where analysis of the genome of an organism can define its biosynthetic capabilities for secondary metabolites. A genome scanning technique that has been developed in our laboratories, and used with our DECIPHER ® technology to analyze the genomes of actinomycetes for their secondary metabolite biosynthetic genes, greatly reduces the amount of sequencing required to define this capability [1,2]. This approach not only ascertains the potential of a producing organism, but it provides a handle to detect, isolate and structurally define a specific metabolite. We have demonstrated this approach in the isolation and structural determination of an antifungal Genomic Analyses Lead to Novel Secondary MetabolitesPart 3

Genetics and Genomics of the Genus Amycolatopsis

“…29 other secondary metabolite gene clusters were found to be present in the genome among which one of the clusters encoded for the synthesis of polyketide synthase compound ECO-0501. As this cluster and corresponding product was already identified and reported in literature [45], the focus shifted to identification of some other biosynthetic gene cluster. A type III PKS/NRPS hybrid gene cluster that showed high similarity to other glycopeptide gene clusters like balhimycin and teicoplanin was identified.…”

Isolation and Identification of Three New 5-Alkenyl-3,3(2H)-furanones from Two Streptomyces species using a Genomic Screening Approach

“…The two singlet peaks for methyl groups at d 1.45 and d 1.44 suggested that compound 1 is an equilibrium mixture of 2R and 2S anomers. This was confirmed by measuring the 1 H NMR in benzene-d 6 , where these two methyl signals occurred at a very different ratio. The 1 H NMR spectrum of 2 was similar to that of 1.…”

“…Analyses of these gene clusters led us to conclude that these microorganisms produce members of a group of very closely related secondary metabolites. Here, we report the isolation and identification of three new 5-alkenyl-3,3(2H)-furanones from these two streptomycetes [6]. …”

Analysis of the Ketosynthase Genes in Streptomyces and Its Implications for Preventing Reinvestigation of Polyketides with Bioactivities

“…Recent molecular approaches have successfully revealed the presence of large numbers of cryptic PKS gene in the genome of Streptomyces spp. Since the presence of novel PKS genes in organisms provides good clues for production of novel polyketides, the sequencing of PKS genes is often used as a screening method for the discovery of novel polyketides (Ayuso et al, 2005;McAlpine et al, 2005;Banskota et al, 2006). The objective of this work was to assess the use of a PCR-based technique in the preliminary screening for microorganisms which were as producer of potential novel polyketide against plant pathogen fungi.…”