Genome sequence of an industrial microorganism Streptomyces avermitilis: deducing the ability of producing secondary metabolites.

Streptomyces avermitilis is a soil bacterium that carries out not only a complex morphological differentiation but also the production of secondary metabolites, one of which, avermectin, is commercially important in human and veterinary medicine. The major interest in this genus Streptomyces is the diversity of its production of secondary metabolites as an industrial microorganism. A major factor in its prominence as a producer of the variety of secondary metabolites is its possession of several metabolic pathways for biosynthesis. Here we report sequence analysis of S. avermitilis, covering 99% of its genome. At least 8.7 million base pairs exist in the linear chromosome; this is the largest bacterial genome sequence, and it provides insights into the intrinsic diversity of the production of the secondary metabolites of Streptomyces. Twenty-five kinds of secondary metabolite gene clusters were found in the genome of S. avermitilis. Four of them are concerned with the biosyntheses of melanin pigments, in which two clusters encode tyrosinase and its cofactor, another two encode an ochronotic pigment derived from homogentiginic acid, and another polyketide-derived melanin. The gene clusters for carotenoid and siderophore biosyntheses are composed of seven and five genes, respectively. There are eight kinds of gene clusters for type-I polyketide compound biosyntheses, and two clusters are involved in the biosyntheses of type-II polyketide-derived compounds. Furthermore, a polyketide synthase that resembles phloroglucinol synthase was detected. Eight clusters are involved in the biosyntheses of peptide compounds that are synthesized by nonribosomal peptide synthetases. These secondary metabolite clusters are widely located in the genome but half of them are near both ends of the genome. The total length of these clusters occupies about 6.4% of the genome.

Actinohivin, a novel anti-HIV protein from an actinomycete that inhibits syncytium formation: isolation, characterization, and biological activities.

Blocking human immunodeficiency virus (HIV) entry into target cells is an important goal of HIV and acquired immune deficiency syndrome (AIDS) therapies. We have searched for anti-HIV substances from microorganisms using a syncytium formation assay system constructed with HeLa/CD4/Lac-Z and HeLa/T-env/Tat cells. We discovered a novel anti-HIV protein that inhibits syncytium formation, designated as actinohivin, from a cultured broth of a soil isolate, actinomycete strain K97-0003. ESI mass spectrometry of actinohivin isolated from the culture filtrate showed an ion with molecular mass of 12,520.3 Da. The amino acid sequence was determined by N-terminal Edman degradation of the intact protein and peptide fragments formed by endoproteinase digestions. Actinohivin consists of a 114-amino-acid chain that exhibits internal sequence triplication. Actinohivin inhibited both T-cell and macrophage tropic syncytium formation, with IC(50) values of 60 and 700 nM, respectively, and the cytopathic effect of HIV-1(IIIB) in MT-4 cells, with IC(50) value of 230 nM.

Zelkovamycin, a new cyclic peptide antibiotic from Streptomyces sp. K96-0670. I. Production, isolation and biological properties.

A new antibiotic termed zelkovamycin was isolated from the fermentation broth of Streptomyces sp. K96-0670 by solvent extraction, ODS column chromatography and preparative HPLC. Zelkovamycin showed antibacterial activity against Xanthomonas oryzae, Acholeplasma laidlawii, Pyricularia oryzae and Staphylococcus aureus.

Ferroverdins, inhibitors of cholesteryl ester transfer protein produced by Streptomyces sp. WK-5344. I. Production, isolation and biological properties.

Streptomyces sp. WK-5344, a soil isolate, was found to produce structurally related inhibitors of cholesteryl ester transfer protein (CETP). New active compounds, designated ferroverdins B and C, were isolated along with known ferroverdin A from the fermentation broth by solvent extraction, ODS column chromatography and silica gel column chromatography. All ferroverdins showed a dose-dependent inhibitory activity against human CETP. The IC50 values were 21, 0.62 and 2.2 microM for ferroverdins A, B and C, respectively, indicating that ferroverdin B is one of the most potent CETP inhibitors of microbial origin.

Amphistin, a new melanogenesis inhibitor, produced by an actinomycete.

A new melanogenesis inhibitor, named amphistin, was isolated from the fermentation broth of an actinomycete strain KP-3052. Amphistin was purified from the culture filtrate by the combination of cation exchange, gel filtration, and aminosilyl silica gel chromatographic methods. The structure of amphistin was elucidated as gamma-(beta-histidinoalanino)homoalanine by NMR experiments including 1H-15N HMBC experiment and other spectroscopic analyses. Amphistin inhibited the melanogenesis of B16 melanoma cells at concentration of 6.8 microM.

Malolactomycins C and D, new 40-membered macrolides active against Botrytis.

A strain of Streptomyces was found to produce two new components of the 40-membered ring macrolides, malolactomycins C and D. They inhibited the growth of Botrytis cinerea in an agar medium and a detached leaf method.

Malolactomycins C and D, new 40-membered macrolides active against Botrytis

A strain of Streptomyces was found to produce two new components of the 40-membered ring macrolides, malolactomycins C and D. They inhibited the growth of Botrytis cinerea in an agar medium and a detached leaf method.

Madindoline, a novel inhibitor of IL-6 activity from Streptomyces sp. K93-0711. I. Taxonomy, fermentation, isolation and biological activities.

Selective growth inhibition against IL-6-dependent cells was detected in fermentation extracts of a microbial strain, K93-0711, which was characterized as Streptomyces species. Active metabolite, termed madindoline A and B, were isolated, and the structure was determined to be 3a-hydroxy-indoline with diketocyclopentene at the N position. Madindoline A and B displayed dose-dependent inhibition of MH60 cells, an IL-6-dependent cell line, in presence of 0.1 U/ml IL-6. The IC50 for madindoline A and B against this cell line was 8 microM and 30 microM, respectively. These compounds did not inhibit the growth of cell lines which are not IL-6 dependent and the growth inhibition of the MH60 cell line was reversed by addition of excess, 0.4 U/ml, of IL-6 to the culture media. These compounds did not show any antimicrobial activity at a concentration of 1,000 micrograms/ml.

New antibiotics phthoxazolins B, C and D produced by Streptomyces sp. KO-7888.

New antibiotics, phthoxazolins B, C and D were isolated from the fermentation broth of Streptomyces sp. KO-7888. They are geometrical isomers of 10-hydroxyphthoxazolin A. They showed selective antifungal activity against Phytophthora parasitica in vitro and modest herbicidal activity in a laboratory test, but the potencies were different among isomers.

Cytosaminomycins, new anticoccidial agents produced by Streptomyces sp. KO-8119. I. Taxonomy, production, isolation and physico-chemical and biological properties.

Streptomyces amakusaensis KO-8119, a soil isolate, was found to produce a series of new anticoccidial compounds. Four active compounds, designated as cytosaminomycins A, B, C and D, were isolated from the fermentation broth of the producing strain by solvent extraction, silica gel column chromatography and preparative HPLC. Cytosaminomycins inhibited the growth of Eimeria tenella in an in vitro assay system using primary chicken embryonic cells as a host. No schizont in the cells was observed at concentrations ranging from 0.3 to 0.6 microgram/ml for cytosaminomycins A, B and C, and at 2.5 micrograms/ml for cytosaminomycin D.

New inhibitors of melanogenesis, OH-3984 K1 and K2. I. Taxonomy, fermentation, isolation and biological characteristics.

Melanogenesis inhibitors, OH-3984 K1 and K2 were isolated from fermentation broth of Streptomyces sp. OH-3984. OH-3984 K1 and K2 inhibited the melanogenesis of B16 melanoma cells at concentrations of 7.5 and 3.8 micrograms/ml, respectively, whereas inhibition of tyrosinase activity has not been observed. The microbial metabolites showed no antimicrobiological activities against Gram-positive and Gram-negative bacteria, fungi or yeast at a concentration of 1,000 micrograms/ml.

Phthoxazolin A, a specific inhibitor of cellulose biosynthesis from microbial origin. I. Discovery, taxonomy of producing microorganism, fermentation, and biological activity.

A new inhibitor of cellulose biosynthesis named phthoxazolin A was discovered as a metabolite of Streptomyces sp. OM-5714. A newly established screening method, which utilized a cellulose-containing fungus, Phytophthora sp. as a test organism, was successfully employed for discovery of the compound. Phthoxazolin A, C16H22N2O3 (MW of 290), is a lipophilic triene compound with an oxazole moiety. It has moderate antifungal activity against Phytophthora spp. only and has potent herbicidal activity. Phosphate-limited fermentation conditions favored production of the active compound.