Antifungal macrodiolide from Streptomyces sp.

Aerobic fermentation cultures of Streptomyces sp. produced an antifungal macrodiolide. This new antibiotic consists of two units of homononactic acid linked to form a cyclic diester. An unknown polypeptide was also isolated in trace quantities. The antibiotic with polypeptide complex showed high levels of antifungal activity compared with that of the macrodiolide alone. The macrodiolide also showed a stimulatory effect on some species of fungi. The production, purification, and characterization of these compounds are reported.     

Production of a streptomycin-Park nucleotide complex by Streptomyces griseus.

A compound (compound X) with antibacterial activity was isolated from early-exponential-phase cultures of the streptomycin producer Streptomyces griseus and from protoplast cultures of the same strain. The protoplast cultures produced a larger amount of compound X than did the young hyphae. Both the mycelia and the protoplasts incorporated 14C-labeled myo-inositol, a precursor of streptomycin, into compound X, which has an amino acid content related to that of the cell wall peptidoglycan of the producer strain. Compound X may contain a streptomycin molecule covalently bound to a cell wall precursor unit, i.e., to a Park nucleotide (J. T. Park, J. Biol. Chem. 194:897-904, 1952), through the glutamic acid of the pentapeptide component.     

Cloning, Expression and Characterization of Thermophilic and Alkalophilic N-acetylglucosaminidase from Streptomyces sp. NK52 for the Targeted Production of N-acetylglucosamine

A chitinolytic Streptomyces strain isolated from an alkaline habitat produced six different isozymes of chitinases. PCR amplification of DNA with chitinase domain specific primers yielded six amplicons of which, a 0.8 Kb fragment was cloned in pQE-30UA direct cloning vector and transformed into E. coli M15 cells (pREP4). The recombinant homodimer protein had a molecular mass of 44 kDa, and the 22 kDa monomers displayed 45 and 60 % activity in the presence of reducing agents. The size of the monomers is close to the predicted putative ORF of 17.8 kDa. The enzyme exhibits extreme pH and temperature optima of 10.0 and 70 °C respectively making it favorable for industrial applications. Its gene sequence revealed no homology with the reported N-acetylglucosaminidases suggesting that it could have novel attributes. This enzyme could be useful in the large scale production of N-acetylglucosamine which is currently having numerous therapeutic and commercial applications.     

Cephamycins, a New Family of β-Lactam Antibiotics I. Production by Actinomycetes, Including Streptomyces lactamdurans sp. n

A number of actinomycetes isolated from soil were found to produce one or more members of a new family of antibiotics, the cephamycins, which are structurally related to cephalosporin C. The cephamycins were produced in submerged fermentation in a wide variety of media by one or more of eight different species of Streptomyces, including a newly described species, S. lactamdurans. These antibiotics exhibit antibacterial activity against a broad spectrum of bacteria which includes many that are resistant to the cephalosporins and penicillins.     

Antifungal Molecules Produced by a New Salt-Tolerant and Alkaliphilic Streptomyces sp. BS30 Isolated from an Arid Soil

Proceedings of the National Academy of Sciences, India Section B: Biological Sciences - A salt-tolerant and alkaliphilic actinomycete BS30 isolated from a Saharan soil sample of Algeria, exhibited...     

Effect of Penicillin on Streptomycin Production by Streptomyces griseus

The correlation between the biosynthesis of the cell wall and the formation of streptomycin (SM) in Streptomyces griseus, influenced by a specific inhibitor of cell wall synthesis, was investigated. Penicillin, in subinhibitory concentrations (1 to 5 mug/ml), was added to cultures of S. griseus in different stages of its life cycle. The inhibitor decreased SM production, when young cultures were treated; however, there was an increase in SM formation when penicillin was added to older cultures. The explanation for these findings is discussed in detail.     

Ahmpatinin iBu,a new HIV-1 protease inhibitor,from Streptomyces sp. CPCC 202950

Ahmpatinin ⁱBu (1) and statinin ⁱBu (2), two new linear peptides, a novel pyrrolidine derivative, (−)-(S)-2-[3-(6-methylheptanamido)-2-oxopyrrolidin-1-yl] acetic acid (3), and three known pepstatin derivatives (4–6) along with their corresponding methanolysis artifacts (7–9) were isolated from Streptomyces sp. CPCC 202950. Their structures were elucidated on the basis of extensive spectroscopic data using Marfey''s analysis, chiral-phase HPLC, and ECD and OR calculation to determine the absolute configurations. Compound 1 contains an unusual amino acid, 4-amino-3-hydroxy-5-(4-methoxyphenyl)pentanoic acid (Ahmppa), and 3 is the first natural product with a 2-(3-amino-2-oxopyrrolidin-1-yl)acetic acid system. Compounds 1, 2, and 4–9 are HIV-1 protease inhibitors. In particular, ahmpatinin ⁱBu (1) exhibits significant inhibitory activity against HIV-1 protease with an IC₅₀ value of 1.79 nM. A preliminary structure–activity relationship is discussed.

Ahmpatinin ⁱBu and statinin ⁱBu, two new linear peptides, were isolated from Streptomyces sp. CPCC 202950. Ahmpatinin ⁱBu exhibited significant inhibitory activity against HIV-1 protease with an IC₅₀ value of 1.79 nM.     

Cycloheximide Production by Streptomyces griseus: Control Mechanisms of Cycloheximide Biosynthesis

Cycloheximide accumulation in a fermentation medium has been shown to be the product of the balance between synthesis and degradation of this antibiotic. Glucose has been shown to prevent cycloheximide degradation. Cycloheximide has been shown to interfere with its own synthesis probably due to feedback inhibition. Approaches for increasing cycloheximide titers in the light of these findings are discussed.     

Characterization of Two Seryl-tRNA Synthetases in Albomycin-Producing Streptomyces sp. Strain ATCC 700974

The Trojan horse antibiotic albomycin, produced by Streptomyces sp. strain ATCC 700974, contains a thioribosyl nucleoside moiety linked to a hydroxamate siderophore through a serine residue. The seryl nucleoside structure (SB-217452) is a potent inhibitor of seryl-tRNA synthetase (SerRS) in the pathogenic bacterium Staphylococcus aureus, with a 50% inhibitory concentration (IC₅₀) of ∼8 nM. In the albomycin-producing Streptomyces sp., a bacterial SerRS homolog (Alb10) was found to be encoded in a biosynthetic gene cluster in addition to another serRS gene (serS1) at a different genetic locus. Alb10, named SerRS2 herein, is significantly divergent from SerRS1, which shows high homology to the housekeeping SerRS found in other Streptomyces species. We genetically and biochemically characterized the two genes and the proteins encoded. Both genes were able to complement a temperature-sensitive serS mutant of Escherichia coli and allowed growth at a nonpermissive temperature. serS2 was shown to confer albomycin resistance, with specific amino acid residues in the motif 2 signature sequences of SerRS2 playing key roles. SerRS1 and SerRS2 are comparably efficient in vitro, but the K_m of serine for SerRS2 measured during tRNA aminoacylation is more than 20-fold higher than that for SerRS1. SB-217452 was also enzymatically generated and purified by two-step chromatography. Its IC₅₀ against SerRS1 was estimated to be 10-fold lower than that against SerRS2. In contrast, both SerRSs displayed comparable inhibition kinetics for serine hydroxamate, indicating that SerRS2 was specifically resistant to SB-217452. These data suggest that mining Streptomyces genomes for duplicated aminoacyl-tRNA synthetase genes could provide a novel approach for the identification of natural products targeting aminoacyl-tRNA synthetases.Aminoacyl-tRNA synthetases (aaRSs) are enzymes that catalyze the ligation of specific amino acids to their cognate tRNAs that are subsequently used in protein synthesis (13). While this process is essential and the reaction mechanisms are universal in all three domains of life, aaRS genes are remarkably evolutionarily divergent, leading to a wide range of aaRS proteins with subtle structural differences in various organisms (19). In most pathogenic bacteria or fungi, aaRSs are encoded by single-copy genes and are attractive antimicrobial drug targets for screening and designing enzyme inhibitors of high specificity (18, 22). For example, the antibiotic pseudomonic acid (mupirocin), a potent inhibitor of bacterial isoleucyl-tRNA synthetase, has been used clinically as a topical agent to prevent Staphylococcus aureus infection (6) and another novel class of aaRS inhibitors has also recently emerged (26). For many environmental bacteria, genomic analyses have revealed two or three homologous genes encoding aaRSs of the same amino acid specificity in one microorganism (21, 37). In several cases, one of these isozymes is demonstrably resistant to an enzyme inhibitor of another (15, 20, 37). This phenomenon is quite common in bacteria such as soil-dwelling actinomycetes that are capable of producing diverse secondary metabolites and antibacterial natural products. A mupirocin-producing Pseudomonas fluorescens uses such a self-resistance mechanism (6, 37), and a similar scenario has been postulated for Streptomyces parvulus producing borrelidin (a threonyl-tRNA synthetase inhibitor) (20). All known Streptomyces species and related bacterial genera host a large array of multiple aaRS gene homologs in their genomes (21). The aaRS genes can either stand alone or associate with biosynthetic gene clusters. Only one of these aaRSs has been biochemically characterized to be drug resistant (15). However, more recent evidence points to an additional physiological role of aaRSs, especially when the second aaRS gene is dedicated to a biosynthetic pathway (9, 10). Because aminoacyl-tRNAs have been shown to serve as substrates in other cellular processes, including phospholipid modification, peptidoglycan biosynthesis, and porphyrin biosynthesis (reviewed in reference 25), a pathway-specific aaRS would obviously meet the need for increasing local concentrations of the charged aminoacyl-tRNA. In Streptomyces viridifaciens, which produces the antibiotic valanimycin, the pathway-specific seryl-tRNA synthetase (SerRS) VlmL is believed to produce l-seryl-tRNA, from which the seryl residue is transferred directly to another reactant in the valanimycin biosynthetic pathway (10). However, vlmL does not encode a valanimycin-resistant SerRS.The Trojan horse antibiotic albomycin, produced by Streptomyces sp. strain ATCC 700974, has a thioribosyl nucleoside moiety linked to an iron-chelating siderophore through a serine residue (Fig. ​(Fig.1)1) (3). The seryl-glycine dipeptide-linked 4′-thioribosyl-N⁴-carbamyl-5-methyl-4-imino-cytidine structure, named SB-217452, has been isolated from a Streptomyces fermentation culture and identified as a potent inhibitor against Staphylococcus aureus SerRS (50% inhibitory concentration [IC₅₀], ∼8 nM) (31). It was speculated that SB-217452 could mimic seryl-adenylate, the intermediate of the SerRS-catalyzed reaction, thus blocking the aminoacylation activity of the enzyme. In fact, albomycin as a whole can be actively taken up by a bacterium through its siderophore-dependent iron acquisition system, which results in an extremely low MIC (11, 23). SB-217452 is released and activated from the prodrug albomycin inside cells by peptidases reported to be PepN in Escherichia coli and PepN and PepA in Salmonella enterica serovar Typhimurium (4). Several natural antimicrobial agents, such as microcin C (17) and agrocin 84 (14), employ similar Trojan horse strategies to transport hydrophilic aaRS inhibitors across the membrane into target cells. To further investigate this important class of emerging antibiotics, we first cloned and sequenced the albomycin biosynthetic gene cluster in Streptomyces sp. strain ATCC 700974 (38). A bacterial SerRS gene homolog (alb10) was identified in the gene cluster. Given several unusual aspects of the serylation systems in bacteria, yeasts, archaea, and mammalian mitochondria (reviewed in reference 34), it was of interest to study the proposed drug-resistant properties of Alb10 and use it as an example to explore the evolutionary divergence of Streptomyces aaRSs. Here we report the in vitro and in vivo characterization of two SerRSs, one pathway specific and the other housekeeping, in the albomycin-producing Streptomyces strain. Our findings now set the stage to elucidate the albomycin biosynthetic pathway, improve the production level of albomycin, and expand the discovery of microbial natural products that are aaRS inhibitors.Open in a separate windowFIG. 1.Structure of the Trojan horse antibiotic albomycin and the SerRS inhibitor SB-217452 linked to the hydroxamate siderophore.     

Cycloheximide Production by Streptomyces griseus: Alleviation of End-Product Inhibition by Dialysis-Extraction Fermentation

The use of dialysis fermentation with the continuous extraction of the dialysate has resulted in a twofold increase in the cycloheximide titer due to relief from product inhibition. Continuous extraction of the dialysate has eliminated the necessity for large reservoir volumes of fermentation medium normally used in dialysis fermentation. The apparatus used in this procedure is described.     

Antibacterial Activity of Streptomyces sp. VITMK1 Isolated from Mangrove Soil of Pichavaram,Tamil Nadu,India

Proceedings of the National Academy of Sciences, India Section B: Biological Sciences - The aim of the present study was to isolate antibacterial compound from Streptomyces sp. VITMK1 isolated from...     

Utilization of Carbon and Nitrogen Sources by Streptomyces kanamyceticus for Kanamycin Production

To select a suitable synthetic medium for kanamycin production, we tested a number of carbon and nitrogen compounds for their effect on growth of Streptomyces kanamyceticus ATCC 12853 as well on kanamycin production. Galactose was found to be the most suitable carbon source, though dextrin, soluble starch, and potato starch gave moderate yields. Sodium nitrate and glycine were adequate nitrogen sources for kanamycin production. There was, however, no direct relation between the growth of the organism and antibiotic formation. The pH of the medium might be an important factor for kanamycin formation, as media giving high kanamycin yields showed an alkaline pH without exception.     

Statistical Modeling for the Optimization of Bioluminescence Production by Newly Isolated Photobacterium sp. NAA-MIE

Proceedings of the National Academy of Sciences, India Section B: Biological Sciences - The utilization of bioluminescent bacteria has served as a powerful bioassay in environmental monitoring as a...     

星点设计-效应面法优化显齿蛇葡萄提取工艺

目的:采用星点设计-效应面法优化显齿蛇葡萄中总黄酮、二氢杨梅素、杨梅苷、杨梅素的超声提取工艺。方法:采用单因素和星点设计进行实验,以乙醇体积分数、溶媒比、提取时间为自变量,总黄酮、二氢杨梅素、杨梅苷、杨梅素得率为因变量,对各水平进行多元线性回归及二项式拟合,通过效应面法优化工艺参数,并对预测值和实际值进行比较分析。结果:确定最优提取工艺为30倍量65%乙醇提取1次,每次30 min。结论:星点设计效应面法可用于显齿蛇葡萄提取工艺的优化,模型预测性良好,提取条件稳定,实验设计方法有较高的可靠性。     

Cloning and characterization of the pyrrolomycin biosynthetic gene clusters from Actinosporangium vitaminophilum ATCC 31673 and Streptomyces sp. strain UC 11065

The pyrrolomycins are a family of polyketide antibiotics, some of which contain a nitro group. To gain insight into the nitration mechanism associated with the formation of these antibiotics, the pyrrolomycin biosynthetic gene cluster from Actinosporangium vitaminophilum was cloned. Sequencing of ca. 56 kb of A. vitaminophilum DNA revealed 35 open reading frames (ORFs). Sequence analysis revealed a clear relationship between some of these ORFs and the biosynthetic gene cluster for pyoluteorin, a structurally related antibiotic. Since a gene transfer system could not be devised for A. vitaminophilum, additional proof for the identity of the cloned gene cluster was sought by cloning the pyrrolomycin gene cluster from Streptomyces sp. strain UC 11065, a transformable pyrrolomycin producer. Sequencing of ca. 26 kb of UC 11065 DNA revealed the presence of 17 ORFs, 15 of which exhibit strong similarity to ORFs in the A. vitaminophilum cluster as well as a nearly identical organization. Single-crossover disruption of two genes in the UC 11065 cluster abolished pyrrolomycin production in both cases. These results confirm that the genetic locus cloned from UC 11065 is essential for pyrrolomycin production, and they also confirm that the highly similar locus in A. vitaminophilum encodes pyrrolomycin biosynthetic genes. Sequence analysis revealed that both clusters contain genes encoding the two components of an assimilatory nitrate reductase. This finding suggests that nitrite is required for the formation of the nitrated pyrrolomycins. However, sequence analysis did not provide additional insights into the nitration process, suggesting the operation of a novel nitration mechanism.     

Sporulation and the Production of Serine Protease and Cephamycin C by Streptomyces lactamdurans

Streptomyces lactamdurans, producer of the antibiotic cephamycin C, excretes at least two proteases. Physiological studies indicate that antibiotic synthesis and serine protease formation are coordinately regulated. Both are produced only after culture growth ends, and they appear with essentially identical kinetics. In addition, strains which produce superior levels of cephamycin C form equally superior levels of the serine protease. Genetic evidence reveals that the syntheses of the antibiotic and serine proteases are associated with sporulation. Mutants which fail to produce aerial hyphae (bald mutants) also fail to synthesize the antibiotic and serine proteases.     

Production of Agar-Derived Antioxidants and Single Cell Detritus from Gracilaria corticata Using Agarase from Microbulbifer sp. CMC-5

Proceedings of the National Academy of Sciences, India Section B: Biological Sciences - Microbulbifer strain CMC-5 reportedly degrades multiple polysaccharides including agar. In present study, the...     

Jejuketomycins A and B,polyketide glycosides with cancer cell migration inhibitory activity from Streptomyces sp. KCB15JA151

Two new polyketide glycosides jejuketomycins A (1) and B (2), were isolated from a culture of Streptomyces sp. KCB15JA151. Their chemical structures including the absolute configurations were determined by detailed analyses of the NMR and HRMS data and ECD calculations and spectral data. Compounds 1 and 2 possess an unusual 6/6/8 tricyclic ring system. Biological evaluation with the wound healing assay and time-lapse cell tracking analysis revealed that compounds 1 and 2 have significant inhibitory activities against cancer cell migration with low cytotoxicity.

Two new polyketide glycosides jejuketomycins A (1) and B (2), were isolated from a culture of Streptomyces sp. KCB15JA151.