Isolation and characterization of new p-Terphenyls with antifungal, antibacterial, and antioxidant activities from halophilic actinomycete Nocardiopsis gilva YIM 90087.

A new p-terphenyl 1 and a novel p-terphenyl derivative 3 bearing a benzothiazole moiety were isolated from halophilic actinomycete Nocardiopsis gilva YIM 90087, along with known p-terphenyl 2, antibiotic novobiocin 4, cyclodipeptides 5-13, and aromatic acids 14 and 15. Their structures were elucidated on the basis of the interpretation of spectral data and by comparison of the corresponding data with those reported previously. The p-terphenyl 1 showed antifungal activity against the three pathogenic fungi, including Fusarium avenaceum, Fusarium graminearum, and Fusarium culmorum, that caused Fusarium head blight with minimal inhibitory concentrations (MICs) of 8, 16, and 128 µg/mL, respectively. Compound 1 showed antifungal activity against Candida albicans with a MIC of 32 µg/mL and antibacterial activity against Bacillus subtilis with a MIC of 64 µg/mL. Novobiocin 4 showed antifungal activity against Pyricularia oryzae with a MIC of 16 µg/mL and antibacterial activity against B. subtilis with a MIC of 16 µg/mL and Staphylococcus aureus with a MIC of 64 µg/mL. The 1,1-diphenyl-2-picryl-hydrazyl assay suggested that 1, 3, and 4 exhibited 54.9% (2 mg/mL), 14.3% (4 mg/mL), and 47.7% (2 mg/mL) free radical scavenging activity, respectively. The positively charged 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid radical (ABTS(+•)) scavenging assay indicated that 1, 3, 4, and 8 exhibited 68.6% (1 mg/mL), 28.4% (2 mg/mL), 78.2% (0.5 mg/mL), and 54.6% (2 mg/mL) ABTS(+•) scavenging capacity, respectively. The superoxide anion radical scavenging assay suggested that 4 exhibited 77.9% superoxide anion radical scavenging capacity at 2 mg/mL. N. gilva YIM 90087 is a new resource for novobiocin 4.

Coumabiocins A-F, aminocoumarins from an organic extract of Streptomyces sp. L-4-4.

Bioassay-directed fractionation of the butanol extract of Streptomyces sp. L-4-4, using the hyphae formation inhibition assay of a prokaryotic whole cell, led to the isolation of six new aminocoumarins, coumabiocins A-F (1-6), along with two known compounds, novobiocin (7) and isonovobiocin (8). Coumabiocins A-E (1-5) contain three structural elements, a central 3-amino-7-hydroxycoumarin that is linked at the 3-amino group to a prenylated 4-hydroxybenzoic acid moiety and at the 7-position to an l-noviosyl sugar, while coumabiocin F (6) lacks the sugar moiety. Their structures were elucidated by spectroscopic methods including 1D- and 2D-NMR techniques and mass spectrometric analyses. Coumabiocins A-E (1-5) exhibited significant inhibitory activity against Streptomyces 85E and gave a 10-15 mm clear zone of inhibition at 20 microg/disk and a 10 mm bald and a 10 mm clear zone of inhibition at 5 and 10 microg/disk, respectively, whereas coumabiocin F (6) was inactive.

Crystallization and preliminary X-ray analysis of the O-methyltransferase NovP from the novobiocin-biosynthetic cluster of Streptomyces spheroides.

Crystals of recombinant NovP (subunit MW = 29 967 Da; 262 amino acids), an S-adenosyl-L-methionine-dependent O-methyltransferase from Streptomyces spheroides, were grown by vapour diffusion. The protein crystallized in space group P2, with unit-cell parameters a = 51.81, b = 46.04, c = 61.22 A, beta = 104.97 degrees. Native data to a maximum resolution of 1.4 A were collected from a single crystal at the synchrotron. NovP is involved in the biosynthesis of the aminocoumarin antibiotic novobiocin that targets the essential bacterial enzyme DNA gyrase.

Characterization of the formation of the pyrrole moiety during clorobiocin and coumermycin A1 biosynthesis.

The aminocoumarin antibiotics clorobiocin and coumermycin A(1) target the B subunit of DNA gyrase by presentation of the 5-methyl-pyrrolyl-2-carboxy ester moiety in the ATP-binding site of the enzyme. The pyrrolyl pharmacophore is derived by a four electron oxidation of a prolyl unit while tethered in phosphopantetheinyl thioester linkage to a peptidyl carrier protein (PCP) subunit. l-Proline is selected and activated as l-prolyl-AMP by adenylation domain enzymes (CloN4 and CouN4) and then installed as the thioester on the holo form of the PCP proteins CloN5 and CouN5. Enzymatic oxidation of the prolyl-S-PCP by the flavoprotein dehydrogenase CloN3 can be followed by rapid quench and subsequent electrospray ionization-Fourier transform mass spectrometry analysis of the acyl-S-protein substrate/product mixture to establish that a two-electron oxidized pyrrolinyl-S-enzyme transiently accumulates on the way to the four-electron oxidized, heteroaromatic pyrrolyl-2-carboxy-S-PCP acyl enzyme product.

TPU-0031-A and B, new antibiotics of the novobiocin group produced by Streptomyces sp. TP-A0556.

Two novel antibiotics, TPU-0031-A and B, were isolated from the culture broth of an actinomycete strain. The producing strain, TP-A0556, was identified as Streptomyces sp. based on the taxonomic study. The new antibiotics were obtained by solvent extraction and chromatographic purification. Spectroscopic analyses showed that TPU-0031-A and B were 7'-demethylnovobiocin and 5"-demethylnovobiocin, respectively. These compounds showed antibiotic activity against Gram-positive and -negative bacteria.

The properties of ion channels formed by the coumarin antibiotic, novobiocin, in lipid bilayers.

The coumarin antibiotic novobiocin forms ion channels of varying conductances in lipid bilayers. The conductances (about 20, 22, 14, 7 and 2 pS for 100 mM NH4Cl, CsCl, KCl, NaCl and LiCl, respectively) and selectivities (cation transference numbers in the range of 0.97-0.98) of one type of novobiocin-induced channel are similar to those found for channels formed by gramicidin A, an antibiotic of very different structure. The conductance of novobiocin channels of this type was independent of the species of the membrane lipid. This observation suggests that novobiocin molecules directly form these channels, and that channels are not formed through defects in lipid structure. The similarity in conductance and ion selectivity between channels induced by novobiocin and those formed by gramicidin A suggests that these structurally different molecules form channels with comparable internal diameter and internal surface charge distribution. Using HPLC purification we argue that the channel-forming activity of novobiocin is related to the activity of the novobiocin molecule itself, and not to a contaminant of the commercially available novobiocin sodium salt preparation.

Induction of resistance to novobiocin in the novobiocin-producing organism Streptomyces niveus.

During growth of Streptomyces niveus wild-type in the novobiocin production medium CDM the resistance of mycelia to novobiocin rises from about 25 micrograms/ml to over 200 micrograms/ml. (S. lividans, a novobiocin-sensitive strain, is resistant to approx. 10 micrograms/ml novobiocin.) The initial period of low level resistance extends from the time of inoculation of the culture until approx. 70 h when the culture is still in the growth phase. High level resistance is initiated before the start of novobiocin production and rises rapidly to a maximum level beyond the end of the growth phase. The rise in pH of the unbuffered CDM medium which occurs during S. niveus fermentation was shown not to be the cause of the change in novobiocin resistance. However, mycelia-free CDM from S. niveus cultures expressing high level novobiocin resistance was shown to contain a factor which induced high level novobiocin resistance in germinating S. niveus spores. Kinetic studies revealed that the inducer first appears in the culture medium before the switch to high level resistance begins and reaches its highest concentration before resistance reaches its maximum level.

[Structure of carminomycins II and III]. / Struktura karminomitsinov II i III.

Carminomycins II and III, the main components of the carminomycin complex were isolated in pure state. Their crystalline exalates and acetate of cardminomycin II were prepared. The PMR spectra of both carminomycins and the 13C-NMR spectra of the oxalates were obtained. The molecular weights of the antibiotics were determined by mass-spectrometry. On the basis of the PMR spectra it was shown that carminomycins II and III had similar structures and differed in the stereoisomerism of the nitrogen-free fragment linked to the amino sugar. This was confirmed by the 13C-NMR spectra. The above fragment (C7H15O3) is analogous to the fragment of baumycins A1 and A2 described earlier.

[Separation of novobiocin, isnovobiocin and descarbamyl novobiocin by a thin-layer chromatographic method]. / Razdelenie novobiotsina, izonovobiotsina i deskarbamilnonovobiotsina metodom khromatografii v tonkom sloe.

A thin-layer chromatographic method for separation of novobiocin, isonovobiocin and descarbamylnovobiocin using Silufol plates was developed. The method is simple and rapid. It provided clear separation of the components and using of diethyl ether, a simple individual separating solvent. The method allowed determination of novobiocin and the products of its isomerization in the fermentation broth, extracts and dry substances, as well as performance of operative control and regulation of the technological processes of fermentation and chemical purification of novobiocin.