Cloning of genes governing the deoxysugar portion of the erythromycin biosynthesis pathway in Saccharopolyspora erythraea (Streptomyces erythreus).

Genes that govern the formation of deoxysugars or their attachment to erythronolide B and 3 alpha-mycarosyl erythronolide B, intermediates of the biosynthesis of the 14-membered macrolide antibiotic erythromycin, were cloned from Saccharopolyspora erythraea (formerly Streptomyces erythreus). Segments of DNA that complement the eryB25, eryB26, eryB46, eryC1-60, and eryD24 mutations blocking the formation of erythronolide B or 3 alpha-mycarosyl erythronolide B, when cloned in Escherichia coli-Streptomyces shuttle cosmids or plasmid vectors that can transform S. erythraea, were located in a ca. 18-kilobase-pair region upstream of the erythromycin resistance (ermE) gene. The eryC1 gene lies just to the 5' side of ermE, and one (or possibly two) eryB gene is approximately 12 kilobase pairs farther upstream. Another eryB gene may be in the same region, while an additional eryB mutation appears to be located elsewhere. The eryD gene lies between the eryB and eryC1 genes and may regulate their function on the basis of the phenotype of an EryD- mutant.     

Biogenesis and regulation of biosynthesis of erythromycins in Saccharopolyspora erythraea: a review

Data on the structure and stages of biosynthesis of erythromycins, relating to (1) successive addition of L-mycarose and D-desosamine to the lactones erythronolide B and mycarosyl-erythronolide B, respectively, and (2) biotransformation of erythromycin D to erythromycin A, are presented. Pathways of biosynthesis of L-mycarose, D-desosamine, and methylmalonyl-CoA and methylpropionyl-CoA precursors of erythronolide B are reviewed, along with the properties of genes coding the enzymes involved. Possible mechanisms of biochemical and gene regulation of erythromycin biosynthesis in Saccharopolyspora erythraea are discussed, including the role of factors ensuring predominant formation of the target product, erythromycin A.     

New 4-O-substituted xylosyl units in the xyloglucan from leaves of Hymenaea courbaril

A homogeneous fucogalactoxyloglucan, isolated from the leaves of Hymenaea courbaril, was analysed by methylation-GC–MS. These procedures involved derived partially O-methylated alditol acetates and acetylated aldononitriles, which demonstrated the presence of both 2-O- and 4-O-substituted Xylp units in the side-chains. The presence of the unusual, latter structure was confirmed by 2D NMR spectroscopy with a correlated HMQC C-4/H-4 signal at δ 77.8/3.73. A similar 4-O-substituted xylosyl structure was present in a decasaccharide Glc₄Xyl₃Gal₂Fuc obtained via endo-glucanase treatment of the polysaccharide, which gave rise to a molecular ion with m/z 1555 (ESI-MS, Na⁺ form).     

Pustulan and branched β-galactofuranan from the phytopathogenic fungus Guignardia citricarpa, excreted from media containing glucose and sucrose

Guignardia citricarpa is a phytopathogenic fungus and the causal agent of citrus black spot. Incubation in a semi-defined media resulted in formation of exopolysaccharides [EPS(s)]. A medium containing glucose gave rise to a (1→6)-linked β-glucan (200 kD), pustulan, which was characterized by NMR and methylation analysis. A sucrose-containing medium provided a homogalactan (376 kD) and methylation analysis showed nonreducing end- (20%), 6-O- (53%) and 5,6-di-O-substituted Galf units (27%). An HMQC spectrum of the homogalactan showed C-1/H-1 signals at δ 108.2/4.820, 108.3/4.820 and 107.1/5.079, corresponding to three types of β- -Galf units. A DEPT analysis showed inverted signals (CH₂) at δ 67.8 and 67.2, corresponding to 6-O-substituted β- -Galf units, whereas a C-5 signal at δ 77.0 suggests 5-O-substitution, confirming a novel structure for a β-galactofuranan.     

An acetophenone and three naphthalides from turkish Rhamnus libanoticus

A new acetophenone glycoside and two new naphthalide glycosides have been isolated from the bark of Turkish Rhamnus libanoticus together with 7-hydroxy-5-methoxyphthalide 7-O-β-D-glucoside. The structures of the new compounds were elucidated by spectroscopic methods as 2,6-dihydroxy-4-methoxyacetophenone 2-O-β- rutinoside, 8,9-dihydroxy-6-methoxynaphthalide 8-O-β-rutinoside, 8,9-dihydroxy-6-methoxynaphthalide 8-O-/3b-D glucoside, respectively.     

Biogenesis and Regulation of Biosynthesis of Erythromycins in Saccharopolyspora erythraea

Data on the structure and stages of biosynthesis of erythromycins, relating to (1) successive addition of L-mycarose and D-desosamine to the lactones erythronolide B and mycarosyl-erythronolide B, respectively, and (2) biotransformation of erythromycin D to erythromycin A, are presented. Pathways of biosynthesis of L-mycarose, D-desosamine, and methylmalonyl-CoA and methylpropionyl-CoA precursors of erythronolide B are reviewed, along with the properties of genes coding the enzymes involved. Possible mechanisms of biochemical and gene regulation of erythromycin biosynthesis in Saccharopolyspora erythraea are discussed, including the role of factors ensuring predominant formation of the target product, erythromycin A.     

Chemical structure of kenaf xylan

Methylation and partial acid hydrolysis of xylans from the bast and core of kenaf (Hibiscus cannabinus) showed that the main chain of these xylans consists of (1 → 4)-linked β-d-xylopyranosyl (Xylp) residues, some of which carry a -1,2-linked 4-O-methyl-glucopyranosyluronic acid (Me-GlcAp) and glucopyranosyluronic acid (GlcAp) residues as side chains. Partial hydrolysis of kenaf xylans afforded two series of aldouronic acids from aldobio- to aldotetraouronic acids. The acids of the first series composed of 4-O-Me-d-GlcAp and d-Xylp residues: 4-O-Me-GlcA-Xyl₃, 4-O-Me-GlcA-Xyl₂ and 4-O-Me-GlcA-Xyl. The second series composed of d-GlcAp and d-Xylp: GlcA-Xyl₃, GlcA-Xyl₂ and GlcA-Xyl.

In addition to these acids, another aldobiouronic acid, 4-O-(-d-GalAp)-d-Xyl was found to be present in the partial hydrolysate.

The molar ratio of GalA, GlcA, 4-O-Me-GlcA, and Xyl residues was calculated to be 1.0:2.0:9.4:119 for the bast xylan and 1.0:1.3:7.9:99.4 for the core xylan.     

Syntheses of branched-chain sugars with push-pull functionality

The reaction of methyl 4,6-O-benzylidene-3(2)-deoxy-- -erythro-hexopyranosid-2(3)-ulose with carbon disulfide, alkyl iodide, and sodium hydride gave methyl 4,6-O-benzylidene-3(2)-[bis(alkylthio)methylene]-3(2)-deoxy-- -erythro-hexopyranosid-2(3)-uloses. Methyl 4,6-O-benzylidene-2-[bis(methylthio)methylene]-2-deoxy-- -erythro-hexopyranosid-3-ulose (5) reacted with aromatic amines to give, in a rearrangement process, N-aryl-2-aryliminomethyl-4,6-O-benzylidene-2-deoxy-- -erythro-hex-1-enopyranosylamin-3-uloses. The reaction of 5 which hydrazine hydrate afforded 5-methylthio-(methyl-4,6-O-benzylidene-2,3-dideoxy-- -erythro-hexopyranosido)[3,2-c]pyrazole.     

Parallel pathways for oxidation of 14-membered polyketide macrolactones in Saccharopolyspora erythraea

The glycosyltransferases OleG1 and OleG2 and the cytochrome P450 oxidase OleP from the oleandomycin biosynthetic gene cluster of Streptomyces antibioticus have been expressed, either separately or from artificial gene cassettes, in strains of Saccharopolyspora erythraea blocked in erythromycin biosynthesis, to investigate their potential for the production of diverse novel macrolides from erythronolide precursors. OleP was found to oxidize 6-deoxyerythronolide B, but not erythronolide B. However, OleP did oxidize derivatives of erythronolide B in which a neutral sugar is attached at C-3. The oxidized products 3-O-mycarosyl-8a-hydroxyerythronolide B, 3-O-mycarosyl-8,8a-epoxyerythronolide B, 6-deoxy-8-hydroxyerythronolide B and the olefin 6-deoxy-8,8a-dehydroerythronolide B were all isolated and their structures determined. When oleP and the mycarosyltransferase eryBV were co-expressed in a gene cassette, 3-O-mycarosyl-6-deoxy-8,8a-dihydroxyerythronolide B was directly obtained. When oleG2 was co-expressed in a gene cassette together with oleP, 6-deoxyerythronolide B was converted into a mixture of 3-O-rhamnosyl-6-deoxy-8,8a-dehydroerythronolide B and 3-O-rhamnosyl-6-deoxy-8,8a-dihydroxyerythronolide B, confirming previous reports that OleG2 can transfer rhamnose, and confirming that oxidation by OleP and attachment of the neutral sugar to the aglycone can occur in either order. Similarly, four different 3-O-mycarosylerythronolides were found to be substrates for the desosaminyltransferase OleG1. These results provide additional insight into the nature of the intermediates in OleP-mediated oxidation, and suggest that oleandomycin biosynthesis might follow parallel pathways in which epoxidation either precedes or follows attachment of the neutral sugar.     

Chemotaxonomic significance of flavonoids and phenolic acids in the Hieracium rohacsense group (Hieracium sect. Alpina; Lactuceae, Compositae)

Five apomictic taxa from the Hieracium rohacsense group were studied for their phenolic constituent composition. The following substances represent dominant compounds in the leaves: chlorogenic acid, 3,5-dicaffeoylquinic acid, luteolin 7-O-β- -glucopyranoside, luteolin 4′-O-β- -glucuronopyranoside and apigenin 4′-O-β- -glucuronopyranoside. Within the group only quantitative differences were found, luteolin 7-O-glucoside being the most important chemotaxonomic marker. Each taxon has its own specific quantitative pattern, invariable within the taxon. Based on these characteristic profiles, H. rohacsense can be distinguished from a closely related and still undescribed taxon from Mt. Pip Ivan. The proportion of luteolin 7-O-glucoside to apigenin 4′-O-glucuronoside also clearly separates the individuals of two morphologically close species—H. ratezaticum and H. pseudocaesium, which corresponds to a few slight but recognisable morphological and phenological characteristics. The ontogenetic stage of leaf development and seasonal variation are also important factors, which must be taken into consideration, as the quantity of the substances changes during leaf ontogeny and with season.     

Highly increased cellular accumulation of vincristine, a useful hydrophobic antitumor-drug, in multidrug-resistant solid cancer cells induced by a simply reduced taxinine

Regio- and/or chemo-selective reductions of taxinine (1a), a taxane diterpenoid readily obtainable from the needles of a Japanese yew (Taxus cuspidata), at the 5-O-cinnamoyl and 4-exo-methylene moieties have been accomplished by the catalytic hydrogenation over Pd/C or Rh/C to obtain 5-O-phenylpropionylated taxinine A (1b), 5-O-cyclohexylpropionylated taxinine A (1c), and 5-O-phenylpropionylated 4,20-dihydrotaxinine A (2a) in almost quantitative yields, respectively. Among them, taxoid 1b was found to be highly effective in increasing the cellular accumulation of vincristine in the multidrug-resistant human ovarian cancer cells compared with the cases of verapamil and the previously reported taxoids.     

Flavonoids of Astilbe and Rodgersia compared to Aruncus

The flavonoid profiles of Astilbe (four taxa studied) and Rodgersia (two taxa studied) are based on simple flavonol glycosides. Astilbe has 3-O-mono-, 3-O-di-, and 3-O-triglycosides of kaempferol, quercetin, and myricetin, while Rodgersia has only mono- and diglycosides of kaempferol and quercetin. Astilbe×arendsii was also shown to accumulate dihydrochalcone glycosides. The flavonoid profile of Rodgersia is the simplest recorded so far in the herbaceous Saxifragaceae. The flavonoids of two species of Aruncus were shown to be based upon kaempferol and quercetin 3-O-mono- and 3-O-diglycosides. One of the species also exhibited an eriodictyol glycoside. The triglycoside differences were not considered important, but the differences in myricetin occurrences were taken as evidence against derivation of Saxifragaceae from an Aruncus-like ancestor. Should such an event be proposed, however, serious consideration would have to be given to the current pattern of myricetin occurrence in the two families.     

Flavonoids of the Hydrangeaceae Dumortier

Fourteen species representing nine genera of the Hydrangeaceae Dumortier were surveyed for their flavonoid pigments. All taxa exhibited profiles based upon common flavonols. Myricetin was seen in two genera: Jamesia and Decumaria. Jamesia was further distinguished by the absence of kaempferol or its glycosides. A complex array of 3-O-mono-, 3-O-di- and 3-O-triglycosides was observed, although not all species had all levels of glycosylation. Decumaria barbara was unique within the species studied in its possession of 3,7-di- and 3,7-triglycosides. The overall pattern of flavonol glycosides observed for the Hydrangeaceae closely resembles that found in herbaceous genera of Saxifragaceae. The comparatively low frequency of myricetin contrasts with its high occurrence in herbaceous genera.     

Choleretic effects of yarrow (Achillea millefolium s.l.) in the isolated perfused rat liver

Different species from the Achillea millefolium aggregate are used against gastrointestinal and hepato-biliary disorders in traditional European medicine. In this work, a fraction enriched in dicaffeoylquinic acids (DCCAs) and luteolin-7-O-β-d-glucuronide was investigated on its choleretic effect in the isolated perfused rat liver (IPRL) compared to cynarin (1,3-DCCA), the main choleretic compound of Cynara scolymus L. A fraction containing 3,4-, 3,5- and 4,5-DCCA and luteolin-7-O-β-d-glucuronide was prepared by solid phase extraction from a 20% methanolic extract of yarrow. A total amount of 48.8% DCCAs and 3.4% luteolin-7-O-β-d-glucuronide was determined by HPLC analysis with cynarin as internal standard. IPRL experiments revealed a dose-dependant increase in bile flow (23–44–47%) by the Achillea fraction. Choleresis was two- to three-fold higher than that of cynarin. The combined effect of DCCAs and luteolin-7-O-β-d-glucuronide stimulated bile flow more effectively than the single compound cynarin. Due to their polar structure, these compounds are quantitatively extracted into teas and tinctures; hence, they seem to be the choleretic active principles in the traditional application forms of yarrow.     

Biotransformation of paeonol by Panax ginseng root and cell cultures

Panax ginseng root and cell cultures were shown to biotransform paeonol (1) into its 2-O-β-d-glucopyranoside (2). P. ginseng root cultures were also able to biotransform paeonol (1) into its 2-O-β-d-xylopyranoside (3), 2-O-β-d-glucopyranosyl(1 → 6)-β-d-glucopyranoside (4) and 2-O-β-d-xylopyranosyl(1 → 6)-β-d-glucopyranoside (5), and its demethylated derivate, 2′,4′-dihydroxyacetophenone (6). Compounds 3 and 4 are new glycosides. It is the first example that the administrated compound was converted into its xylopyranoside by plant biotransformation.     

Pregnane glycosides from fruits of Balanites aegyptiaca

From the mesocarp of Balanites aegyptiaca fruits, two pregnane glycosides were isolated. One is new and identified as pregn-5-ene-3β,16β,20(R)-triol 3-O-(2,6-di-O--l-rhamnopyranosyl)-β-d-glucopyranoside (balagyptin), while the other is known and assigned as pregn-5-ene-3β,16β,20(R)-triol 3-O-β-d-glucopyranoside.     

Microbial metabolism of 1-aminoanthracene by Beauveria bassiana

The carcinogen and mutagen, 1-aminoanthracene, was efficiently metabolized by the fungal strain Beauveria bassiana ATCC 7159 to yield three new metabolites identified as 1-acetamido-5-[(4′-O-methyl-β-d-glucopyranosyl)oxy]anthracene, 1-acetamido-8-[(4′-O-methyl-β-d-glucopyranosyl)oxy]anthraquinone, and 1-acetamido-6-[(4′-O-methyl-β-d-glucopyranosyl)oxy]anthraquinone, together with 1-acetamidoanthracene and 1-acetamidoanthraquinone. Formation of these metabolites suggests that the metabolic pathways of 1-aminoanthracene in B. bassiana ATCC 7159 involve acetylation, oxidation, hydroxylation, and O-methylglucosylation.     

Synthesis of a di- and a tri-saccharide related to the K-antigen of Klebsiella type 10 and a study of their inhibition in the precipitin reaction

Syntheses of methyl 3-O-- -galactopyranosyl-- -mannopyranoside (10) and methyl 3-O-- -galactopyranosyl-2-O-(β- -glucopyranosyluronic acid)-- -mannopyranoside (11) in good yield are described. Both 10 and 11 significantly inhibit antigen-antibody precipitation in the Klebsiella Type 10 immune system. The results provide more evidence for the structure (1) of the antigen from Klebsiella K-10 and its immunodominant grouping.     

Flavonoids of Helianthus series Microcephali

Ray flower and leaf flavonoids were investigated for the three species of Helianthus series Microcephali. Ray flowers of all species contain coreopsin, sulphurein, and quercetin 7-O-glucoside; those of H. microcephalus also contain quercetin 3-O-glucoside. A mixture of flavonoid aglycones, mostly methoxylated flavones, occurs in leaves of H. microcephalus, but not in H. glaucophyllus or H. laevigatus which also lack the glandular trichomes that in Helianthus are typically associated with flavonoid aglycones. The presence of compounds with the 6,8,4′ pattern of methoxylation in H. microcephalus suggests that the series is more similar in flavonoids to series Angustifolii than to series Corona-solis.