Cyclopeptides from the Mushroom Pathogen Fungus Cladobotryum varium.

Three new cyclopeptides with serial Phe residues were identified with the aid of HPLC-DAD analysis, from the culture broth of Cladobotryum varium, a fungal pathogen causing mushroom cobweb disease. Cladoamides A (1) and B (2) have two consecutive N-methylphenylalanine units in the destruxin class cyclic depsipentapeptide framework, while cladoamide C (3) has a three consecutive Phe motif in a cyclopentapeptide structure. Of these three cyclopeptides, 1 showed potent autophagy-inducing activity at 10 µg/mL, comparable to a positive control, rapamycin. For the determination of the absolute configurations of the Ile residues in 1 and 3, new conditions for separating Ile and allo-Ile, using a pentafluorophenyl-bonded solid phase and methanolic solvent, were established within the analytical scheme of the advanced Marfey's method, thus offering a convenient alternative to the C3 Marfey's method, which requires elution with a three-solvent mixture. The sequence of two d-Phe and one l-Phe in 3 was determined through NMR chemical shift prediction by DFT-based calculations and chemical synthesis, which demonstrated the significance of noncovalent interactions in the accurate calculation of stable conformers for peptides with multiple aromatic rings.

Hikiamides A-C, Cyclic Pentadepsipeptides from Fusarium sp. TAMA 456.

Three new cyclic pentadepsipeptides, hikiamides A-C (1-3), were isolated from the culture extract of Fusarium sp. TAMA 456. The structures were determined by spectroscopic analysis using NMR and MS, and the absolute configurations were established by using Marfey's method and chiral HPLC analysis. Hikiamides induced the differentiation of murine ST-13 preadipocytes into mature adipocytes at 2 µM and adiponectin mRNA expression (5- to 13-fold higher than control). They also induced PPAR-γ-dependent gene expression at a concentration from 0.63 to 10 µM in a gene reporter assay.

Isolation and biosynthesis of preussin B, a pyrrolidine alkaloid from Simplicillium lanosoniveum.

A new pyrrolidine alkaloid, preussin B (1), was isolated from the culture extract of the fungus Simplicillium lanosoniveum TAMA 173 along with the known congener preussin (2). The structure and absolute configuration of 1 were determined by spectroscopic analysis and spectral comparison with 2. Feeding experiments with 13C-labeled precursors revealed that the pyrrolidine ring of 1 was assembled from acetate and l-phenylalanine by a PKS-NRPS hybrid biosynthetic pathway.

A high-throughput screen for inhibitors of the prolyl isomerase, Pin1, identifies a seaweed polyphenol that reduces adipose cell differentiation.

The peptidyl prolyl cis/trans isomerase Pin1 enhances the uptake of triglycerides and the differentiation of fibroblasts into adipose cells in response to insulin stimulation. Pin1 downregulation could be a potential approach to prevent and treat obesity-related disorders. In order to identify an inhibitor of Pin1 that exhibited minimal cytotoxicity, we established a high-throughput screen for Pin1 inhibitors and used this method to identify an inhibitor from 1,056 crude fractions of two natural product libraries. The candidate, a phlorotannin called 974-B, was isolated from the seaweed, Ecklonia kurome. 974-B inhibited the differentiation of mouse embryonic fibroblasts and 3T3-L1 cells into adipose cells without inducing cytotoxicity. We discovered the Pin1 inhibitor, 974-B, from the seaweed, E. kurome, and showed that it blocks the differentiation of fibroblasts into adipose cells, suggesting that 974-B could be a lead drug candidate for obesity-related disorders.

Marianins A and B, prenylated phenylpropanoids from Mariannaea camptospora.

Marianins A (1) and B (2), two new prenylated phenylpropanoids, were isolated from the culture extract of the fungus Mariannaea camptospora. Structures of marianins were elucidated by interpretation of NMR and other spectroscopic data. 1 is a 5-methylcoumarin bearing two prenyloxy groups, while 2 is an orcinol derivative substituted with a 3,3-dimethyl-4-pentenoyl chain. 2 is possibly derived from 1 through a Claisen rearrangement of the prenyl group, followed by lactone hydrolysis and decarboxylation. These compounds showed weak antibacterial activity against Micrococcus luteus.

New analogs of pochonicine, a potent ß-N-acetylglucosaminidase inhibitor from fungus Pochonia suchlasporia var. suchlasporia TAMA 87.

Three novel analogs of pochonicine (1) were isolated from a solid fermentation culture of the fungal strain Pochonia suchlasporia var. suchlasporia TAMA 87, and their structures were elucidated as 7-deoxypochonicine (2), 6-deoxypochonicine (3), and 6,7-dideoxypochonicine (4). These analogs were found to possess the same stereochemistry as pochonicine. Comparison of ß-N-acetylglucosaminidase (GlcNAcase) inhibitory activity between these analogs and pochonicine suggested that the C-6 hydroxy group of pochonicine was essential to its potent GlcNAcase inhibitory activity and that the C-7 hydroxy group also contributed to the activity, but to a lesser extent than the C-6 hydroxy group.

Pithohirolide, an antimicrobial tetradepsipeptide from a fungus Pithomyces chartarum.

Pithohirolide (1), a new depsipeptide, was isolated from an ascomycetous fungus Pithomyces chartarum TAMA 581. The planar structure of 1 was elucidated on the basis of NMR and MS analyses and the absolute configuration was determined by the advanced Marfey's analysis, chiral-phase HPLC analysis, and synthesis of degradation product. Compound 1 possesses a cyclic structure comprising (S)-2-hydroxy-3-phenylpropanoic acid, (S)-3-hydroxy-3-phenylpropanoic acid, (S)-2-hydroxyisovaleric acid, and N-methyl-L-alanine, connected via three ester and one amide linkages. Compound 1 exhibited antimicrobial activity against Staphylococcus aureus and Saccharomyces cerevisiae at MIC 3.1 µg ml-1.

The resorcylic acid lactone hypothemycin selectively inhibits the mitogen-activated protein kinase kinase-extracellular signal-regulated kinase pathway in cells.

The resorcylic acid lactone hypothemycin has been shown to inactivate protein kinases by binding to a cysteine conserved in 46 protein kinases, including mitogen-activated protein kinase kinase (MEK), extracellular signal-regulated kinase (ERK) and platelet-derived growth factor receptor (PDGFR). We assessed the selectivity of hypothemycin in cellular contexts. Hypothemycin normalized the morphology and inhibited anchorage-independent growth of Ki-ras transformed normal rat kidney (NRK) cells with selectivity and potency comparable to or greater than that of the MEK inhibitor U0126. In Ki-ras-transformed and phorbol 12-myristate 13-acetate (PMA)-treated NRK cells, hypothemycin blocked ERK activation but showed a minimal effect on autophosphorylation of protein kinase D1 (PKD1), another kinase containing the conserved cysteine. Hypothemycin potently inhibited PDGFR autophosphorylation and activation of the MEK-ERK pathway in platelet-derived growth factor (PDGF)-treated NRK cells. However, the phosphoinositide-3-kinase (PI3K) pathway was only modestly attenuated. Hypothemycin also inhibited growth factor- and anchorage-independent growth of human cancer cell lines with a constitutively active MEK-ERK pathway. Although hypothemycin has the potential to inactivate various protein kinases, the results indicate that in intracellular environments, hypothemycin can inhibit the MEK-ERK axis with sufficient selectivity to normalize transformed phenotypes of cells dependent on this pathway.

A new asteltoxin analog with insecticidal activity from Pochonia suchlasporia TAMA 87.

A new asteltoxin analog, named asteltoxin H (1), was isolated by the solid-state fermentation of the fungus Pochonia suchlasporia var. suchlasporia TAMA 87. The chemical structure of 1 was deduced by spectroscopic methods, including 1D and 2D NMR, HRESIMS, and UV-Vis analyses. Compound 1 showed insecticidal activity against prepupae of the blowfly, Lucilia sericata, with an LD50 value of 0.94 µg/mg prepupal body weight.

Pochonicine, a polyhydroxylated pyrrolizidine alkaloid from fungus Pochonia suchlasporia var. suchlasporia TAMA 87 as a potent beta-N-acetylglucosaminidase inhibitor.

A new polyhydroxylated pyrrolizidine alkaloid designated as pochonicine (1) was isolated from a solid fermentation culture of the fungal strain Pochonia suchlasporia var. suchlasporia TAMA 87. The structure of 1 was determined using NMR and MS techniques as (1R*, 3S*, 5S*, 6S*, 7R*, 7a S*)-5-acetamidomethyl-3-hydroxymethyl-1,6,7-trihydroxypyrrolizidine. Pochonicine (1) showed potent inhibition against beta-N-acetylglucosaminidases (GlcNAcases) of various organisms including insects, fungi, mammals, and a plant but no inhibition against beta-glucosidase of almond, alpha-glucosidase of yeast, or chitinase of Bacillus sp. The GlcNAcase inhibitory activity of pochonicine (1) was comparable to nagstatin, a potent GlcNAcase inhibitor of natural origin.