Marfey's Method Research Articles

Trichodestruxins A-D: Cytotoxic Cyclodepsipeptides from the Endophytic Fungus Trichoderma harzianum.

Four new cyclodepsipeptides, trichodestruxins A-D (1-4), together with destruxin E2 chlorohydrin (5) and destruxin A2 (6), were isolated from the plant endophytic fungus Trichoderma harzianum by a bioassay-guided fractionation method. Their planar structures were elucidated on the basis of 1D and 2D NMR and MS/MS spectroscopic analyses. The stereochemical configuration was established by application of the advanced Marfey's method, J-based configuration analysis, Mosher's method, and chemical derivatizations. Trichodestruxin B contains hydroxy acid fragments of the THPA unit, while trichodestruxins A and C contain the β-methylproline moiety. All cyclodepsipeptides displayed cytotoxicity against HT-29, A549, and/or P388 cell lines with IC50 values of 0.7-19.1 μM.

Zelkovamycins B-E, Cyclic Octapeptides Containing Rare Amino Acid Residues from an Endophytic Kitasatospora sp.

Four unusual cyclopeptides, zelkovamycins B-E (1-4), were isolated from an endophytic Kitasatospora sp. Zelkovamycin B was featured by an unprecedented 3-methyl-5-hydroxypyrrolidine-2,4-dione ring system linked to the cyclopeptide skeleton. Their structures and full configurations were established by spectroscopic analysis, Marfey's method, and NMR calculations. A plausible biosynthetic pathway for zelkovamycins was proposed based on gene cluster analysis. Zelkovamycin E displayed potent inhibitory activity against H1N1 influenza A virus.

Phakefustatins A-C: Kynurenine-Bearing Cycloheptapeptides as RXRα Modulators from the Marine Sponge Phakellia fusca.

A neutral-loss scanning mass method was used to explore new kynurenine-containing cycloheptapeptides, phakefustatins A-C (1-3), from the marine sponge Phakellia fusca. Their structures were elucidated by spectroscopic analysis and the advanced Marfey's method. 1 was total synthesized via a final-stage ozonolysis strategy by the combination of solid/solution-phase synthesis. Phakefustatin A (1) was identified as a RXRα modulator to inhibit cancer cell growth, and its pharmacophores could be Kyn and guanidine groups.

Isolation and characterization of a new cyclic lipopeptide surfactin from a marine-derived Bacillus velezensis SH-B74.

A marine-sediment-derived bacterium Bacillus velezensis SH-B74 can produce cyclic lipopeptides (CLPs). This study presented the isolation, characterization, and activity evaluation of a new CLP from the bacterial cultures of the strain SH-B74. Multiple chromatographic methods (solid-phase extraction and reversed-phase high-performance liquid chromatography) were applied to the purifying procedure of CLP, and the structural characterization of the new CLP was conducted by various spectroscopy (1D and 2D nuclear magnetic resonance together with Fourier transform infrared spectroscopy) and spectrometry (liquid chromatography-mass spectrometry, high-resolution mass spectrometry and tandem mass spectrometry) techniques as well as Marfey's method. The results displayed that the new CLP (anteiso-C15 Ile2,7 surfactin, 1) consists of a peptidic backbone of L-Glu1, L-Ile2, D-Leu3, L-Val4, L-Asp5, D-Leu6, L-Ile7, and an anteiso-C15 type saturated fatty acid chain. Further activity assay showed that the new CLP displays activity on the inhibition of the appressoria formation of rice blast causal pathogen Magnaporthe oryzae. To sum up, the results presented the perspective of potential application of the new CLP as a green agrichemical to control M. oryzae.

Basidiosins A and B: Cyclopentapeptides from the entomophthoralean fungus Basidiobolus meristosporus

Two new cyclopentapeptides, basidiosins A and B (1 and 2) were isolated from the mycelia extracts of entomophthoralean fungus Basidiobolus meristosporus RCEF 4516. The structures were determined based on spectroscopic methods, and the absolute configurations were assigned by Marfey's method on their acid hydrolyzates. Compounds 1 and 2 were identified as cyclo(L-Thr-L-Leu- L-Ile-D-Tyr-D-Thr) and cyclo(L-Thr-L-Leu-L-Val-D-Val-D-Ser), respectively. They were evaluated for the biological activities including antibacterial, antifungal and antioxidative activities. Furthermore, the biosynthetic pathway of 1 was proposed by bioinformatic analysis. This is the first study on the isolation of natural products from Basidiobolus fungus.

Structural Revision of Lydiamycin A by Reinvestigation of the Stereochemistry.

Lydiamycin A (1) is a previously reported piperazic acid-bearing cyclic peptide from Streptomyces. The absolute configuration of lydiamycin A has not been fully determined despite the fact that multiple total syntheses have been reported. The absolute configuration of 1 was reinvestigated by the advanced Marfey's method, chemical derivatization, and quantum-mechanics-based computational analysis, eventually resulting in a structural revision and establishment of the complete configuration. Lydiamycin A displayed weak antituberculosis activity in vitro.

Heterologous expression of a cryptic gene cluster from Grimontia marina affords a novel tricyclic peptide grimoviridin.

Microviridins are a class of ribosomally synthesized and post-translationally modified peptides (RiPPs) that have been isolated from a wide variety of cyanobacterial strains. There are similar gene clusters of RiPPs distributed in the genomes of bacteria belonging to the phyla Proteobacteria and Bacteroidetes. A cryptic gene cluster for the production of microviridin-type peptide was found in the genome of the marine γ-Proteobacterium Grimontia marina. Heterologous production of new microviridin-type peptide named grimoviridin was accomplished in Escherichia coli using the biosynthetic gene cluster of G. marina. The structure of grimoviridin was determined by analysis of MS and NMR data. Grimoviridin contained one isopeptide and two ester bonds, which had exactly the same bridging pattern as other microviridin-type peptides. The absolute stereochemistries of constituent amino acids were determined to be all L-forms by modified Marfey's method. Grimoviridin showed potent inhibitory activity against trypsin with an IC50 value of 238nM. This is the first report of heterologous production of microviridin-type peptide using a biosynthetic gene cluster from a Proteobacterium. Key points • Heterologous production afforded new microviridin-type peptide named grimoviridin. • This is the first report of microviridin-type peptide from proteobacterial origin. • Grimoviridin showed potent inhibitory activity against trypsin.

Thioamycolamides A-E, Sulfur-Containing Cycliclipopeptides Produced by the Rare Actinomycete Amycolatopsis sp.

A series of novel sulfur-containing cycliclipopeptides named thioamycolamides A-E, with thiazoline, thioether rings, and fatty acid moieties, were identified from the culture broth of the rare actinomycete Amycolatopsis sp. 26-4. The planar structural elucidation was accomplished by HRMS and 1D/2D NMR spectroscopic data analyses. The absolute configurations were unambiguously determined by Marfey's method, CD spectroscopy, and synthesis of partial structures. Moreover, their growth inhibitory activities against human tumor cell lines were investigated.

Meliponamycins: Antimicrobials from Stingless Bee-Associated Streptomyces sp.

Social insects establish complex interactions with microorganisms, some of which play defensive roles in colony protection. The important role of pollinators such as the stingless bee Melipona scutellaris in nature encouraged us to pursue efforts to study its associated microbiota. Here we describe the discovery of two novel cyclic hexadepsipeptides, meliponamycin A (1) and meliponamycin B (2), from Streptomyces sp. ICBG1318 isolated from M. scutellaris nurse bees. Their structures were established by interpretation of NMR and MS data, and the absolute configuration of the constituent amino acids was determined by the advanced Marfey's method. Compounds 1 and 2 showed strong activity against the entomopathogen Paenibacillus larvae and human pathogens Staphylococcus aureus and Leishmania infantum.

Heptavalinamide A, an Extensively N-Methylated Linear Nonapeptide from a Cyanobacterium Symploca sp. and Development of a Highly Sensitive Analysis of N,N-Dimethylvaline by LCMS.

An extensively N-methylated linear nonapeptide heptavalinamide A (1) was isolated from the marine cyanobacterium Symploca sp. collected at Kabira Reef of Ishigaki Island, Okinawa. The amino acid sequence of 1 was assigned by interpretation of 2D NMR and MS/MS data. The absolute configurations of the constituent amino acids were determined by the application of Marfey's method. A method to assign the configuration of N,N-dimethylvaline by LCMS is discussed.

Phototemtide A, a Cyclic Lipopeptide Heterologously Expressed from Photorhabdus temperata Meg1, Shows Selective Antiprotozoal Activity.

A new cyclic lipopeptide, phototemtide A (1), was isolated from Escherichia coli expressing the biosynthetic gene cluster pttABC from Photorhabdus temperata Meg1. The structure of 1 was elucidated by HR‐ESI‐MS and NMR experiments. The absolute configurations of amino acids and 3‐hydroxyoctanoic acid in 1 were determined by using the advanced Marfey's method and comparison after total synthesis of 1, respectively. Additionally, three new minor derivatives, phototemtides B–D (2–4), were identified by detailed HPLC–MS analysis. Phototemtide A (1) showed weak antiprotozoal activity against Plasmodium falciparum, with an IC50 value of 9.8 μm. The biosynthesis of phototemtides A–D (1–4) was also proposed.

14-Residue peptaibol velutibol A from Trichoderma velutinum: its structural and cytotoxic evaluation.

Velutibol A (1), a new 14-residue peptaibol was isolated from the Himalayan cold habitat fungus Trichoderma velutinum. The structural characterization was carried out by 1D and 2D NMR studies, and tandem mass studies, and Marfey's method aided in determining the stereochemistry of the amino acids. The CD analysis revealed folding of the peptide in a 310-helical conformation. The intramolecular H-bonding was determined by an NMR-VT experiment. Cytotoxic evaluation was carried out against a panel of cancer cell lines. The cell cycle assay was carried out on human myeloid leukaemia (HL-60) cells and revealed the formation of apoptotic bodies and DNA damage in a dose-dependent manner. Three other peptaibols namely velutibol B (2), velutibol C (3), and velutibol D (4) were also isolated in trace amounts from the psychotropic fungus and characterized through tandem mass spectroscopy and Marfey's analysis.

Structure, Biosynthesis, and Bioactivity of Photoditritide from Photorhabdus temperata Meg1.

A new cyclic peptide photoditritide (1), containing two rare amino acid d-homoarginine residues, was isolated from Photorhabdus temperata Meg1 after the nonribosomal peptide synthetase encoding gene pdtS was activated via promoter exchange. The structure of 1 was elucidated by HR-MS and NMR experiments. The absolute configurations of amino acids were determined according to the advanced Marfey's method after hydrolysis of 1. Bioactivity testing of 1 revealed potent antimicrobial activity against Micrococcus luteus with an MIC value of 3.0 μM and weak antiprotozoal activity against Trypanosoma brucei rhodesiense with an IC50 value of 13 μM. Additionally, the biosynthetic pathway of 1 was also proposed.

Scytodecamide from the Cultured Scytonema sp. UIC 10036 Expands the Chemical and Genetic Diversity of Cyanobactins.

Cyanobactins are a large family of cyanobacterial ribosomally synthesized and post-translationally modified peptides (RiPPs) often associated with biological activities, such as cytotoxicity, antiviral, and antimalarial activities. They are traditionally described as cyclic molecules containing heterocyclized amino acids. However, this definition has been recently challenged by the discovery of short, linear cyanobactins containing three to five amino acids as well as cyanobactins containing no heterocyclized residues. Herein we report the discovery of scytodecamide (1) from the freshwater cyanobacterium Scytonema sp. UIC 10036. Structural elucidation based on mass spectrometry, 1D and 2D NMR spectroscopy, and Marfey's method revealed 1 to be a linear decapeptide with an N-terminal N-methylation and a C-terminal amidation. The genome of Scytonema sp. UIC 10036 was sequenced, and bioinformatic analysis revealed a cyanobactin-like biosynthetic gene cluster consistent with the structure of 1. The discovery of 1 as a novel linear peptide containing an N-terminal N-methylation and a C-terminal amidation expands the chemical and genetic diversity of the cyanobactin family of compounds.

Trikoramide A, a Prenylated Cyanobactin from the Marine Cyanobacterium Symploca hydnoides.

A new cyclic decapeptide, trikoramide A (1), has been isolated from samples of the marine cyanobacterium Symploca hydnoides, collected from Bintan Island, Indonesia. Trikoramide A (1) is a C-prenylated cyclotryptophan-containing cyanobactin. Its planar structure was deduced by 1D and 2D NMR spectroscopy as well as HR-MS/MS data. In addition, its absolute configuration was determined by Marfey's method and 2D NOESY NMR spectroscopic analysis. Compound 1 possessed cytotoxicity against the MOLT-4 and AML2 cancer cell lines with IC50 values of 4.8 and 8.2 μM, respectively.

Natural Hydroxamate-Containing Siderophore Acremonpeptides A–D and an Aluminum Complex of Acremonpeptide D from the Marine-Derived Acremonium persicinum SCSIO 115

Four new hydroxamate-containing natural product cyclopeptides designated acremonpeptides A-D (1-4), together with Al(III)-acremonpeptide D (5) were obtained from the marine fungus Acremonium persicinum SCSIO 115. The planar structures of 1-5 were established on the basis of HRMS as well as 1D and 2D NMR data sets. Moreover, the amino acid absolute configurations were determined using Marfey's method. Compounds 1-5 all feature three 2-amino-5-(N-hydroxyacetamido)pentanoic acid (N5-hydroxy-N5-acetyl-l-ornithine) metal ion chelating moieties. Beyond their discovery and structure elucidation, in vitro bioassays revealed acremonpeptides A (1), B (2), and Al(III)-acremonpeptide D (5) as moderate antiviral agents for herpes simplex virus 1 with EC50 values of 16, 8.7, and 14 μM, respectively.

Neuroprotective Glycosylated Cyclic Lipodepsipeptides, Colletotrichamides A-E, from a Halophyte-Associated Fungus, Colletotrichum gloeosporioides JS419.

Glutamate neurotoxicity has been implicated in neuronal death in both acute CNS injury and in chronic neurodegenerative diseases. Five unique cyclic depsipeptides with neuroprotective activity, colletotrichamides A-E (1-5), were isolated from cultures of a halophyte Suaeda japonica-associated fungus, Colletotrichum gloeosporioides JS419. Spectroscopic analysis revealed that they were glycosylated cyclic lipodepsipeptides. Their relative configurations were determined by ROESY and J-based configuration analysis, and absolute configurations were established by chemical reactions including modified Mosher's method, advanced Marfey's method, and sugar derivatization. This is the first report of a glycosylated dimethyl-trioxygenated dodecanoyl moiety, and the relative as well as absolute stereochemistry was elucidated herein for the first time. Colletotrichamide C exhibited strong neuroprotective activity against glutamate in hippocampal HT22 cells.

Friomaramide, a Highly Modified Linear Hexapeptide from an Antarctic Sponge, Inhibits Plasmodium falciparum Liver-Stage Development.

The cold waters of Antarctica are known to harbor a rich biodiversity. Our continuing interest in the chemical analysis of Antarctic invertebrates has resulted in the isolation of friomaramide (1), a new, highly modified hexapeptide, from the Antarctic sponge Inflatella coelosphaeroides. The structure of friomaramide was determined using spectroscopic methods and its configuration established by Marfey's method. Friomaramide, which bears the unusual permethylation of the amino acid backbone and is the longest polypeptide bearing a tryptenamine C-terminus, blocks >90% of Plasmodium falciparum liver-stage parasite development at 6.1 μM.

Sarpeptins A and B, Lipopeptides Produced by Streptomyces sp. KO-7888 Overexpressing a Specific SARP Regulator.

Whole genome analysis of Streptomyces sp. KO-7888 has revealed various pathway-specific transcriptional regulatory genes associated with silent biosynthetic gene clusters. A Streptomyces antibiotic regulatory protein gene, speR, located adjacent to a novel nonribosomal peptide synthetase (NRPS) gene cluster, was overexpressed in the wild-type strain. The resulting recombinant strain of Streptomyces sp. KO-7888 produced two new lipopeptides, sarpeptins A and B. Their structures were elucidated by high-resolution electrospray ionization mass spectrometry, NMR analysis, and the advanced Marfey's method. The distinct modular sections of the corresponding NRPS biosynthetic gene cluster were characterized, and the assembly line for production of the lipopeptide chain was proposed.

Cipralphelin, a new anti-oxidative N-cinnamoyl tripeptide produced by the deep sea-derived fungal strain Penicillium brevicompactum FKJ-0123.

A new N-cinnamoyl tripeptide, designated cipralphelin (1), was isolated from a cultured broth of Penicillium brevicompactum FKJ-0123 by physicochemical (PC) screening. Compound 1 was purified by silica gel and ODS column chromatography followed by preparative HPLC. The structure of 1 was determined as N-cinnamoyl-prolyl-alanyl-phenylalanine methyl ester by nuclear magnetic resonance and mass spectrometry analyses. The absolute configurations of three amino acids were determined by an advanced Marfey's method applied to the hydrolysate of 1. Compound 1 was evaluated for its cytotoxicity, anti-microbial activity, and ability to scavenge or quench reactive oxygen species (ROS) such as superoxide anion radicals, hydroxy radicals, and singlet oxygen. Compound 1 exhibited potent scavenging activity against hydroxy radicals.