Akkermansia muciniphila phospholipid induces homeostatic immune responses.

Multiple studies have established associations between human gut bacteria and host physiology, but determining the molecular mechanisms underlying these associations has been challenging1-3. Akkermansia muciniphila has been robustly associated with positive systemic effects on host metabolism, favourable outcomes to checkpoint blockade in cancer immunotherapy and homeostatic immunity4-7. Here we report the identification of a lipid from A. muciniphila's cell membrane that recapitulates the immunomodulatory activity of A. muciniphila in cell-based assays8. The isolated immunogen, a diacyl phosphatidylethanolamine with two branched chains (a15:0-i15:0 PE), was characterized through both spectroscopic analysis and chemical synthesis. The immunogenic activity of a15:0-i15:0 PE has a highly restricted structure-activity relationship, and its immune signalling requires an unexpected toll-like receptor TLR2-TLR1 heterodimer9,10. Certain features of the phospholipid's activity are worth noting: it is significantly less potent than known natural and synthetic TLR2 agonists; it preferentially induces some inflammatory cytokines but not others; and, at low doses (1% of EC50) it resets activation thresholds and responses for immune signalling. Identifying both the molecule and an equipotent synthetic analogue, its non-canonical TLR2-TLR1 signalling pathway, its immunomodulatory selectivity and its low-dose immunoregulatory effects provide a molecular mechanism for a model of A. muciniphila's ability to set immunological tone and its varied roles in health and disease.

Collinsella aerofaciens Produces a pH-Responsive Lipid Immunogen.

Some members of the human gut microbiota profoundly influence their host's physiology, health, and therapeutic responses, but the responsible molecules and mechanisms are largely unknown. As part of a project to identify immunomodulators produced by gut microbes, we analyzed the metabolome of Collinsella aerofaciens, an actinomycete that figures prominently in numerous association studies. The associations are typically positive correlations of C. aerofaciens with pro-inflammatory responses and undesirable outcomes, but an association with favorable responses to PD-1/PD-L1 cancer immunotherapy is a notable exception. A phenotypic assay-guided screen using dendritic cells (mBMDCs) and cytokine readouts identified the active compound, which was structurally characterized as a lysoglycoglycerolipid with an acetal-bearing ß-galactofuranose head group (CaLGL-1, 1). The structural assignment was confirmed through total synthesis. Assays with tlr2-/-, tlr4-/-, and wt mBMDCs revealed TLR2-dependent signaling. CaLGL-1 is produced by a conversion of a bacterially biosynthesized plasmalogen (CaPlsM, 3) to CaLGL-1 (1) in a low-pH environment.

Synthesis and Structure Revision of Naturally Occurring Homoisoflavane (+)-Dracaeconolide B.

Dracaeconolide B (1), a naturally occurring homoisoflavane, was isolated from the red resin of Dracaena cochinchinensis. Efforts have been made to elucidate the exact structure of compound 1 since it was confirmed that dracaeconolide B did not contain a 7-hydroxy-5,8-dimethoxy moiety. The structure of dracaeconolide B was revised by synthesis of three homoisoflavanes containing a 5,6,7-trioxygenated moiety each and analysis by NMR spectroscopy. The revised structure of dracaeconolide B was proposed as 3-(4-hydroxybenzyl)-7-hydroxy-5,6-dimethoxychromane. Noyori's Ru-catalyzed asymmetric transfer hydrogenation was used to synthesize (+)-dracaeconolide B. The absolute configuration of the compound was revised to S based on the results obtained by the electronic circular dichroism calculation. We examined the antiangiogenic activity of (S)- and (R)-dracaeconolide B and of synthetic 5,6,7- and 5,7,8-trioxygenated homoisoflavanes. The results can potentially help in the synthesis of related natural products and support drug discovery to treat neovascular eye diseases.

Lyme Disease, Borrelia burgdorferi, and Lipid Immunogens.

The human immune system detects potentially pathogenic microbes with receptors that respond to microbial metabolites. While the overall immune signaling pathway is known in considerable detail, the initial molecular signals, the microbially produced immunogens, for important diseases like Lyme disease (LD) are often not well-defined. The immunogens for LD are produced by the spirochete Borrelia burgdorferi, and a galactoglycerolipid (1) has been identified as a key trigger for the inflammatory immune response that characterizes LD. This report corrects the original structural assignment of 1 to 3, a change of an α-galactopyranose to an α-galactofuranose headgroup. The seemingly small change has important implications for the diagnosis, prevention, and treatment of LD.

Bacterial diketopiperazines stimulate diatom growth and lipid accumulation.

Diatoms are photosynthetic microalgae that fix a significant fraction of the world's carbon. Because of their photosynthetic efficiency and high-lipid content, diatoms are priority candidates for biofuel production. Here, we report that sporulating Bacillus thuringiensis and other members of the Bacillus cereus group, when in co-culture with the marine diatom Phaeodactylum tricornutum, significantly increase diatom cell count. Bioassay-guided purification of the mother cell lysate of B. thuringiensis led to the identification of two diketopiperazines (DKPs) that stimulate both P. tricornutum growth and increase its lipid content. These findings may be exploited to enhance P. tricornutum growth and microalgae-based biofuel production. As increasing numbers of DKPs are isolated from marine microbes, the work gives potential clues to bacterial-produced growth factors for marine microalgae.

Actinoflavosides B-D, Flavonoid Type Glycosides from Tidal Mudflat-Derived Actinomyces.

Three new secondary metabolites, actinoflavosides B-D (1-3), were discovered in the culture broth of two actinomycete strains (JML48 and JMS33) that were isolated from tidal mudflat sediment in Muan, Republic of Korea. The planar structures of the actinoflavosides were elucidated by MS, UV, and NMR analyses. The stereochemistry of an aminosugar, 2,3,6-trideoxy-3-amino-ribopyranoside in the actinoflavosides was determined by J-based configuration analysis using values obtained from DQF-COSY experiments and modified Mosher's method. Actinoflavosides B-D (1-3) displayed antibacterial activity against Pseudomonas aeruginosa, and actinoflavoside D (3) significantly increased IL-2 production in mouse splenocytes.

Taeanamides A and B, Nonribosomal Lipo-Decapeptides Isolated from an Intertidal-Mudflat-Derived Streptomyces sp.

Two new lipo-decapeptides, namely taeanamides A and B (1 and 2), were discovered from the Gram-positive bacterium Streptomyces sp. AMD43, which was isolated from a mudflat sample from Anmyeondo, Korea. The exact molecular masses of 1 and 2 were revealed by high-resolution mass spectrometry, and the planar structures of 1 and 2 were elucidated using NMR spectroscopy. The absolute configurations of 1 and 2 were determined using a combined analysis of 1H-1H coupling constants and ROESY correlations, the advanced Marfey's method, and bioinformatics. The putative nonribosomal peptide synthetase pathway for the taeanamides was identified by analyzing the full genome sequence data of Streptomyces sp. AMD43. We also found that taeanamide A exhibited mild anti-tuberculosis bioactivity, whereas taeanamide B showed significant bioactivity against several cancer cell lines.

Targeted Degradation of the Oncogenic Phosphatase SHP2.

SHP2 is a protein tyrosine phosphatase that plays a critical role in the full activation of the Ras-MAPK pathway upon stimulation of receptor tyrosine kinases, which are frequently amplified or mutationally activated in human cancer. In addition, activating mutations in SHP2 result in developmental disorders and hematologic malignancies. Several allosteric inhibitors have been developed for SHP2 and are currently in clinical trials. Here, we report the development and evaluation of a SHP2 PROTAC created by conjugating RMC-4550 with pomalidomide using a PEG linker. This molecule is highly selective for SHP2, induces degradation of SHP2 in leukemic cells at submicromolar concentrations, inhibits MAPK signaling, and suppresses cancer cell growth. SHP2 PROTACs serve as an alternative strategy for targeting ERK-dependent cancers and are useful tools alongside allosteric inhibitors for dissecting the mechanisms by which SHP2 exerts its oncogenic activity.

Unprecedented Noncanonical Features of the Nonlinear Nonribosomal Peptide Synthetase Assembly Line for WS9326A Biosynthesis.

Systematic inactivation of nonribosomal peptide synthetase (NRPS) domains and translocation of the thioesterase (TE) domain revealed several unprecedented nonlinear NRPS assembly processes during the biosynthesis of the cyclodepsipeptide WS9326A in Streptomyces sp. SNM55. First, two sets of type ΙΙ TE (TEΙΙ)-like enzymes mediate the shuttling of activated amino acids between two sets of stand-alone adenylation (A)-thiolation (T) didomain modules and an "A-less" condensation (C)-T module with distinctive specificities and flexibilities. This was confirmed by the elucidation of the affinities of the A-T didomains for the TEΙΙs and its structure. Second, the C-T didomain module operates iteratively and independently from other modules in the same protein to catalyze two chain elongation cycles. Third, this biosynthetic pathway includes the first example of module skipping, where the interpolated C and T domains are required for chain transfer.

Donghaecyclinones A-C: New Cytotoxic Rearranged Angucyclinones from a Volcanic Island-Derived Marine Streptomyces sp.

Chemical profiling of the Streptomyces sp. strain SUD119, which was isolated from a marine sediment sample collected from a volcanic island in Korea, led to the discovery of three new metabolites: donghaecyclinones A-C (1-3). The structures of 1-3 were found to be rearranged, multicyclic, angucyclinone-class compounds according to nuclear magnetic resonance (NMR) and mass spectrometry (MS) analyses. The configurations of their stereogenic centers were successfully assigned using a combination of quantum mechanics-based computational methods for calculating the NMR shielding tensor (DP4 and CP3) as well as electronic circular dichroism (ECD) along with a modified version of Mosher's method. Donghaecyclinones A-C (1-3) displayed cytotoxicity against diverse human cancer cell lines (IC50: 6.7-9.6 µM for 3).

Rhizolutin, a Novel 7/10/6-Tricyclic Dilactone, Dissociates Misfolded Protein Aggregates and Reduces Apoptosis/Inflammation Associated with Alzheimer's Disease.

Rhizolutin (1) was discovered as a natural product of ginseng-rhizospheric Streptomyces sp. WON17. Its structure features an unprecedented 7/10/6-tricyclic dilactone carbon skeleton composed of dimethylcyclodecatriene flanked by a 7-membered and a 6-membered lactone ring based on spectroscopic analysis. During an unbiased screening of natural product libraries, this novel compound was found to dissociate amyloid-ß (Aß) plaques and tau tangles, which are key pathological hallmarks of Alzheimer's disease (AD). Rhizolutin treatment of APP/PS1 double transgenic mice with AD significantly dissociated hippocampal plaques. In vitro, rhizolutin substantially decreased Aß-induced apoptosis and inflammation in neuronal and glial cells. Our findings introduce a unique chemical entity that targets Aß and tau concurrently by mimicking misfolded protein clearance mechanisms of immunotherapy, which is prominently investigated in clinical trials.

Depsidomycins B and C: New Cyclic Peptides from a Ginseng Farm Soil-Derived Actinomycete.

LC/MS-based chemical profiling of a ginseng farm soil-derived actinomycete strain, Streptomyces sp. BYK1371, enabled the discovery of two new cyclic heptapeptides, depsidomycins B and C (1 and 2), each containing two piperazic acid units and a formyl group at their N-terminus. The structures of 1 and 2 were elucidated by a combination of spectroscopic and chemical analyses. These new compounds were determined to possess d-leucine, d-threonine, d-valine, and S-piperazic acid based on the advanced Marfey's method and a GITC (2,3,4,6-tetra-O-acetyl-ß-d-glucopyranosyl isothiocyanate) derivatization of their hydrolysates, followed by LC/MS analysis. Depsidomycins B and C displayed significant antimetastatic activities against metastatic breast cancer cells (MDA-MB-231).

Structural Revision of Baulamycin A and Structure-Activity Relationships of Baulamycin A Derivatives.

Total synthesis of the proposed structure of baulamycin A was performed. The spectral properties of the synthetic compound differ from those reported for the natural product. On the basis of comprehensive NMR study, we proposed two other possible structures for natural baulamycin A. Total syntheses of these two substances were performed, which enabled assignment of the correct structure of baulamycin A. Key features of the convergent and fully stereocontrolled route include Evans Aldol and Brown allylation reactions to construct the left fragment, a prolinol amide-derived alkylation/desymmetrization to install the methyl-substituted centers in the right fragment, and finally, a Carreira alkynylation to join both fragments. In addition, we have determined the inhibitory activities of novel baulamycin A derivatives against the enzyme SbnE. This SAR study provides useful insight into the design of novel SbnE inhibitors that overcome the drug resistance of pathogens, which cause life-threatening infections.

Anti-Neurodegenerative Biflavonoid Glycosides from Impatiens balsamina.

Four biflavonoid glycosides, balsamisides A-D (1-4), and nine known compounds (5-13) were obtained from the white petals of Impatiens balsamina. The 2D structures of the purified phytochemicals were established using conventional NMR techniques in addition to the new long-range HSQMBC NMR experiment. Acid hydrolysis followed by experimental and quantum-mechanics-based ECD data analysis permitted full configurational assignment of the purified metabolites. Compounds 1-13 were assessed for their potential to impede the generation of nitric oxide in lipopolysaccharide-stimulated BV2 cells. They were also investigated for potential neuroprotective activity using C6 cells and cytotoxicity against some human tumor cell lines, but were inactive (IC50 > 10 µM) against all the cell lines.

QM-HiFSA-Aided Structure Determination of Succinilenes A-D, New Triene Polyols from a Marine-Derived Streptomyces sp.

Based on profiles of secondary metabolites produced by marine bacteria obtained using LC/MS, succinilenes A-D (1-4), new triene polyols, were discovered from a culture of a Streptomyces strain SAK1, which was collected in the southern area of Jeju Island, Republic of Korea. The gross structures of 1-4 were primarily determined through analysis of NMR spectra. The double bond geometries of the succinilenes, which could not be established from conventional ¹H NMR spectra because of the highly overlapped olefinic signals, were successfully deciphered using the recently developed quantum-mechanics-driven ¹H iterative full spin analysis (QM-HiFSA). Succinilenes A-C (1-3) displayed inhibitory effects against lipopolysaccharide (LPS)-induced nitric oxide (NO) production, indicating their anti-inflammatory significance. These three compounds (1-3) commonly bear a succinic acid moiety, although succinilene D (4), which did not inhibit NO production, does not have this moiety in its structure. The absolute configurations of succinilenes A-D (1-4) were established through J-based configuration analysis, the modified Mosher's method following methanolysis, and CD spectral analysis.

Mohangic Acids A-E, p-Aminoacetophenonic Acids from a Marine-Mudflat-Derived Streptomyces sp.

Mohangic acids A-E (1-5) were isolated from a marine Streptomyces sp. collected from a mudflat in Buan, Republic of Korea. Comprehensive spectroscopic analysis revealed that the mohangic acids are new members of the p-aminoacetophenonic acid class. The relative and absolute configurations of the mohangic acids were determined by J-based configuration analysis and by the application of bidentate chiral NMR solvents followed by (13)C NMR analysis, chemical derivatization, and circular dichroism spectroscopy. Mohangic acid E (5), which is the first glycosylated compound in the p-aminoacetophenonic acid family, displayed significant quinone reductase induction activity.

Hormaomycins B and C: New Antibiotic Cyclic Depsipeptides from a Marine Mudflat-Derived Streptomyces sp.

Alterations in microbial culture conditions may trigger the production of diverse bioactive secondary metabolites. While applying various culture conditions and monitoring secondary metabolite profiles using LC/MS, hormaomycins B and C (1 and 2) were discovered from a marine mudflat-derived actinomycete, Streptomyces sp., collected in Mohang, Korea. The planar structures of the hormaomycins, which bear structurally-unique units, such as 4-(Z)-propenylproline, 3-(2-nitrocyclopropyl)alanine, 5-chloro-1-hydroxypyrrol-2-carboxylic acid and b-methylphenylalanine, were established as the first natural analogues belonging to the hormaomycin peptide class. The absolute configurations of 1 and 2 were deduced by comparing their CD spectra with that of hormaomycin. These hormaomycins exhibited significant inhibitory effects against various pathogenic Gram-positive and Gram-negative bacteria.

A new benzofuran glycoside and indole alkaloids from a sponge-associated rare actinomycete, Amycolatopsis sp.

Three new secondary metabolites, amycofuran (1), amycocyclopiazonic acid (2), and amycolactam (3), were isolated from the sponge-associated rare actinomycete Amycolatopsis sp. Based on combined spectroscopic analyses, the structures of 1-3 were determined to be a new benzofuran glycoside and new indole alkaloids related to cyclopiazonic acids, a class that has previously only been reported in fungi. The absolute configurations of 1 and 3 were deduced by ECD calculations, whereas that of 2 was determined using the modified Mosher method. Amycolactam (3) displayed significant cytotoxicity against the gastric cancer cell line SNU638 and the colon cancer cell line HCT116.

Separacenes A-D, novel polyene polyols from the marine actinomycete, Streptomyces sp.

Separacenes A-D (1-4), novel polyene polyols, were isolated from Streptomyces sp. collected from the southern area of Jeju Island, Korea. The chemical structures of 1-4 were established by NMR, mass, UV, and IR spectroscopy as well as the modified Mosher's method. Separacenes A-B (1-2), which share an identical planar structure but possess different relative configurations, bear tetraene units flanked by two diol moieties, whereas the stereoisomeric separacenes C-D (3-4) possess a triene moiety between two diol substructures. Separacenes A-D each contain a terminal olefinic methylene. Separacene A displayed inhibitory activity against Candida albicans isocitrate lyase and weak cytotoxicity against both the colon carcinoma cell line HCT-116 and the lung cancer cell line A549.

Asymmetric Synthesis of Four Stereoisomers of 2,2-Dimethyl-3-hydroxy-4-(1'-angeloyloxy)-6-acetylchromane from Ageratina grandifolia and Plausible Absolute Stereochemistry of the Natural Product.

2,2-Dimethyl-3-hydroxy-4-(1'-angeloyloxy)-6-acetylchromane is a natural product isolated from Ageratina grandifolia that exhibits inhibitory activity against yeast α-glucosidase. Initially, its structure was proposed to be 4-hydroxy-3-((S)-1'-angeloyloxy-(R)-2',3'-epoxy-3'-methyl)butylacetophenone with an epoxide, but the structure was later revised to 2,2-dimethyl-3R-hydroxy-4S-(1-angeloyloxy)-6-acetylchromane. In this study, we present a total synthesis of 2,2-dimethyl-3-hydroxy-4-(1'-angeloyloxy)-6-acetylchromane from A. gradifolia and its stereoisomers. The key features of their synthesis include Sharpless asymmetric dihydroxylation of a readily available benzopyran substrate and subsequent Mitsunobu or Steglich reaction to provide both cis- and trans-isomers with chiral control. The absolute stereochemistry of the natural product was determined to be 2,2-dimethyl-3S-hydroxy-4R-(1'-angeloyloxy)-6-acetylchromane based on optical rotations of the synthesized compounds. The absolute configuration of the synthesized stereoisomers was confirmed by Mosher ester analysis. In addition, we provided ECD spectra for the four stereoisomers, which will allow verification of the absolute configuration of the natural product. Synthesis of all four stereoisomers of 2,2-dimethyl-3-hydroxy-4-(1'-angeloyloxy)-6-acetylchromane would facilitate the exploration of their potential biomedical applications.