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Tayler, NM, Boya CA, Herrera L, Moy J, Ng M, Pineda L, Almanza A, Rosero S, Coronado LM, Correa R, Santamaria R, Caballero Z, Durant-Archibold AA, Currency KJT, Balunas MJ, Gerwick WH, Spadafora A, Gutierrez M, SpadaforaId C.  2019.  Analysis of the antiparasitic and anticancer activity of the coconut palm (Cocos nucifera L. ARECACEAE) from the natural reserve of Punta Patino, Darien. Plos One. 14   10.1371/journal.pone.0214193   AbstractWebsite

Cocos nucifera (C. nucifera) (the coconut palm tree) has been traditionally used to fight a number of human diseases, but only a few studies have tested its components against parasites such as those that cause malaria. In this study, C. nucifera samples were collected from a private natural reserve in Punta Patino, Darien, Panama. The husk, leaves, pulp, and milk of C. nucifera were extracted and evaluated against the parasites that cause Chagas' disease or American trypanosomiasis (Trypanosoma cruzi), leishmaniasis (Leishmania donovani) and malaria (Plasmodium falciparum), as well as against a line of breast cancer cells. While there was no activity in the rest of the tests, five and fifteen-minute aqueous decoctions of leaves showed antiplasmodial activity at 10% v/v concentration. Removal of some HPLC fractions resulted in loss of activity, pointing to the presence of synergy between the components of the decoction. Chemical molecules were separated and identified using an ultra-performance liquid chromatography (UPLC) approach coupled to tandem mass spectrometry (LC-MS/MS) using atmospheric pressure chemical ionization quadrupole-time of flight mass spectrometry (APCI-Q-TOF-MS) and molecular networking analysis, revealing the presence of compounds including polyphenol, flavone, sterol, fatty acid and chlorophyll families, among others.

Hou, XM, Wang CY, Gerwick WH, Shao CL.  2019.  Marine natural products as potential anti-tubercular agents. European Journal of Medicinal Chemistry. 165:273-292.   10.1016/j.ejmech.2019.01.026   AbstractWebsite

Tuberculosis has been one of the greatest global health challenges of all time. Although the current first-line anti-tuberculosis (anti-TB) medicines used in the clinic have reduced mortality, multidrug-resistance and extensively drug-resistance forms of the disease have now spread worldwide and become a global problem. Even so, few new clinically approved drugs have emerged during the past 30 years. Highly biodiverse marine organisms have received considerable attention for drug discovery in the past couple of decades, and emerging TB drug resistance has motivated interest in assessing marine natural products (MNPs) in the treatment of this disease. So far, more than 170 compounds have been isolated from marine organisms with anti-TB properties, ten of which exhibit potent activity and have the potential for further development. This review systematically surveys MNPs with anti-TB activity and illustrates the impact of these compounds on drug discovery research against tuberculosis. (C) 2019 Elsevier Masson SAS. All rights reserved.

Caro-Diaz, EJE, Valeriote FA, Gerwick WH.  2019.  Highly convergent total synthesis and assignment of absolute configuration of majusculamide D, a potent and selective cytotoxic metabolite from Moorea sp. Organic Letters. 21:793-796.   10.1021/acs.orglett.8b04050   AbstractWebsite

The total synthesis of majusculamide D (MJS-D) is described, a lipopentapeptide originally isolated from Lyngbya majuscula and reisolated from a Moorea sp. MJS-D possesses selective and potent cancer cell toxicity. A scalable and convergent strategy with a minimal number of purifications produced significant quantities of MJM-D for in vivo evaluations. The absolute configuration of the 1,3-dimethyl-octanamide motif was determined by synthesis of this fragment via ZACA chemistry.

Chen, HM, Yang R, Chen JJ, Luo QJ, Cui XS, Yan XJ, Gerwick WH.  2019.  1-Octen-3-ol, a self-stimulating oxylipin messenger, can prime and induce defense of marine alga. Bmc Plant Biology. 19   10.1186/s12870-019-1642-0   AbstractWebsite

BackgroundShort chain oxylipins in plants as the main volatile organic carbon have been speculated to playing an important role for plant innate immunity, however, not yet intensively studied and far away established as the fully recognized algae defense signals.ResultsThe production of 1-octen-3-ol is self-amplified via the fatty acid-oxylipin metabolic cycle through positive feedback loop. Production of 1-octen-3-ol may act as a messenger that induces P. haitanensis to be in a primed state and ready for defense by upregulating the synthesis of methyl jasmonic acid, indole-3-acetic acid, and gibberellin A3. Production of these oxylipins also adjust the redox state in cells, resulting in host defense activation.ConclusionsWe provide the first demonstration that 1-octen-3-ol from P. haitanensis, can act as a self-stimulating community messenger. The multiple effects of 1-octen-3-ol may explain why P. haitanensis, a very ancient lineage within plant kingdom, thrives in the niche of intertidal zones.

Choi, H, Engene N, Byrum T, Wang SH, Oh DC, Gerwick WH.  2019.  Dragocins A-D, structurally intriguing cytotoxic metabolites from a Panamanian marine cyanobacterium. Organic Letters. 21:266-270.   10.1021/acs.orglett.8b03712   AbstractWebsite

Dragocins A-D (1-4) were isolated from a dark-red wooly textured marine cyanobacterium collected in Boca del Drago, Panama. Dragocins A-C (1-3) possessed 2,3-dihydroxypyrrolidine, 1-hydroxy-5-O-Me-benzoyl, and 4'-substituted-beta-ribofuranose moieties that connected to form a nine-membered macrocyclic ring. Dragocins A-C are members of a unique hybrid structural class with substitution at the C-4' position of a ribofuranose unit. Of the four new compounds, dragocin A was the most potent cytotoxin to human H-460 lung cancer cells

Almaliti, J, Miller B, Pietraszkiewicz H, Glukhov E, Naman CB, Kline T, Hanson J, Li XF, Zhou SH, Valeriote FA, Gerwick WH.  2019.  Exploration of the carmaphycins as payloads in antibody drug conjugate anticancer agents. European Journal of Medicinal Chemistry. 161:416-432.   10.1016/j.ejmech.2018.10.024   AbstractWebsite

Antibody-drug conjugates (ADCs) represent a new dimension of anticancer chemotherapeutics, with warheads to date generally involving either antitubulin or DNA-directed agents to achieve low-to sub-nanomolar potency. However, other potent cytotoxins working by different pharmacological mechanisms are under investigation, such as alpha,beta-epoxyketone based proteasome inhibitors. These proteasome active agents are an emerging class of anticancer drug that possesses ultra-potent cytotoxicity to some cancer cell lines. The carmaphycins are representatives of this latter class that we isolated and characterized from a marine cyanobacterium, and these as well as several synthetic analogues exhibit this level of potency. In the current work, we investigated the use of these highly potent cytotoxic compounds as warheads in the design of novel ADCs. We designed and synthesized a library of carmaphycin B analogues that contain amine handles, enabling their attachment to an antibody linker. The basicity of these incorporated amine handles was shown to strongly affect their cytotoxic properties. Linear amines resulted in the greatest reduction in cytotoxicity whereas less basic aromatic amines retained potent activity as demonstrated by a 4-sulfonylaniline derivative. These investigations resulted in identifying the P2 residue in the carmaphycins as the most suitable site for linker attachment point, and hence, we synthesized a highly potent analogue of carmaphycin B that contained a 4-sulfonylaniline handle as an attachment point for the linker antibody. (C) 2018 Elsevier Masson SAS. All rights reserved.

Ong, JFM, Goh HC, Lim SC, Pang LM, Chin JSF, Tan KS, Liang ZX, Yang L, Glukhov E, Gerwick WH, Tan LT.  2019.  Integrated genomic and metabolomic approach to the discovery of potential anti-quorum sensing natural products from microbes associated with marine samples from Singapore. Marine Drugs. 17   10.3390/md17010072   AbstractWebsite

With 70% of the Earth's surface covered in water, the marine ecosystem offers immense opportunities for drug discovery and development. Due to the decreasing rate of novel natural product discovery from terrestrial sources in recent years, many researchers are beginning to look seaward for breakthroughs in new therapeutic agents. As part of an ongoing marine drug discovery programme in Singapore, an integrated approach of combining metabolomic and genomic techniques were initiated for uncovering novel anti-quorum sensing molecules from bacteria associated with subtidal samples collected in the Singapore Strait. Based on the culture-dependent method, a total of 102 marine bacteria strains were isolated and the identities of selected strains were established based on their 16S rRNA gene sequences. About 5% of the marine bacterial organic extracts showed quorum sensing inhibitory (QSI) activity in a dose-dependent manner based on the Pseudomonas aeruginosa QS reporter system. In addition, the extracts were subjected to mass spectrometry-based molecular networking and the genome of selected strains were analysed for known as well as new biosynthetic gene clusters. This study revealed that using integrated techniques, coupled with biological assays, can provide an effective and rapid prioritization of marine bacterial strains for downstream large-scale culturing for the purpose of isolation and structural elucidation of novel bioactive compounds.

McAlpine, JB, Chen SN, Kutateladze A, MacMillan JB, Appendino G, Barison A, Beniddir MA, Biavatti MW, Bluml S, Boufridi A, Butler MS, Capon RJ, Choi YH, Coppage D, Crews P, Crimmins MT, Csete M, Dewapriya P, Egan JM, Garson MJ, Genta-Jouve G, Gerwick WH, Gross H, Harper MK, Hermanto P, Hook JM, Hunter L, Jeannerat D, Ji NY, Johnson TA, Kingston DGI, Koshino H, Lee HW, Lewin G, Li J, Linington RG, Liu MM, McPhail KL, Molinski TF, Moore BS, Nam JW, Neupane RP, Niemitz M, Nuzillard JM, Oberlies NH, Ocampos FMM, Pan G, Quinn RJ, Reddy DS, Renault JH, Rivera-Chavez J, Robien W, Saunders CM, Schmidt TJ, Seger C, Shen B, Steinbeck C, Stuppner H, Sturm S, Taglialatela-Scafati O, Tantillo DJ, Verpoorte R, Wang BG, Williams CM, Williams PG, Wist J, Yue JM, Zhang C, Xu ZR, Simmler C, Lankin DC, Bisson J, Pauli GF.  2019.  The value of universally available raw NMR data for transparency, reproducibility, and integrity in natural product research. Natural Product Reports. 36:35-107.   10.1039/c7np00064b   AbstractWebsite

With contributions from the global natural product (NP) research community, and continuing the Raw Data Initiative, this review collects a comprehensive demonstration of the immense scientific value of disseminating raw nuclear magnetic resonance (NMR) data, independently of, and in parallel with, classical publishing outlets. A comprehensive compilation of historic to present-day cases as well as contemporary and future applications show that addressing the urgent need for a repository of publicly accessible raw NMR data has the potential to transform natural products (NPs) and associated fields of chemical and biomedical research. The call for advancing open sharing mechanisms for raw data is intended to enhance the transparency of experimental protocols, augment the reproducibility of reported outcomes, including biological studies, become a regular component of responsible research, and thereby enrich the integrity of NP research and related fields.

Moss, NA, Leao T, Rankin MR, McCullough TM, Qu PP, Korobeynikov A, Smith JL, Gerwick L, Gerwick WH.  2018.  Ketoreductase domain dysfunction expands chemodiversity: Malyngamide biosynthesis in the cyanobacterium Okeania hirsuta. Acs Chemical Biology. 13:3385-3395.   10.1021/acschembio.8b00910   AbstractWebsite

Dozens of type A malyngamides, principally identified by a decorated six-membered cyclohexanone head group and methoxylated lyngbic acid tail, have been isolated over several decades. Their environmental sources include macro- and microbiotic organisms, including sea hares, red alga, and cyanobacterial assemblages, but the true producing organism has remained enigmatic. Many type A analogues display potent bioactivity in human-health related assays, spurring an interest in this molecular class and its biosynthetic pathway. Here, we present the discovery of the type A malyngamide biosynthetic pathway in the first sequenced genome of the cyanobacterial genus Okeania. Bioinformatic analysis of two cultured Okeania genome assemblies identified 62 and 68 kb polyketide synthase/nonribosomal peptide synthetase (PKS/NRPS) pathways with unusual loading and termination genes. NMR data of malyngamide C acetate derived from C-13-substrate-fed cultures provided evidence that an intact octanoate moiety is transferred to the first KS module via a LipM homologue originally associated with lipoic acid metabolism and implicated an inactive ketoreductase (KR0) as critical for six-membered ring formation, a hallmark of the malyngamide family. Phylogenetic analysis and homology modeling of the penultimate KR0 domain inferred structural cofactor binding and active site alterations as contributory to domain dysfunction, which was confirmed by recombinant protein expression and NADPH binding assay. The carbonyl retained from this KR0 ultimately enables an intramolecular Knoevenagel condensation to form the characteristic cyclohexanone ring. Understanding this critical step allows assignment of a biosynthetic model for all type A malyngamides, whereby well-characterized tailoring modifications explain the surprising proliferation and diversity of analogues.

Ding, CYG, Pang LM, Liang ZX, Goh KKK, Glukhov E, Gerwick WH, Tan LT.  2018.  MS/MS-based molecular networking approach for the detection of aplysiatoxin-related compounds in environmental marine cyanobacteria. Marine Drugs. 16   10.3390/md16120505   AbstractWebsite

Certain strains of cyanobacteria produce a wide array of cyanotoxins, such as microcystins, lyngbyatoxins and aplysiatoxins, that are associated with public health issues. In this pilot study, an approach combining LC-MS/MS and molecular networking was employed as a rapid analytical method to detect aplysiatoxins present in four environmental marine cyanobacterial samples collected from intertidal areas in Singapore. Based on 16S-ITS rRNA gene sequences, these filamentous cyanobacterial samples collected from Pulau Hantu were determined as Trichodesmium erythraeum, Oscillatoria sp. PAB-2 and Okeania sp. PNG05-4. Organic extracts were prepared and analyzed on LC-HRMS/MS and Global Natural Product Social Molecular Networking (GNPS) for the presence of aplysiatoxin-related molecules. From the molecular networking, six known compounds, debromoaplysiatoxin (1), anhydrodebromoaplysiatoxin (2), 3-methoxydebromoaplysiatoxin (3), aplysiatoxin (4), oscillatoxin A (5) and 31-noroscillatoxin B (6), as well as potential new analogues, were detected in these samples. In addition, differences and similarities in molecular networking clusters related to the aplysiatoxin molecular family were observed in extracts of Trichodesmium erythraeum collected from two different locations and from different cyanobacterial species found at Pulau Hantu, respectively.

Skiba, MA, Bivins MM, Schultz JR, Bernard SM, Fiers WD, Dan QY, Kulkarni S, Wipf P, Gerwick WH, Sherman DH, Aldrich CC, Smith JL.  2018.  Structural basis of polyketide synthase o-methylation. Acs Chemical Biology. 13:3221-3228.   10.1021/acschembio.8b00687   AbstractWebsite

Modular type I polyketide synthases (PKSs) produce some of the most chemically complex metabolites in nature through a series of multienzyme modules. Each module contains a variety of catalytic domains to selectively tailor the growing molecule. PKS O-methyltransferases (O-MTs) are predicted to methylate beta-hydroxyl or beta-keto groups, but their activity and structure have not been reported. We determined the domain boundaries and characterized the catalytic activity and structure of the StiD and StiE O-MTs, which methylate opposite beta-hydroxyl stereocenters in the myxobacterial stigmatellin biosynthetic pathway. Substrate stereospecificity was demonstrated for the StiD O-MT. Key catalytic residues were identified in the crystal structures and investigated in StiE O-MT via site-directed mutagenesis and further validated with the cyanobacterial CurL O-MT from the curacin biosynthetic pathway. Initial structural and biochemical analysis of PKS O-MTs supplies a new chemoenzymatic tool, with the unique ability to selectively modify hydroxyl groups during polyketide biosynthesis .

Via, CW, Glukhov E, Costa S, Zimba PV, Moeller PDR, Gerwick WH, Bertin MJ.  2018.  The metabolome of a cyanobacterial bloom visualized by MS/MS-based molecular networking reveals new neurotoxic smenamide analogs (C, D, and E). Frontiers in Chemistry. 6   10.3389/fchem.2018.00316   AbstractWebsite

Members of the cyanobacterial genus Trichodesmium are well known for their substantial impact on nitrogen influx in ocean ecosystems and the enormous surface blooms they form in tropical and subtropical locations. However, the secondary metabolite composition of these complex environmental bloom events is not well known, nor the possibility of the production of potent toxins that have been observed in other bloom-forming marine and freshwater cyanobacteria species. In the present work, we aimed to characterize the metabolome of a Trichodesmium bloom utilizing MS/MS-based molecular networking. Furthermore, we integrated cytotoxicity assays in order to identify and ultimately isolate potential cyanotoxins from the bloom. These efforts led to the isolation and identification of several members of the smenamide family, including three new smenamide analogs (1-3) as well as the previously reported smenothiazole A-hybrid polyketide-peptide compounds. Two of these new smenamides possessed cytotoxicity to neuro-2A cells (1 and 3) and their presence elicits further questions as to their potential ecological roles. HPLC profiling and molecular networking of chromatography fractions from the bloom revealed an elaborate secondary metabolome, generating hypotheses with respect to the environmental role of these metabolites and the consistency of this chemical composition across genera, space and time.

Huang, QL, Chen J, Zhang WJ, Zhou BP, Zhang CL, Gerwick WH, Cao ZY.  2018.  Alkaloids from Corydalis decumbens suppress neuronal excitability in primary cultures of mouse neocortical neurons. Phytochemistry. 150:85-92.   10.1016/j.phytochem.2018.03.006   AbstractWebsite

Eight previously undescribed alkaloids, named corydemine, dihydrocorydemine, corydedine, 8,13-dioxo-14-hydroxytetrahydropalmatine, egenine-alpha-N-oxide, egenine-beta-N-oxide, 7'-O-ethylegenine-alpha-N-oxide, and 7'-O-ethylegenine-beta-N-oxide, together with three known ones, muramine, L-tetrahydropalmatine, and (+)-egenine, were isolated from the bulbs of Corydalis decumbens. Their structures were elucidated by comprehensive spectroscopic analysis and chemical correlation. The isolated compounds were tested for their ability to modulate neuronal excitability in primary cultured neocortical neurons. Four of the compounds, corydemine, dihydrocorydemine, muramine, and L-tetrahydropalmatine, inhibited neuronal excitability with IC50 values of 3.6, 16.7, 13.5 and 14.0 mu M, respectively. (C) 2018 Elsevier Ltd. All rights reserved.

Skiba, MA, Sikkema AP, Moss NA, Lowell AN, Su M, Sturgis RM, Gerwick L, Gerwick WH, Sherman DH, Smith JL.  2018.  Biosynthesis of t-Butyl in apratoxin A: functional analysis and architecture of a PKS loading module. Acs Chemical Biology. 13:1640-1650.   10.1021/acschembio.8b00252   AbstractWebsite

The unusual feature of a t-butyl group is found in several marine-derived natural products including apratoxin A, a Sec61 inhibitor produced by the cyanobacterium Moorea bouillonii PNG 5-198. Here, we determine that the apratoxin A t-butyl group is formed as a pivaloyl acyl carrier protein (ACP) by AprA, the polyketide synthase (PKS) loading module of the apratoxin A biosynthetic pathway. AprA contains an inactive "pseudo" GCN5-related N-acetyltransferase domain (Psi GNAT) flanked by two methyltransferase domains (MT1 and MT2) that differ distinctly in sequence. Structural, biochemical, and precursor incorporation studies reveal that MT2 catalyzes unusually coupled decarboxylation and methylation reactions to transform dimethylmalonyl-ACP, the product of MT1, to pivaloyl-ACP. Further, pivaloyl-ACP synthesis is primed by the fatty acid synthase malonyl acyltransferase (FabD), which compensates for the Psi GNAT and provides the initial aryl-transfer step to form AprA malonyl-ACP. Additionally, images of AprA from negative stain electron microscopy reveal multiple conformations that may facilitate the individual catalytic steps of the multienzyme module.

Wang, JL, Zheng JC, Huang CH, Zhao JY, Lin JJ, Zhou XZ, Naman CB, Wang N, Gerwick WH, Wang QW, Yan XJ, Cui W, He S.  2018.  Eckmaxol, a phlorotannin extracted from ecklonia maxima, produces anti-ss-amyloid oligomer neuroprotective effects possibly via directly acting on glycogen synthase kinase 3 ss. Acs Chemical Neuroscience. 9:1349-1356.   10.1021/acschemneuro.7b00527   AbstractWebsite

Alzheimer's disease is a progressive neurodegenerative disorder that mainly affects the elderly. Soluble ss-amyloid oligomer, which can induce neurotoxicity, is generally regarded as the main neurotoxin in Alzheimer's disease. Here we report that eckmaxol, a phlorotannin extracted from the brown alga Ecklonia maxima, could produce neuroprotective effects in SH-SY5Y cells. Eckmaxol effectively prevented but did not rescue ss-amyloid oligomer-induced neuronal apoptosis and increase of intracellular reactive oxygen species. Eckmaxol also significantly reversed the decreased expression of phospho-Ser9-glycogen synthase kinase 3 ss and increased expression of phospho-extracellular signal-regulated kinase, which was induced by A ss oligomer. Moreover, both glycogen synthase kinase 3 ss and mitogen activated protein kinase inhibitors produced neuroprotective effects in SH-SY5Y cells. Furthermore, eckmaxol showed favorable interaction in the ATP binding site of glycogen synthase kinase 3 ss and mitogen activated protein kinase. These results suggested that eckmaxol might produce neuroprotective effects via concurrent inhibition of glycogen synthase kinase 3 ss and extracellular signal-regulated kinase pathways, possibly via directly acting on glycogen synthase kinase 3 ss and mitogen activated protein kinase. Based on the central role that ss-amyloid oligomers play in the pathogenesis of Alzheimer's disease and the high annual production of Ecklonia maxima for alginate and other nutritional ingredients, this report represents a new candidate for the treatment of Alzheimer's disease, and also expands the potential application of Ecklonia maxima and its constituents in the field of pharmacology.

O'Rourke, A, Kremb S, Duggan BM, Sioud S, Kharbatia N, Raji M, Emwas AH, Gerwick WH, Voolstra CR.  2018.  Identification of a 3-Alkylpyridinium Compound from the Red Sea Sponge Amphimedon chloros with In Vitro Inhibitory Activity against the West Nile Virus NS3 Protease. Molecules. 23   10.3390/molecules23061472   AbstractWebsite

Viruses are underrepresented as targets in pharmacological screening efforts, given the difficulties of devising suitable cell-based and biochemical assays. In this study we found that a pre-fractionated organic extract of the Red Sea sponge Amphimedon chloros was able to inhibit the West Nile Virus NS3 protease (WNV NS3). Using liquid chromatography-mass spectrometry (LC-MS) and nuclear magnetic resonance (NMR) spectroscopy, the identity of the bioactive compound was determined as a 3-alkylpyridinium with m/z = 190.16. Diffusion Ordered Spectroscopy (DOSY) NMR and NMR relaxation rate analysis suggest that the bioactive compound forms oligomers of up to 35 kDa. We observed that at 9.4 mu g/mL there was up to 40-70% inhibitory activity on WNV NS3 protease in orthogonal biochemical assays for solid phase extracts (SPE) of A. chloros. However, the LC-MS purified fragment was effective at inhibiting the protease up to 95% at an approximate amount of 2 mu g/mL with negligible cytotoxicity to HeLa cells based on a High-Content Screening (HCS) cytological profiling strategy. To date, 3-alkylpyridinium type natural products have not been reported to show antiviral activity since the first characterization of halitoxin, or 3-alkylpyridinium, in 1978. This study provides the first account of a 3-alkylpyridinium complex that exhibits a proposed antiviral activity by inhibiting the NS3 protease. We suggest that the here-described compound can be further modified to increase its stability and tested in a cell-based assay to explore its full potential as a potential novel antiviral capable of inhibiting WNV replication.

Kristoffersen, V, Rama T, Isaksson J, Andersen JH, Gerwick WH, Hansen E.  2018.  Characterization of rhamnolipids produced by an Arctic marine bacterium from the Pseudomonas fluorescence group. Marine Drugs. 16   10.3390/md16050163   AbstractWebsite

The marine environment is a rich source of biodiversity, including microorganisms that have proven to be prolific producers of bioactive secondary metabolites. Arctic seas are less explored than warmer, more accessible areas, providing a promising starting point to search for novel bioactive compounds. In the present work, an Arctic marine Pseudomonas sp. belonging to the Pseudomonas (P.) fluorescence group was cultivated in four different media in an attempt to activate biosynthetic pathways leading to the production of antibacterial and anticancer compounds. Culture extracts were pre-fractionated and screened for antibacterial and anticancer activities. One fraction from three of the four growth conditions showed inhibitory activity towards bacteria and cancer cells. The active fractions were dereplicated using molecular networking based on MS/MS fragmentation data, indicating the presence of a cluster of related rhamnolipids. Six compounds were isolated using HPLC and mass-guided fractionation, and by interpreting data from NMR and high-resolution MS/MS analysis; the structures of the compounds were determined to be five mono-rhamnolipids and the lipid moiety of one of the rhamnolipids. Molecular networking proved to be a valuable tool for dereplication of these related compounds, and for the first time, five mono-rhamnolipids from a bacterium within the P. fluorescence group were characterized, including one new mono-rhamnolipid.

Tao, YW, Li PL, Zhang DJ, Glukhov E, Gerwick L, Zhang C, Murray TF, Gerwick WH.  2018.  Samholides, swinholide-related metabolites from a marine cyanobacterium cf. Phormidium sp. Journal of Organic Chemistry. 83:3034-3046.   10.1021/acs.joc.8b00028   AbstractWebsite

Cancer cell cytotoxicity was used to guide the isolation of nine new swinholide-related compounds, named samholides A-I (1-9), from an American Samoan marine cyanobacterium cf. Phormidium sp. Their structures were determined by extensive analysis of 1D and 2D NMR spectroscopic data. The new compounds share an unusual 20-demethyl 44-membered lactone ring composed of two monomers, and they demonstrate structural diversity arising from geometric isomerization of double bonds, sugar units with unique glyceryl moieties and varied methylation patterns. All of the new samholides were potently active against the H-460 human lung cancer cell line with IC50 values ranging from 170 to 910 nM. The isolation of these new swinholide-related compounds from a marine cyanobacterium reinvigorates questions concerning the evolution and biosynthetic origin of these natural products.

Shao, CL, Mou XF, Cao F, Spadafora C, Glukhov E, Gerwick L, Wang CY, Gerwick WH.  2018.  Bastimolide B, an antimalarial 24-membered marine macrolide possessing a tert-butyl group. Journal of Natural Products. 81:211-215.   10.1021/acs.jnatprod.7b00917   AbstractWebsite

We reported previously the discovery of the potent antimalarial 40-membered macrolide bastimolide A (1) from the tropical marine cyanobacterium Okeania hirsute. Continued investigation has led to the discovery of a new analogue, bastimolide B (2), a 24-membered polyhydroxy macrolide with a long aliphatic chain and unique terminal tertbutyl group. Its complete structure was determined by a combination of extensive spectroscopic methods and comparative analysis of its methanolysis products with those of bastimolide A. A methanolysis mechanism for bastimolide A is proposed, and one unexpected isomerization product of the C2-C3 double bond, 2-(E)-bastimolide A (3), was obtained. Bastimolide B (2) showed strong antimalarial activity against chloroquine-sensitive Plasmodium falciparum strain HB3. A preliminary investigation of the structure activity relationship based on six analogues revealed the importance of the double bond as well as the 1,3-diol and 1,3,5-triol functionalities.

Moss, NA, Leao T, Glukhov E, Gerwick L, Gerwick WH.  2018.  Collection, Culturing, and Genome Analyses of Tropical Marine Filamentous Benthic Cyanobacteria. Marine Enzymes and Specialized Metabolism, Pt A. 604( Moore BS, Ed.).:3-43., San Diego: Elsevier Academic Press Inc   10.1016/bs.mie.2018.02.014   Abstract

Decreasing sequencing costs has sparked widespread investigation of the use of microbial genomics to accelerate the discovery and development of natural products for therapeutic uses. Tropical marine filamentous cyanobacteria have historically produced many structurally novel natural products, and therefore present an excellent opportunity for the systematic discovery of new metabolites via the information derived from genomics and molecular genetics. Adequate knowledge transfer and institutional know-how are important to maintain the capability for studying filamentous cyanobacteria due to their unusual microbial morphology and characteristics. Here, we describe workflows, procedures, and commentary on sample collection, cultivation, genomic DNA generation, bioinformatics tools, and biosynthetic pathway analysis concerning filamentous cyanobacteria.

Skiba, MA, Sikkema AP, Moss NA, Tran CL, Sturgis RM, Gerwick L, Gerwick WH, Sherman DH, Smith JL.  2017.  A mononuclear iron-dependent methyltransferase catalyzes initial steps in assembly of the apratoxin A polyketide starter unit. Acs Chemical Biology. 12:3039-3048.   10.1021/acschembio.7b00746   AbstractWebsite

Natural product biosynthetic pathways contain a plethora of enzymatic tools to carry out difficult biosynthetic transformations. Here, we discover an unusual mononuclear iron dependent methyltransferase that acts in the initiation steps of apratoxin A biosynthesis (AprA MT1). Fe3+-replete AprA MT1 catalyzes one or two methyl transfer reactions on the substrate malonyl-ACP (acyl carrier protein), whereas Co2+, Fe2+, Mn2+, and Ni2+ support only a single methyl transfer. MT1 homologues' exist;Within the "GNAT" (GCNS-related N-acetyltransferase) loading modules of several modular biosynthetic pathways with propionyl, isobutyryt or pivaloyl starter units. GNAT domains are thought to catalyze decarboXylation of malonyl-CoA and acetyl transfer to a carrier protein. In AprA, the GNAT domain lacks both decarboxylation and acyl transfer activity. A crystal structure of the AprA MT1-GNAT di-domain with bound Mn2+, malonate, and the methyl donor S-adenosylmethionine (SAM) reveals that the malonyl substrate is a bidentate metal ligand, indicating that the metal acts as a Lewis acid to promote methylation of the malonyl alpha-carbon. The GNAT domain is truncated relative to functional homologues. These results afford an expanded understanding of MT1-GNAT structure and activity arid permit the functional annotation of homologous GNAT loading modules both with and without methyltransferases, additionally revealing their rapid evolutionary adaptation in different biosynthetic contexts.

Zhang, C, Idelbayev Y, Roberts N, Tao YW, Nannapaneni Y, Duggan BM, Min J, Lin EC, Gerwick EC, Cottrell GW, Gerwick WH.  2017.  Small Molecule Accurate Recognition Technology (SMART) to Enhance Natural Products Research. Scientific Reports. 7   10.1038/s41598-017-13923-x   AbstractWebsite

Various algorithms comparing 2D NMR spectra have been explored for their ability to dereplicate natural products as well as determine molecular structures. However, spectroscopic artefacts, solvent effects, and the interactive effect of functional group(s) on chemical shifts combine to hinder their effectiveness. Here, we leveraged Non-Uniform Sampling (NUS) 2D NMR techniques and deep Convolutional Neural Networks (CNNs) to create a tool, SMART, that can assist in natural products discovery efforts. First, an NUS heteronuclear single quantum coherence (HSQC) NMR pulse sequence was adapted to a state-of-the-art nuclear magnetic resonance (NMR) instrument, and data reconstruction methods were optimized, and second, a deep CNN with contrastive loss was trained on a database containing over 2,054 HSQC spectra as the training set. To demonstrate the utility of SMART, several newly isolated compounds were automatically located with their known analogues in the embedded clustering space, thereby streamlining the discovery pipeline for new natural products.

Gerwick, WH.  2017.  The Face of a Molecule. Journal of Natural Products. 80:2583-2588.   10.1021/acs.jnatprod.7b00624   AbstractWebsite

Recent technological advances in mass spectrometry and NMR spectroscopy have enabled new approaches for the rapid and insightful profiling of natural product mixtures. MALDI-MS with the provision of biosynthetic heavy-isotope labeled precursors can be a powerful method by which to interrogate a natural product metabolome and to gain insight into its unique constituents; this is illustrated herein by the detection, isolation, and characterization of cryptomaldamide. MS/MS-based Molecular Networks, facilitated by the Global Natural Products Social (GNPS) platform, is rapidly changing the way in which we dereplicate known natural products in mixtures, find new analogues in desired structure classes, and identify fundamentally new chemical entities. This method can be linked to genomic information to assist in genome-driven natural products discovery and is illustrated here with the characterization of the columbamides. Similarly, algorithmic interpretation of NMR data is facilitating the automatic identification or classification of new natural products. We developed such a tool named the Small Molecule Accurate Recognition Technology (SMART), which employs a convolutional neural network to classify HSQC spectra of organic molecules using pattern recognition principles. The discovery and rapid classification of several new peptides from a marine cyanobacterium as members of the viequeamide class provides an example of its utility in natural products research. These three illustrations represent different methods by which to look at the external features of a chemical substance and derive valuable insights into its identity or, as described herein, the "face of a molecule".

Fang, F, Zhao JY, Ding LJ, Huang CH, Naman CB, He S, Wu B, Zhu P, Luo QJ, Gerwick WH, Yan XJ, Wang QW, Zhang ZJ, Cui W.  2017.  5-Hydroxycyclopenicillone, a New beta-Amyloid Fibrillization Inhibitor from a Sponge-Derived Fungus Trichoderma sp HPQJ-34. Marine Drugs. 15   10.3390/md15080260   AbstractWebsite

A new cyclopentenone, 5-hydroxycyclopeni cillone (1), was isolated together with three known compounds, ar-turmerone (2), citreoisocoumarin (3), and 6-O-methyl-citreoisocoumarin (4), from a culture of the sponge-derived fungus Trichoderma sp. HPQJ-34. The structures of 1-4 were characterized using comprehensive spectroscopic analyses. The absolute configuration of 1 was determined by comparison of electronic circular dichroism (ECD) spectra with literature values used for the reported analogue, cyclopenicillone (5), which was not isolated in this research. Compound 1 was shown to scavenge 2,2-diphenyl-1-picrylhydrazyl free radicals, and decrease beta-amyloid (A beta) fibrillization in vitro. Moreover, 1 significantly reduced H2O2-induced neurotoxicity in SH-SY5Y cells. These findings suggested that compound 1, a newly discovered cyclopentenone, has moderate anti-oxidative, anti-A fi fibrillization properties and neuroprotective effects, and might be a good free radical scavenger.

Naman, CB, Almaliti J, Armstrong L, Caro-Diaz EJ, Pierce ML, Glukhov E, Fenner A, Spadafora C, Debonsi HM, Dorrestein PC, Murray TF, Gerwick WH.  2017.  Discovery and Synthesis of Caracolamide A, an Ion Channel Modulating Dichlorovinylidene Containing Phenethylamide from a Panamanian Marine Cyanobacterium cf. Symploca Species. Journal of Natural Products. 80:2328-2334.   10.1021/acs.jnatprod.7b00367   AbstractWebsite

A recent untargeted metabolomics investigation into the chemical profile of 10 organic extracts from cf. Symploca spp. revealed several interesting chemical leads for further natural product drug discovery. Subsequent target directed isolation efforts with one of these, a Panamanian marine cyanobacterium cf. Symploca sp., yielded a phenethylamide metabolite that terminates in a relatively rare gemdichlorovinylidene moiety, caracolarnide A (1), along with a known isotactic polymethoxy-1-alkene (2). Detailed NMR and HRESIMS analyses were used to determine the structures of these molecules, and compound 1 was confirmed by a three step synthesis. Pure compound 1 was shown to have in vitro calcium influx and calcium channel oscillation modulatory activity when tested as low as 10 pM using cultured murine cortical neurons, but was not cytotoxic to NCI-H460 human non-small-cell lung cancer cells in vitro (IC50 > 10 mu M).