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Current research energies are fixated on the synthesis of environmentally friendly and non-hazardous products, which include finding and recognizing biosurfactants that can substitute synthetic surfactants. Microbial biosurfactants are surface-active compounds synthesized intracellularly or extracellularly. To use biosurfactants in various industries, it is essential to understand scientific engagements that demonstrate its potentials as real advancement in the 21st century. Other than applying a substantial effect on the world economic market, engineered hyper-producing microbial strains in combination with optimized cultivation parameters have made it probable for many industrial companies to receive the profits of 'green' biosurfactant innovation. There needs to be an emphasis on the worldwide state of biosurfactant synthesis, expression of biosurfactant genes in expressive host systems, the recent developments, and prospects in this line of research. Thus, molecular dynamics with respect to genetic engineering of biosynthetic genes are proposed as new biotechnological tools for development, improved synthesis, and applications of biosurfactants. For example, mutant and hyper-producing recombinants have been designed efficaciously to advance the nature, quantity, and quality of biosurfactants. The fastidious and deliberate investigation will prompt a comprehension of the molecular dynamics and phenomena in new microorganisms. Throughout the decade, valuable data on the molecular genetics of biosurfactant have been produced, and this solid foundation would encourage application-oriented yields of the biosurfactant production industry and expand its utilization in diverse fields. Therefore, the conversations among different interdisciplinary experts from various scientific interests such as microbiology, biochemistry, molecular biology, and genetics are indispensable and significant to accomplish these objectives.

Here, we identified sequence variations of CYP72A69 and UGT73F4 genes from the Sojeongja and Haepum cultivars, and revealed that accumulation patterns of soyasaponin Aa and Ab were different in the Sojeongja and Haepum cultivars. Several genes related in soyasaponin biosynthetic pathway showed significant changes in their expression patterns in two soybean cultivars and soyasaponin contents were also altered under several abiotic stress conditions, albeit no significant correlations between soyasaponin contents and biosynthetic gene expression were observed. We also demonstrated that UGT73F4 gene could be responsible for biosynthesis of soyasaponin Aa in the Socheongja by investigating gene expression and soyasaponin accumulations. Our results provide a molecular basis to improve the commercial properties of soyasaponin in Korean domestic soybean cultivars.

While exploring phytotoxic metabolites from phytopathogenic fungi in the 1970s, we became interested in biosynthetic enzymes that catalyze Diels-Alder reactions involving biosynthesis of several phytotoxins that we isolated. Target enzymes were successfully characterized, and this triggered the identification of various Diels-Alderases in a recent decade. Through our Diels-Alderase project in 1990s, we recognized a highly efficient expression system of various biosynthetic genes with Aspergillus oryzae as a host. With the development of tools such as genomic data and bioinformatics analysis to identify biosynthetic gene clusters for natural products, we developed a highly reliable methodology such as hot spot knock-in to elucidate the biosynthetic pathways of representative fungal metabolites including phytotoxic substances. This methodology allows total biosynthesis of natural products and genome mining using silent biosynthetic gene clusters to obtain novel bioactive metabolites. Further applications of this technology are discussed.

Anthocyanins confer the wide range of colors for plants and also play beneficial health roles as potentially protective factors against heart disease and cancer. Brassica juncea is cultivated as an edible oil resource and vegetable crop worldwide, thus elucidating the anthocyanin biosynthetic pathway would be helpful to improve the nutritional quality of Brassica juncea through the breeding and cultivating of high anthocyanin content varieties. Herein, 129 genes in B. juncea were identified as orthologs of 41 anthocyanin biosynthetic genes (ABGs) in Arabidopsis thaliana by comparative genomic analyses. The B. juncea ABGs have expanded by whole genome triplication and subsequent allopolyploidizatoin, but lost mainly during the whole genome triplication between B. rapa/B. nigra and A. thaliana, rather than the allopolyploidization process between B. juncea and B. rapa/B. nigra, leading to different copy numbers retention of A. thaliana homologous genes. Although the overall expansion levels ABGs were similar to the whole genome, more negative regulatory genes were retained in the anthocyanin biosynthesis regulatory system. Transcriptional analysis of B. juncea with different anthocyanin accumulation showed that BjDFR, BjTT19, BjTT8 are significantly up-regulated in plants with purple leaves as compared with green leaves. The overexpression of BjTT8 and these target genes which were involved in late anthocyanin biosynthesis and transport might account for increasing levels of anthocyanin accumulation in purple leaves. Our results could promote the understanding of the genetic mechanism of anthocyanin biosynthesis in B. juncea.

Advances in next-generation DNA sequencing technologies, bioinformatics, and mass spectrometry-based metabolite detection have ushered in a new era of natural product discovery. Microbial secondary metabolomes are complex, especially when otherwise silent biosynthetic genes are activated, and there is therefore a need for data analysis software to explore and map the resulting multidimensional datasets. To that end, we herein report the Metabolomics Explorer (MetEx), a publicly available web application for the analysis of parallel liquid chromatography-coupled mass spectrometry (LC-MS)-based metabolomics data. MetEx is a highly interactive application that facilitates visualization and analysis of complex metabolomics datasets, consisting of retention time, m/z, and MS intensity features, as a function of hundreds of conditions or elicitors. The software enables prioritization of leads from three-dimensional maps, extraction of two-dimensional slices from various higher order plots, organization of datasets by elicitor chemotypes, customizable library-based dereplication, and automatically scored lead selection. We describe the application of MetEx to the first UPLC-MS-guided high-throughput elicitor screen in which Burkholderia gladioli was challenged with 750 elicitors, and the resulting profiles were interrogated by UPLC-Qtof-MS and subsequently analyzed with the app. We demonstrate the utility of MetEx by reporting elicitors for several cryptic metabolite groups and by uncovering new natural products that remain to be characterized. MetEx is available at https://mo.princeton.edu/MetEx/.

Heparin is an essential anticoagulant drug discovered over a century ago. Heparin is the second most highly used natural drug and remains a mainstay of therapy with an expected global market share of more than $14 billion in the next 10 years. However, it is still naturally derived from unsustainable animal sources, such as bovine lungs and porcine intestines, as an unfractionated, heterogeneous complex mixture with unpredictable pharmacokinetic properties. Extensive research has been done in devising bioengineering and chemical approaches to produce structurally specific heparin and heparin-like polymers. Though several challenges remain, one of the main bottlenecks is the rapid, high-yield production of recombinant heparosan, a heparin precursor, which is originally isolated from a pathogenic E. coli K5 strain. Herein, we outline the methods for producing metabolically engineered size-specific heparosan, by transforming the essential heparosan biosynthetic genes into nonpathogenic E.coli strain BL21(DE3), in a highly controlled manner. The methods described herein are promising and can be easily scaled up for large-scale production of heparin-like structures.

Environmental microorganisms continue to serve as a major source of bioactive natural products (NPs) and as an inspiration for many other scaffolds in the toolbox of modern medicine. Nearly all microbial NP-inspired therapies can be traced to field expeditions to collect samples from the environment. Despite the importance of these expeditions in the search for new drugs, few studies have attempted to document the extent to which NPs or their corresponding production genes are distributed within a given environment. To gain insights into this, the geographic occurrence of NP ketosynthase (KS) and adenylation (A) domains was documented across 53 and 58 surface sediment samples, respectively, covering 59,590 square kilometers of Lake Huron. Overall, no discernible NP geographic distribution patterns were observed for 90,528 NP classes of nonribosomal peptides and polyketides detected in the survey. While each sampling location harbored a similar number of A domain operational biosynthetic units (OBUs), a limited overlap of OBU type was observed, suggesting that at the sequencing depth used in this study, no single location served as a NP "hotspot". These data support the hypothesis that there is ample variation in NP occurrence between sampling sites and suggest that extensive sample collection efforts are required to fully capture the functional chemical diversity of sediment microbial communities on a regional scale.

Milbemycins and their semisynthetic derivatives are recognized as effective and eco-friendly pesticides, whereas the high price limits their widespread applications in agriculture. One of the pivotal questions is the accumulation of milbemycin-like by-products, which not only reduces the yield of the target products milbemycin A3/A4, but also brings difficulty to the purification. With other analogous by-products abolished, α9/α10 and β-family milbemycins remain to be eliminated. Herein, we solved these issues by engineering of post-modification steps. First, Cyp41, a CYP268 family cytochrome P450, was identified to participate in α9/α10 biosynthesis. By deleting cyp41, milbemycin α9/α10 was eliminated with an increase of milbemycin A3/A4 titer from 2382.5 ± 55.7 mg/L to 2625.6 ± 64.5 mg/L. Then, MilE, a CYP171 family cytochrome P450, was determined to be responsible for the generation of the furan ring between C6 and C8a of milbemycins. By further overexpression of milE, the production of β-family milbemycins was reduced by 77.2%. Finally, the titer of milbemycin A3/A4 was increased by 53.1% to 3646.9 ± 69.9 mg/L. Interestingly, overexpression of milE resulted in increased transcriptional levels of milbemycin biosynthetic genes and production of total milbemycins, which implied that the insufficient function of MilE was a limiting factor to milbemycin biosynthesis. Our research not only provides an efficient engineering strategy to improve the production of a commercially important product milbemycins, but also offers the clues for future study about transcriptional regulation of milbemycin biosynthesis.

In the course of our studies on the heterologous expression of giant biosynthetic genes, we discovered a novel cryptic biosynthetic gene cluster in Streptomyces rochei IFO12908. During our efforts to express biosynthetic genes using the host SUKA strain derived from Streptomyces avermitilis, a novel polyene macrolactam compound designated as JBIR-156 was produced. We report herein the cloning and heterologous expression of the JBIR-156 biosynthetic gene cluster, and the isolation, structure determination, and cytotoxic activity of this novel compound.

Glucosinolates (GSLs) are important secondary metabolites in Brassicales related to insect and disease resistance, flavor formation, and human health. Here, we determined the GSL profile with sinigrin as the predominant GSL in Brassica nigra. A total of 184 GSL biosynthetic genes (BniGSLs) were identified in B. nigra by a genome-wide search for orthologs of 82 of the 95 known GSL genes in Arabidopsis thaliana. Transcriptome data demonstrated that at least one BniGSL was highly expressed in stems and leaves at each step of the sinigrin synthesis pathway, which ensured the synthesis of a large amount of sinigrin in B. nigra. Among these key candidates of BniGSLs, the high expression of BniMAM1-2, BniCYP79F1, and BniAOP2-1/2, and the absence of MAM3 and AOP3, may contribute remarkably to the synthesis and accumulation of sinigrin. In addition, the low expression of some key BniGSLs partially explains the low content of indolic and aromatic GSLs in B. nigra. This study provided a genetic explanation for the formation of the unique GSL profile with sinigrin as the main GSL in B. nigra. The results of this study will be valuable for further functional analysis of BniGSLs and genetic improvement of GSLs in B. nigra and other Brassica species.

Aberrant splicing of pre-mRNA is implicated in many human genetic disorders. Small molecules that target the spliceosome are important leads as therapeutics and research tools, and one compound of significant interest is the polyketide natural product pladienolide B. Here, we describe the reactivation of quiescent pladienolide B production in the domesticated lab strain Streptomyces platensis AS6200 by overexpression of the pathway-specific activator PldR. The resulting dysregulation of the biosynthetic genes led to the accumulation and isolation of five additional intermediate or shunt metabolites of pladienolide B biosynthesis, including three previously unreported congeners. These compounds likely comprise the entire pladienolide biosynthetic pathway and demonstrate the link between polyketide tailoring reactions and bioactivity, particularly the importance of the 18,19-epoxide. Each congener demonstrated specific inhibitory activity against mammalian cell lines, with successive modifications leading to increased activity (IC50: 8 mM to 5 μM)., Reactivation of quiescent polyketide production in a domesticated lab strain.

Covering: up to September 2020 Mushroom-forming fungi of the division Basidiomycota have traditionally been recognised as prolific producers of structurally diverse and often bioactive secondary metabolites, using the methods of chemistry for research. Over the past decade, -omics technologies were applied on these fungi, and sophisticated heterologous gene expression platforms emerged, which have boosted research into the genetic and biochemical basis of the biosyntheses. This review provides an overview on experimentally confirmed natural product biosyntheses of basidiomycete polyketides, amino acid-derived products, terpenoids, and volatiles. We also present challenges and solutions particular to natural product research with these fungi. 222 references are cited.

Summary We demonstrate the production of cholesterol by Enhygromyxa salina, a heterotrophic marine bacterium. Cholesterol is a highly modified sterol that is preserved as cholestane in the rock record. Cholestane is often indicative of heterotrophic microbial eukaryotes. The production of cholesterol by E. salina is the first reported instance of cholesterol synthesis by a bacterium. Through genomic analyses, we show that E. salina has cholesterol biosynthetic genes that are homologous to the eukaryotic cholesterol pathway. However, we do observe some biosynthetic proteins that are unique to E. salina. Further, extensive lipid analyses demonstrate that cholesterol in this organism is conjugated to either a lipid or protein moiety and does not exist as a free sterol. Conjugated sterols play significant physiological roles in eukaryotes and understanding the nature of conjugated cholesterol in E. salina may provide insight into the physiological roles of these lipids in bacteria. Through these studies, we should be able to better assess what implications, if any, cholesterol production by bacteria has on our interpretation of sterol biomarkers in the rock record.

Paeonia ostii seeds were identified as novel sources of edible plant oil with a high proportion of α-linolenic acid, a type of n-3 fatty acid with many health benefits. Due to the unreliability of seed oil content and quality, it is necessary to discover the mechanism underlying lipid biosynthesis in Paeonia ostii seeds. This study aimed to identify the key genes involved in lipid biosynthesis in Paeonia ostii seeds by analyzing the relationship among the seed characteristics and the expression patterns of lipid genes in Paeonia ostii during seed development. Preliminary research on Paeonia ostii seed development was carried out from 10 days after pollination until maturity, focusing on phenology, oil content and lipid profiles. In addition, we investigated the spatiotemporal expression of 36 lipid biosynthetic genes in Paeonia ostii by using quantitative real-time PCR. The results suggested that the development of Paeonia ostii seeds from pollination to maturity could be divided into three periods. The 36 lipid genes showed various spatiotemporal expression patterns and five gene groups with distinct temporal patterns during seed development were identified by clustering analysis of expression data. Furthermore, the relationships between gene expression and lipid/fatty acid accumulation and some candidate key lipid genes were discussed. This study provided the global patterns of fatty acid and lipid biosynthesis-related gene expression, which are critical to understanding the molecular basis of lipid biosynthesis and identifying the lipid accumulation rate-limiting genes during seed development.

Colchicine is among the oldest plant natural products (NPs) still used for treating a broad spectrum of human diseases including gout and other articular inflammation disorders. This molecule is synthesized by several herbaceous species related to the Liliaceae family, but in very low quantities in whole plants. As for many pharmaceutical compounds from plants, the production of colchicine still depends on the natural resource from which it is extracted. From the past decade, metabolic engineering has progressively become a credible alternative for the cost-effective large-scale production of several valuable NPs. In the same vein, Nett and colleagues recently reported an unprecedented advance in the field for colchicine. By using a combination of transcriptomics, metabolomics and pathway reconstitution, Sattely's group deciphered a near-complete biosynthetic pathway to colchicine without prior knowledge of biosynthetic genes. Besides constituting a benchmark for the elucidation of natural product biosynthetic pathways, it opens unprecedented perspectives regarding metabolic engineering of colchicine biosynthesis.

8-Azaguanine (1) is a special 1,2,3-triazole containing natural product that possesses potent antibacterial and antitumor activities. In the present study, the entire 8-azaguanine biosynthetic gene cluster was located from Streptomyces CGMCC4.1633. Targeted gene disruption, heterologous expression analysis, and feeding experiments identified crucial genes for 8-azaguanine production. Moreover, we characterized the structure of two novel metabolites, analyzed NO (or reactive nitrogen species) related genes 8-azgA/B and radical SAM enzyme homologous 8-AzgG, and verified the non-enzymatic ring formation reaction of 8-azaguanine 1,2,3-triazole. All of the data and presumptions provide insight into the timing and mechanism of the enzymatic and non-enzymatic pathway that produce 8-azaguanine-type 1,2,3-triazole., Graphical Abstract

The root of Phlomis umbrosa has traditionally been used as a medicine in South Asian nations to treat colds and bone fractures, to staunch bleeding, and as an anti-inflammatory, and such use continues today. We identified 10 genes that are involved in terpenoid biosynthesis, while using the Illumina/Solexa HiSeq2000 platform. We investigated the transcript levels of the 10 genes using quantitative real-time PCR and quantified the level of terpenoid accumulation in different organs of P. umbrosa while using high-performance liquid chromatography. The transcript levels of PuHDR and PuHMGR1 were the highest among the studied genes. Sesamoside, an iridoid glycoside, appeared in higher quantity than shanzhiside methylester, umbroside (8-O-acetyl shanzhiside methyl ester), and acteoside. We speculate that PuHDR and PuHMGR1 may contribute to terpenoid biosynthesis in P. umbrosa. This study highlights the molecular mechanisms that underlie iridoid glycoside biosynthesis in P. umbrosa.

Indolmycin is an antibiotic from Streptomyces griseus ATCC 12648 with activity against Helicobacter pylori, Plasmodium falciparum, and methicillin-resistant Staphylococcus aureus. Here we describe the use of the indolmycin biosynthetic genes in E. coli to make indolmycenic acid, a chiral intermediate in indolmycin biosynthesis, which can then be converted to indolmycin through a three-step synthesis. To expand indolmycin structural diversity, we introduce a promiscuous tryptophanyl-tRNA synthetase gene (trpS) into our E. coli production system and feed halogenated indoles to generate the corresponding indolmycenic acids, ultimately allowing us to access indolmycin derivatives through synthesis. Bioactivity testing against methicillin-resistant Staphylococcus aureus showed modest antibiotic activity for 5-, 6-, and 7-fluoro-indolmycin., A semi-synthetic system for producing indolmycin, an antibiotic, was developed and used to make indole-substituted, halogenated derivatives of indolmycin, some with modest bioactivity against methicillin-resistant Staphylococcus aureus.

The content of pullulan and melanin in 500 mutants of Aureobasidum pullulans obtained by ultraviolet mutagenesis were examined and statistically analyzed, and a strong positive correlation was found between them. The result was further confirmed by culturing wild type strain As3.3984 in different media. Then we constructed melanin-deletion mutant As-Δalb1 and pullulan-deletion mutant As-Δpul. As-Δalb1 was a melanin-free strain with the yield of pullulan decreased by 41.01%. The supplementation of melanin in the culture of As-Δalb1 increased the production of pullulan. As-Δpul synthesized neither pullulan nor melanin and recovered melanin synthesis by adding pullulan to the medium. The results suggested that high concentration- of pullulan induced morphological transformation and synthesis of melanin, and melanin promoted the synthesis of pullulan. The pullulan biosynthetic genes, upt, pgm, ugp, and pul, were down-regulated, while the negative regulatory gene of pullulan synthesis, creA, was up-regulated by melanin deficiency.

A fluorometabolite, 5-fluoro-5-deoxy-D-ribulose (5-FDRul), from the culture broth of the soil bacterium Streptomyces sp. MA37, was identified through a combination of genetic manipulation, chemo-enzymatic synthesis and NMR comparison. Although 5-FDRul has been chemically synthesized before, it was not an intermediate or a shunt product in previous studies of fluorometalism in S. cattleya. Our study of MA37 demonstrates that 5-FDRul is a naturally occurring fluorometabolite, rendering it a new addition to this rare collection of natural products. The genetic inactivation of key biosynthetic genes involved in the fluorometabolisms in MA37 resulted in the increased accumulation of unidentified fluorometabolites as observed from 19F-NMR spectral comparison among the wild type (WT) of MA37 and the mutated variants, providing evidence of the presence of other new biosynthetic enzymes involved in the fluorometabolite pathway in MA37.

Streptomycetes are an important source of natural products potentially applicable in the pharmaceutical industry. Many of these drugs are secondary metabolites whose biosynthetic genes are very often poorly expressed under laboratory cultivation conditions. In many cases, antibiotic-resistant mutants exhibit increased production of natural drugs, which facilitates the identification and isolation of new substances. In this study, we report the induction of a type II polyketide synthase gene cluster in the marine strain Streptomyces albus subsp. chlorinus through the selection of streptomycin-resistant mutants, resulting in overproduction of the novel compound fredericamycin C2 (1). Fredericamycin C2 (1) is structurally related to the potent antitumor drug lead fredericamycin A.

Flavonoids represent a large diverse group of natural products that are used as a traditional medicine against various infectious diseases. They possess many biological activities including antimicrobial, antioxidant, anti-inflammatory, anti-cancer and anti-diabetic activities. Commercially, flavonoids are mainly obtained from plants, however, several challenges are faced during their extraction. Microorganisms have been known as natural sources of a wide range of bioactive compounds including flavonoids. Actinobacteria are the most prolific group of microorganisms for the production of bioactive secondary metabolites, thus facilitating the production of flavonoids. The screening programs for bioactive compounds revealed the potential application of actinobacteria to produce flavonoids with interesting biological activities, especially anticancer activities. Since marine actinobacteria are recognized as a potential source of novel anticancer agents, they are highly expected to be potential producers of anticancer flavonoids with unusual structures and properties. In this review, we highlight the production of flavonoids by actinobacteria through classical fermentation, engineering of plant biosynthetic genes in a recombinant actinobacterium and the de novo biosynthesis approach. Through these approaches, we can control and improve the production of interesting flavonoids or their derivatives for the treatment of cancer.

Environmental changes resulted in a variety of stresses in plants of which heavy metal stress holds important position, affect the growth and development and trigger a series of morphological, physiological, biochemical and molecular changes in plants. When exposed heavy metal stress, the complex dynamickinetics of polyamine biosynthesis was observed. Polyamines are small organic polycations present in all organisms and have a leading role in signaling, plant growth and development and deliver tolerance to a cultivar against stresses. High accumulation of polyamines (putrescine, spermidine and spermine) in plantsduring heavy metal stress has been well reported and is correlated with increased tolerance to different plants under stressed condition. Genetic engineering of polyamine biosynthetic genes in crop plants is the way to create resistance heavy metal toxicity.

Ganoderic triterpenoid (GT) is a promising anti-tumour constituent of Ganoderma lucidum. GT accumulation occurs following stressors that include various elicitors or signalling molecules. Understanding the signal transduction pathways underlying elicitor-induced GT production will be helpful for commercial GT production. In this study, the activation of GT biosynthetic genes and GT accumulation mediated by biotic and abiotic elicitors will be investigated. Jasmonic acid (JA) is a well-known signalling molecule that is primarily involved in GT accumulation. Ca2+ spiking and reactive oxygen species (ROS), nitric oxide (NO) and cAMP production are other messengers that mediate GT production. This review highlights the elicitation strategies for GT production based on the principle of putative signal transduction pathways and the fundamental role of JA, ROS, NO, Ca2+, cAMP and their crosstalk in G. lucidum cells. In the future, elucidating the regulatory networks that control the secondary metabolite biosynthesis is therefore an important goal to manipulate the accumulation of GT.

A new series comprising phenylacetyl-homoserine lactones (HSLs), caffeoyl-HSL and feruloyl-HSL, was biologically synthesized using an artificial de novo biosynthetic pathway. We developed an Escherichia coli system containing artificial biosynthetic pathways that yield phenylacetyl-HSLs from simple carbon sources. These artificial biosynthetic pathways contained the LuxI-type synthase gene (rpaI) in addition to caffeoyl-CoA and feruloyl-CoA biosynthetic genes, respectively. Finally, the yields for caffeoyl-HSL and feruloyl-HSL were 97.1 ± 10.3 and 65.2 ± 5.7 mg/l, respectively, by tyrosine-overproducing E. coli with a L-methionine feeding strategy. In a quorum sensing (QS) competition assay, feruloyl-HSL and p-coumaroyl-HSL antagonized the QS receptor TraR in Agrobacterium tumefaciens NT1, whereas caffeoyl-HSL did not.

In the present study, efforts were made to determine effect of different light wavelengths on carotenoid biosynthesis in Scutellaria baicalensis, a promising traditional herbal medicine used in North America and Asia. The variation in the transcriptional level of carotenoid biosynthesis genes and carotenoid contents in the callus was analyzed after exposure to light from red (wavelength, 660 nm), blue (wavelength, 470 nm), or white (wavelength, 380 nm) light-emitting diodes. The transcription levels of phytoene synthase (SbPSY), ξ-carotene desaturase (SbZDS), β-ring carotene hydroxylase (SbCHXB), and zeaxanthin epoxidase (SbZEP) were upregulated on exposure to all three lights, with the greatest increase in expression observed upon exposure to blue light. In contrast, the transcriptional levels of two carotenoid cleavage dioxygenases (SbCCD1 and SbCCD4) decreased under all three light treatments. Increased production of zeaxanthin, β-carotene, and 9-cis-β-carotene was observed on exposure to all 3 lights. The total carotenoid production rose by 27.35% under blue light treatment, while the white and red lights did not have any considerable effect. Our findings may establish new approaches for increasing carotenoid production in S. baicalensis as well as in other plants.

This review analyzes the advances in understanding the natural signaling pathways and environmental factors regulating stilbene biosynthesis. We also discuss the studies reporting on stilbene content and repertoire in plants. Stilbenes, including the most-studied stilbene resveratrol, are a family of phenolic plant secondary metabolites that have been the subject of intensive research due to their valuable pharmaceutical effects and contribution to plant disease resistance. Understanding the natural mechanisms regulating stilbene biosynthesis in plants could be useful for both the development of new plant protection strategies and for commercial stilbene production. In this review, we focus on the environmental factors and cell signaling pathways regulating stilbene biosynthesis in plants and make a comparison with the regulation of flavonoid biosynthesis. This review also analyzes the recent data on stilbene biosynthetic genes and summarizes the available studies reporting on both stilbene content and stilbene composition in different plant families.

Filamentous fungi produce a wide range of natural products that are commonly used in various industrial contexts (e.g., pharmaceuticals and insecticides). Meroterpenoids are natural products of interest because of their various biological activities. Among the meroterpenoids, there is a group of insecticidal compounds known as the austinoids. These compounds have also been studied because of their intriguing spiro-lactone ring formation along with various modifications. Here, we present an extension of the original austinol/dehydroaustinol biosynthesis pathway from Aspergillus nidulans in the recently identified filamentous fungus Aspergillus calidoustus. Besides the discovery and elucidation of further derivatives, genome mining led to the discovery of new putative biosynthetic genes. The genes involved in the biosynthesis of later austinoid products were characterized, and among them was a second polyketide synthase gene in the A. calidoustus cluster that was unusual because it was a noninterative polyketide synthase producing a diketide. This diketide product was then loaded onto the austinoid backbone, resulting in a new insecticidal derivative, calidodehydroaustin.

Lasso peptides are ribosomally synthesized and posttranslationally modified peptides with diverse biological functions. Recent genome mining has revealed that many species of actinomycetes possibly contain biosynthetic gene clusters of lasso peptides. With genome mining for lasso peptide biosynthesis, we screened several actinomycetes for lasso peptide production using high-performance liquid chromatography and electrospray ionization–mass spectrometry. Consequently, Streptomyces cattleya was identified as a producer of a new lasso peptide named cattlecin. Analysis of amino acid content on cattlecin indicated the presence of four moles each of Asp and His, three moles each of Gly and Tyr, and one mole of Ser. Tandem mass spectrometry (MS/MS) analysis of cattlecin revealed C-terminal sequence of WHHGWYGWWDD. The peptide sequence (SYHWGDYHDWHHGWYGWWDD) was the expected amino acid sequence of cattlecin based on genome mining. As a result of MS/MS analysis, the amine residue of the first Ser was proposed to form a macrolactam ring with the β-carboxyl residue of the ninth Asp. The biosynthetic gene cluster of cattlecin comprised four genes: catA, catC, catB1, and catB2, which is typical of a lasso peptide biosynthetic gene cluster in actinomycetes.

One strain many compounds (OSMAC) strategy was an effective method to activate the silent biosynthetic genes of microorganisms. Comparison with our previous investigations on the secondary metabolites of the marine-derived fungus Aspergillus sp. SCSIO 41501, in this study, three new cyclopentenone derivatives, aspergispones A-C (1–3), and five new cyclohexenone derivatives, aspergispones D-H (4–8), together with two known analogues, were further isolated from the fungal strain by altering culture medium compositions. The structures of new compounds were elucidated by extensive spectroscopic analysis. And the absolute configurations of 4 and 7 were further confirmed by single-crystal X-ray diffraction experiments.

The interspecies communication roles of γ-butyrolactones (GBLs) have been described for a long time but are still poorly understood. Herein, we analyzed more than 1000 Streptomyces strains and noticed a big quantitative gap between the strains with GBL biosynthetic genes and the strains with GBL receptor genes, which implies the wide-spread of GBLs as interspecies signals in Streptomyces and their great potential in the activation of silent natural product gene clusters. Streptomyces albidoflavus J1074, which has one GBL receptor gene but no GBL biosynthetic gene, was chosen as a target to study the possible interspecies communication roles of GBLs. At first, the GBL biosynthetic genes from Streptomyces coelicolor M145 were expressed in S. albidoflavus J1074, which enabled the S. albidoflavus strains to synthesize Streptomyces coelicolor butanolides (SCBs) and activated the production of paulomycins. Further studies showed that this activation process requires the participation of the GBL receptor gene XNR_4681. The results suggest that the expression of exogenous GBL biosynthetic genes can modulate the metabolisms of GBL non-producing strains, and this regulation role might be meaningful for silent gene cluster activation in Streptomyces. At final, we synthesized racemic-SCB2 and tried to simplify the activation process by adding SCB2 directly to S. albidoflavus J1074, which unfortunately failed to induce paulomycin production.

Current understanding of the role of plant hormones during cereal grain filling is confounded by contradictory reports on hormone production that is based on poor methodology. We report here on the accurate measurement of indole-3-acetic acid (IAA) and abscisic acid (ABA) by combined liquid chromatography-tandem mass spectrometry in multiple reaction-monitoring mode with heavy isotope labelled internal standards. ABA and IAA contents of superior versus inferior rice grains (ABA maxima 159 ng g–1 FW and 109 ng g–1 FW, IAA maxima 2 µg g–1 FW and 1.7 µg g–1 FW respectively) correlated with the expression of biosynthetic genes and with grain fill. Results confirm that grain ABA is produced primarily by OsNCED2(5), but suggest that ABA import and metabolism also play important roles in ABA regulation. The IAA content of grains is primarily influenced by OsYUC9 and OsYUC11. However, the distinct expression profile of OsYUC12 suggests a specific role for IAA produced by this enzyme. Co-expression of OsYUC12 with OsIAA29 indicates their involvement in a common signalling pathway. Co-expression and cis-element analysis identified several aleurone-specific transcriptional regulators as well as glutelin as strong candidates for detailed investigation for direct regulation by the auxin-signalling pathway.

Background: Pueraria candollei var. mirifica , a Thai medicinal plant used traditionally as a rejuvenating herb, is known as a rich source of phytoestrogens, including isoflavonoids and the highly estrogenic miroestrol and deoxymiroestrol. Although these active constituents in P. candollei var. mirifica have been known for some time, actual knowledge regarding their biosynthetic genes remains unknown. Results: A de novo transcriptome analysis was conducted using combined P. candollei var. mirifica tissues of young leaves, mature leaves, tuberous cortices, and cortex-excised tubers.A total of 166,923 contigs was assembled for functional annotation using protein databases and as a library for identification of genes that are potentially involved in the biosynthesis of isoflavonoids and miroestrol. Twenty-one differentially expressed genes from four separate libraries were identified as candidatesinvolved in these biosyntheticpathways, and their respective expressions were validated by quantitative real-time reverse transcription polymerase chain reaction. Notably, isoflavonoid profiling generated by LC-MS/MS was positively correlated with expression levels of isoflavonoid biosynthetic genes across the four types of tissues. Moreover, we identified R2R3 MYB transcription factors that may be involved in the regulation of isoflavonoid biosynthesis in P. candollei var. mirifica . To confirm the function of a key-isoflavone biosynthetic gene, P. candollei var. mirifica isoflavone synthase identified in our library was transiently co-expressed with an Arabidopsis MYB12 transcription factor ( At MYB12) in Nicotiana benthamiana leaves. Remarkably, the combined expression of these proteins ledto the production of the isoflavone genistein. Conclusions: Our results provide compelling evidence regarding the integration of transcriptome and metabolome as a powerful tool for unraveling biosynthetic pathways in plants.

Covering: 2005 up to 2019Natural products are of paramount importance in human medicine. Not only are most antibacterial and anticancer drugs derived directly from or inspired by natural products, many other branches of medicine, such as immunology, neurology, and cardiology, have similarly benefited from natural product-based drugs. Typically, the genetic material required to synthesize a microbial specialized product is arranged in a multigene biosynthetic gene cluster (BGC), which codes for proteins associated with molecule construction, regulation, and transport. The ability to connect natural product compounds to BGCs and vice versa, along with ever-increasing knowledge of biosynthetic machineries, has spawned the field of genomics-guided natural product genome mining for the rational discovery of new chemical entities. One significant challenge in the field of natural product genome mining is how to rapidly link orphan biosynthetic genes to their associated chemical products. This review highlights state-of-the-art genetic platforms to identify, interrogate, and engineer BGCs from diverse microbial sources, which can be broken into three stages: (1) cloning and isolation of genomic loci, (2) heterologous expression in a host organism, and (3) genetic manipulation of cloned pathways. In the future, we envision natural product genome mining will be rapidly accelerated by de novo DNA synthesis and refactoring of whole biosynthetic pathways in combination with systematic heterologous expression methodologies.

New technology for the derivatization of peptide natural products is required for drug development. Despite the recent advances in the genome sequencing technique enabling us to search for the biosynthetic genes for wide variety of natural products, the technical methods to get access to them are limited. A class of RiPPs, a recently emerged natural product family such as thioviridamide, is one of those possessing such unexplored chemical space. In this paper, we report a streamlined method to generate new thioviridamide derivatives and to assess their biological activities. Heterologous expression of 42 constructs in an engineered Streptomyces avermitilis host gave 35 designed thioviridamide derivatives, along with several unprecedented analogues. Moreover, cytotoxicity assay revealed that several derivatives showed more potent activities than those of prethioviridamide. These results indicate that this strategy can become one of the potential ways to produce supreme unnatural products.

Chlorophyll (Chl) biosynthesis requires highly efficient metabolic regulation during chloroplast biogenesis in angiosperms. The intermediates of Chl biosynthesis and Chls are easily activated by li...

Terrestrial bacteria, especially actinomycetes, are known to be prolific producers of volatile compounds. We show here that bacteria from ocean sediments can also release complex bouquets of volatiles. The actinomycete Salinispora tropica produces cyclohexenyl compounds not previously known in nature, such as methyl cyclohex-2-ene-1-carboxylate (9), methyl 2-(cyclohex-2-en-1-yl)acetate (10), methyl (E/Z)-2-(cyclohex-2-en-1-ylidene)acetate (11/12), and related alcohols 8 and 13. These compounds were identified by GC/MS and confirmed by synthesis. In addition, rare spiroacetals, aromatic compounds, short-chain acids and esters, alcohols, and various cyclic compounds were produced by the bacteria. The biosynthesis of the cyclohexenyl compounds is closely coupled to that of cyclohexenylalanine (4), a building block of salinosporamide A, a proteasome inhibitor produced by S. tropica. Analysis of S. tropica strains that harbor knockouts of the salinosporamide biosynthetic genes salX and salD, coupled with feeding experiments, revealed that 3-(cyclohex-2-en-1-yl)-2-oxopropanoic acid (60) and 3-(cyclohex-2-en-1-ylidene)-2-oxopropanoic acid (isomers 61 and 62) are important intermediates in the biosynthesis of salinosporamide A, 4, and 8-13.

Astragalosides (AGs) are triterpene saponins that uniquely occur in Astragalus species with multiple pharmacological activities. For the first time, we reported an elicitation strategy, using UV radiation (UV-A, UV-B, and UV-C), to enhance AG production in Astragalus membranaceus hairy root cultures (AMHRCs). The treatment of 32 day-old AMHRCs with 54 kJ/m 2 dose of UV-B radiation was capable of achieving the maximum production of AGs, i.e. 3.431 ± 0.092 mg/g dry weight (DW), which was 1.30-fold higher as compared to the level in non-treated control (2.645 ± 0.073 mg/g DW). Expressions of eleven genes involved in AG biosynthetic pathway were monitored by quantitative real-time PCR (qRT-PCR). Transcriptional results showed that only the HMGR gene was significantly up-regulated in response to UV-B elicitation. The present study demonstrated that UV-B elicitation is a feasible strategy to promote AG production in AMHRCs and that HMGR represents a potential key gene in AG biosynthetic pathway.

Light quality and intensity have severe impact on Steroidal glycoalkaloid (SGA) biosynthesis, causing the quality degradation of potato by greening and accelerating SGA-induced toxicity. In this study, Atlantic and Haryoung cultivars responded differently when exposed to seven light qualities viz., purple, red, blue, green, yellow, UV and fluorescent lights. Haryoung tuber exhibited a 44 % lesser SGA accumulation in yellow light compared to fluorescent light. Further screening of selected light sources by investigating the expression pattern of key genes and SGA contents, showed lower transcript levels of key genes in yellow light which in-turn reflected in the least amount of SGA accumulation with a 22 % lesser SGA levels in yellow light compared to fluorescent light. This suppression in the SGA accumulation suggests that yellow light can be used as an alternative light source instead of fluorescent light which would help in minimizing the quality degradation and SGA- induced toxicity.

Pasteurella multocida produces a capsule composed of different polysaccharides according to the capsular serotype (A, B, D, E, and F). Hyaluronic acid (HA) is a component of certain capsular types of this bacterium, especially capsular type A. Previously, 2 HA biosynthetic genes from a capsular type A strain were studied for the industrial-scale improvement of HA production. Molecular comparison of these genes across different capsular serotypes of P multocida has not been reported. This study aimed to compare 8 HA biosynthetic genes ( pgi, pgm, galU, hyaC, glmS, glmM, glmU, and hyaD) of 22 P multocida strains (A:B:D:F = 6:6:6:4) with those of other organisms using sequence and structural bioinformatics analyses. These 8 genes showed a high level of within-species similarity (98%-99%) compared with other organisms. Only the last gene of 4 strains with capsular type F (HN07, PM70, HNF01, and HNF02) significantly differed from those of other strains (82%). Analysis of amino acid patterns together with phylogenetic results showed that the HA biosynthetic genes of the type A were closely related within the group. The genes in the capsular type F strain were notably similar to those of the capsular type A strain. Protein structural analysis supported structural similarities of the encoded enzymes between the strains of capsular types A, B, D, and F, except for the Pgm, GlmS, GlmU, and HyaD proteins. Our bioinformatics analytic workflow proposed that variations observed within these genes could be useful for genetic engineering–based improvement of hyaluronic acid–producing enzymes.

Covering: 1960s to end of August 2018 Plant glandular trichomes (GTs) are adaptive structures that are well known as "phytochemical factories" due to their impressive capacity to biosynthesize and store large quantities of specialized natural products. The natural products in GTs are chemically diverse and mostly function as defense chemicals, therefore GTs are frequently regarded as "the first defense line" of plants against biotic and abiotic stresses. More importantly, many GT natural products are commercially desirable, thanks to their significant biological activities, thus attracting extensive interest in their biosynthesis. Consequently, it is well known that plant GTs are not only important reservoirs of biologically active natural products but are also a valuable bank of novel biosynthetic genes and enzymes. The non-volatile or oxygenated natural products in plant GTs, which need longer biosynthetic pathways and more energy from the plants, are of particular interest due to their more extensive biological activities and high commercial value. This review mainly focuses on these non-volatile natural products in plant GTs, including their chemistry, biological activities and biosynthesis. The methods employed for investigating natural products and their biosynthesis in plant GTs are also comprehensively discussed.

Covering: up to 2018 Ovothiols are sulfur-containing natural products biosynthesized by marine invertebrates, microalgae, and bacteria. These compounds are characterized by unique chemical properties suggestive of numerous cellular functions. For example, ovothiols may be cytoprotectants against oxidative stress, serve as building blocks of more complex structures and may act as molecular messengers for inter- and intracellular signaling. Detailed understanding of ovothiol physiological role in marine organisms may unearth novel concepts in cellular redox biochemistry and highlight the therapeutic potential of this antioxidant. The recent discovery of ovothiol biosynthetic genes has paved the way for a systematic investigation of ovothiol-modulated cellular processes. In this highlight we review the early research on ovothiol and we discuss key questions that may now be addressed using genome-based approaches. This highlight article provides an overview of recent progress towards elucidating the biosynthesis, function and potential application of ovothiols.

Seed germination plays important roles in the establishment of seedlings and their subsequent growth; however, seed germination is inhibited by salinity, and the inhibitory mechanism remains elusive. Our results indicate that NaCl treatment inhibits rice seed germination by decreasing the contents of bioactive gibberellins (GAs), such as GA1 and GA4, and that this inhibition can be rescued by exogenous bioactive GA application. To explore the mechanism of bioactive GA deficiency, the effect of NaCl on GA metabolic gene expression was investigated, revealing that expression of both GA biosynthetic genes and GA-inactivated genes was up-regulated by NaCl treatment. These results suggest that NaCl-induced bioactive GA deficiency is caused by up-regulated expression of GA-inactivated genes, and the up-regulated expression of GA biosynthetic genes might be a consequence of negative feedback regulation of the bioactive GA deficiency. Moreover, we provide evidence that NaCl-induced bioactive GA deficiency inhibits rice seed germination by decreasing α-amylase activity via down-regulation of α-amylase gene expression. Additionally, exogenous bioactive GA rescues NaCl-inhibited seed germination by enhancing α-amylase activity. Thus, NaCl treatment reduces bioactive GA content through promotion of bioactive GA inactivation, which in turn inhibits rice seed germination by decreasing α-amylase activity via down-regulation of α-amylase gene expression.

Covering: 2006 to 2016 The computational mining of genomes has become an important part in the discovery of novel natural products as drug leads. Thousands of bacterial genome sequences are publically available these days containing an even larger number and diversity of secondary metabolite gene clusters that await linkage to their encoded natural products. With the development of high-throughput sequencing methods and the wealth of DNA data available, a variety of genome mining methods and tools have been developed to guide discovery and characterisation of these compounds. This article reviews the development of these computational approaches during the last decade and shows how the revolution of next generation sequencing methods has led to an evolution of various genome mining approaches, techniques and tools. After a short introduction and brief overview of important milestones, this article will focus on the different approaches of mining genomes for secondary metabolites, from detecting biosynthetic genes to resistance based methods and “evo-mining” strategies including a short evaluation of the impact of the development of genome mining methods and tools on the field of natural products and microbial ecology.

Amaryllidaceae alkaloids are an example of the vast diversity of secondary metabolites with great therapeutic promise. The identification of novel compounds in this group with over 300 known structures continues to be an area of active study. The recent identification of norbelladine 4′-O-methyltransferase (N4OMT), an Amaryllidaceae alkaloid biosynthetic enzyme, and the assembly of transcriptomes for Narcissus sp. aff. pseudonarcissus and Lycoris aurea highlight the potential for discovery of Amaryllidaceae alkaloid biosynthetic genes with new technologies. Recent technical advances of interest include those in enzymology, next generation sequencing, genetic modification, nuclear magnetic resonance spectroscopy (NMR), and mass spectrometry (MS).

Apples with red fruit flesh are desirable for breeding because they contain more health-beneficial compounds. That color trait is also considered more attractive to consumer. To obtain experimental evidence about the molecular mechanism for this pigmentation, we investigated how the practice of bagging fruit might affect levels of phenolic compounds as well as the expression of anthocyanin biosynthetic and regulatory genes. Two red-fleshed varieties from Xinjiang, P.R. China, were studied – ‘Xiahongrou’ and ‘No.1 Hongxun’. HPLC analysis showed that the contents of anthocyanins in both peel and flesh were decreased by bagging, but were increased again when fruits were re-exposed to sunlight. Except for F3H in ‘No.1 Hongxun’, transcription levels for most anthocyanin biosynthetic genes in the flesh were enhanced after bag removal. MYB10 is a key transcriptional factor in the anthocyanin pathway of these two apple varieties, particularly in the flesh. These results suggest that the development of red coloring when bags are removed could be a result of light-induced coordinated expression by MYB10 and anthocyanin biosynthetic genes in red-fleshed apples.

Withanolides are highly oxygenated natural products. These C28 steroids with ergostane-based skeletons functionalized at C-22 and C-26 form six-membered δ-lactone rings. Withanolides containing a δ-lactone side chain often occur in Solanaceae and have a variety of biological activities because of their complicated structures. Characteristic spectroscopic behaviors and biosynthesis of withanolides are conducive to their structural elucidation and “biomimetic synthesis”, respectively. However, the last review to summarize their spectroscopic features and biosynthesis was in 1996. Since then, many withanolides with novel structures have been described by their spectra with biosynthesis investigated with many bioassays. This review surveys δ-lactone withanolides and emphasizes their spectral features, configurations and biosynthetic genes. The period reviewed includes through January 2014. We also include phytochemical species.