In this review, we address recent advances made in pathway engineering, directed evolution, and systems/synthetic biology approaches employed in the production and modification of flavonoids from microbial cells. The review is divided into two major parts. In the first, various metabolic engineering and system/synthetic biology approaches used for production of flavonoids and derivatives are discussed broadly. All the manipulations/engineering accomplished on the microorganisms since 2000 are described in detail along with the biosynthetic pathway enzymes, their sources, structures of the compounds, and yield of each product. In the second part of the review, post-modifications of flavonoids by four major reactions, namely glycosylations, methylations, hydroxylations and prenylations using recombinant strains are described.

To assess the effect of sleep on functional residual capacity (FRC) in normal subjects and asthmatic patients, 10 adult subjects (5 asthmatic patients with nocturnal worsening, 5 normal controls) were monitored overnight in a horizontal volume-displacement body plethysmograph. With the use of a single inspiratory occlusion technique, we determined that when supine and awake, asthmatic patients were hyperinflated relative to normal controls (FRC = 3.46 +/- 0.18 and 2.95 +/- 0.13 liters, respectively; P less than 0.05). During sleep FRC decreased in both groups, but the decrease was significantly greater in asthmatic patients such that during rapid-eye-movement (REM) sleep FRC was equivalent between the asthmatic and normal groups (FRC = 2.46 +/- 0.23 and 2.45 +/- 0.09 liters, respectively). Specific pulmonary conductance decreased progressively and significantly in the asthmatic patients during the night, falling from 0.047 +/- 0.007 to 0.018 +/- 0.002 cmH2O-1.s-1 (P less than 0.01). There was a significant linear relationship through the night between FRC and pulmonary conductance in only two of the five asthmatic patients (r = 0.55 and 0.65, respectively). We conclude that 1) FRC falls during sleep in both normal subjects and asthmatic patients, 2) the hyperinflation observed in awake asthmatic patients is diminished during non-REM sleep and eliminated during REM sleep, and 3) sleep-associated reductions in FRC may contribute to but do not account for all the nocturnal increase in airflow resistance observed in asthmatic patients with nocturnal worsening.

A UDP-glycosyltransferase from Bacillus licheniformis was exploited for the glycosylation of phloretin. The in vitro glycosylation reaction confirmed the production of five phloretin glucosides, including three novel glucosides. Consequently, we demonstrated the application of the same glycosyltransferase for the efficient whole-cell biocatalysis of phloretin in engineered Escherichia coli. P hloretin is a dihydrochalcone, an intermediate of the biosynthetic pathway of flavonoids in plants, which is abundantly present in the peel of apple (1, 2) and in strawberries (3). They occur in different glycosidic forms, such as naringin dihydrochalcone, phlorizin, and phloretin-4=-O-glucoside, in the different parts of the plants, contributing to various physiological properties of the plants, as well as to their color. Phloretin and its glycosides have been determined to have beneficial biological activities. Studies have uncovered that phloretin has inhibitory activity against glucose cotransporter 1 (4, 5), antioxidant activity (6), and activity to suppress the tumor necrosis factor alpha-induced inflammatory response, ameliorate inflammation of the colon, positively affect body weight loss (7), modulate Ca 2ϩ -activated K ϩ channels, and increase endothelial nitric oxide production, which might help to protect against atherosclerosis (8). Importantly, phloretin has other biological functions, like anticarcinogenic (9) and estrogenic activities (10) and inhibition of cardiovascular disease (11, 12). Irrespective of their diverse physiological and pharmacological activities, the use of most of the natural polyphenols as drugs and food additives has been limited because of their water insolubility and low absorbability. Glycosylation enhances the bioavailability and pharmacological properties of compounds by increasing their solubility and stability (13,14). Importantly, the sugar moieties of the glycosides often participate in the specific recognition of their biological targets and help to determine their efficacy in drug development (14, 15). According to the CAZy database (http://www .cazy.org/) (16,17,18), glycosyltransferase family 1 (GT1) proteins contain the UDP-glycosyltransferases that are common in all domains of life (19) and predominantly recognize small molecules as the sugar acceptors. A recent report showed that YjiC, a Bacillus licheniformis UDP-glycosyltransferase that falls in the GT1 family of proteins, can glycosylate at different hydroxyl positions of geldanamycin analogs (20). Here, we report the use of this glycosyltransferase for the biosynthesis of diverse phloretin glucosides in vitro and the subsequent application of YjiC for in vivo production of phloretin glucosides in an Escherichia coli mutant generating a cytoplasmic pool of UDP-glucose, since the YjiC-homologous glycosyltransferases from other Bacillus species were found to have flexible glycosyltransferase activities toward different flavonoid groups of compounds. Moreover, we found that by reversing the glycosylation reaction, the enz...

Quercetin, a flavonol aglycone, is one of the most abundant flavonoids with high medicinal value. The bioavailability and pharmacokinetic properties of quercetin are influenced by the type of sugars attached to the molecule. To efficiently diversify the therapeutic uses of quercetin, Escherichia coli was harnessed as a production factory by the installation of various plant and bacterial UDP-xylose sugar biosynthetic genes. The genes encoding for the UDP-xylose pathway enzymes phosphoglucomutase (nfa44530), glucose-1-phosphate uridylyltransferase (galU), UDP-glucose dehydrogenase (calS8), and UDP-glucuronic acid decarboxylase (calS9) were overexpressed in E. coli BL21 (DE3) along with a glycosyltransferase (arGt-3) from Arabidopsis thaliana. Furthermore, E. coli BL21(DE3)/∆pgi, E. coli BL21(DE3)/∆zwf, E. coli BL21(DE3)/∆pgi∆zwf, and E. coli BL21(DE3)/∆pgi∆zwf∆ushA mutants carrying the aforementioned UDP-xylose sugar biosynthetic genes and glycosyltransferase and the galU-integrated E. coli BL21(DE3)/∆pgi host harboring only calS8, calS9, and arGt-3 were constructed to enhance whole-cell bioconversion of exogeneously supplied quercetin into 3-O-xylosyl quercetin. Here, we report the highest production of 3-O-xylosyl quercetin with E. coli BL21 (DE3)/∆pgi∆zwf∆ushA carrying UDP-xylose sugar biosynthetic genes and glycosyltransferase. The maximum concentration of 3-O-xylosyl quercetin achieved was 23.78 mg/L (54.75 μM), representing 54.75 % bioconversion, which was an ~4.8-fold higher bioconversion than that shown by E. coli BL21 (DE3) with the same set of genes when the reaction was carried out in 5-mL culture tubes with 100 μM quercetin under optimized conditions. Bioconversion was further improved by 98 % when the reaction was scaled up in a 3-L fermentor at 36 h.

Several photocatalytic nanoparticles are synthesized and studied for potential application for the degradation of organic and biological wastes. Although these materials degrade organic compounds by advance oxidation process, the exact mechanisms of microbial decontamination remains partially known. Understanding the real mechanisms of these materials for microbial cell death and growth inhibition helps to fabricate more efficient semiconductor photocatalyst for large-scale decontamination of environmental wastewater or industries and hospitals/biomedical labs generating highly pathogenic bacteria and toxic molecules containing liquid waste by designing a reactor. Recent studies on microbial decontamination by photocatalytic nanoparticles and their possible mechanisms of action is highlighted with examples in this mini review.

The very well-known bioactive natural product, resveratrol (3,5,4′-trihydroxystilbene), is a highly studied secondary metabolite produced by several plants, particularly grapes, passion fruit, white tea, and berries. It is in high demand not only because of its wide range of biological activities against various kinds of cardiovascular and nerve-related diseases, but also as important ingredients in pharmaceuticals and nutritional supplements. Due to its very low content in plants, multi-step isolation and purification processes, and environmental and chemical hazards issues, resveratrol extraction from plants is difficult, time consuming, impracticable, and unsustainable. Therefore, microbial hosts, such as Escherichia coli, Saccharomyces cerevisiae, and Corynebacterium glutamicum, are commonly used as an alternative production source by improvising resveratrol biosynthetic genes in them. The biosynthesis genes are rewired applying combinatorial biosynthetic systems, including metabolic engineering and synthetic biology, while optimizing the various production processes. The native biosynthesis of resveratrol is not present in microbes, which are easy to manipulate genetically, so the use of microbial hosts is increasing these days. This review will mainly focus on the recent biotechnological advances for the production of resveratrol, including the various strategies used to produce its chemically diverse derivatives.

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We report the production of astragalin (AST) from regiospecific modifications of naringenin (NRN) in Escherichia coli BL21(DE3). The exogenously supplied NRN was converted into dihydrokaempferol (DHK) and then kaempferol (KMF) in the presence of flavanone-3-hydroxylase (f3h) and flavonone synthase (fls1) from Arabidopsis thaliana, respectively. KMF was further modified to produce AST by 3-O-glucosylation utilizing the endogeneous UDP-glucose in presence of UGT78K1 from Glycine max. To increase the intracellular UDP-glucose concentration by channeling the carbon flux toward UDP-glucose at the branch point of glucose-6-phosphate (G6P), the chromosomal glucose phosphate isomerase (pgi) and D-glucose-6-phosphate dehydrogenase (zwf) were knocked-out in E. coli BL21(DE3). The two enzymes directly involved in the synthesis of UDP-glucose from G6P, phosphoglucomutase (nfa44530) from Nocardia farcinia and glucose-1-phosphate uridylyltransferase (galU) from E. coli K12 were overexpressed, which successfully diverted the carbon flow from glycolysis to the synthesis of UDP-glucose. Furthermore, to prevent the dissociation of UDP-glucose into UDP and glucose, the UDP-glucose hydrolase (ushA) was deleted. The E. coli ΔpgiΔzwfΔushA mutant harboring the UDP-glucose biosynthetic pathway and the aforementioned genes for the regiospecific glucosylation produced 109.3 mg/L (244 µM) of AST representing 48.8% conversion from 500 µM of NRN in 60 h without any supplementation of extracellular UDP-glucose.