In modern human civilization, the demand for lipids has become upswing for several purposes: nutritional supplements, the production of foods, surfactants, lubricants, and biofuels. With the gradual rise in population, shortage, and deterioration of arable land due to anthropogenic activities, traditional lipids production methods alone cannot conciliate future demand. Various microbial genera related to algae, bacteria, fungi, and yeast can synthesize and accumulate lipids in their bodies. Currently, microbial lipids have emerged as a sustainable successor of plant-derived lipids. However, the commercial scale production of microbial lipids faces some problems such as inadequate level of lipid accumulation in the microbial cells, lipid extraction, and operational cost associated with microbial cultivation. Thus, there is an urgent need to construct oleaginous microbes with modified, preferable features. With the modern biotechnological tools, the insights of the complex microbial genetic makeup and metabolic pathways become explored, allowing various genetic engineering (GE) and metabolic engineering (ME) approaches to develop microbes of desired lipid production abilities with the required lipid profile and physiological qualities. The current review mainly deals with the basic lipid metabolic pathways run in the various groups of microbes, properties of lipids they synthesized, state-of-the-art GE and ME techniques useful for the lipid overproduction in oleaginous microbes, and explain challenging futuristic developmental directions.
Currently, extreme weather events caused by climate change, such as heat waves, drought, frost, and heavy precipitation, have become a threat to agriculture by detrimentally affecting plant productivity and quality. The overuse of synthetic fertilizers is another major concern damaging the soil quality and water and air quality. In this regard, biostimulants could be a promising and potent solution to address these environmental concerns and meet the need for developing sustainable and green modern agriculture. Biostimulants that are primarily composed of natural substances and/or microorganisms can be broadly divided into non-microbial and microbial categories. In this review, the applications of the main types of biostimulants to plant growth and development are discussed, and the possible associated mechanisms of action are described as well. Furthermore, the current status and challenges relating to commercialization and large-scale implementation under changing climate conditions are covered. Overall, this review article could offer insights and knowledge of biostimulants’ uses in agriculture for both academia and industrial sectors.
Sai Mod/Poro Apong and Rohi are fermented indigenous rice beers of the Mising tribe and Kacharis of Northeast India. The rice beer has its own significance as it contains various herbs that add flavor and medicinal properties to the product. In the current study, the overall acceptability of Rohi mod and Sai mod was recorded to be 4.25 ± 0.65 and 5.5 ± 0.41, respectively. Both versions of the beer have significant variations in terms of physicochemical properties and are recorded as (pH: 3.86 ± 0.43 and 3.72 ± 0.22; Moisture content: 85.5% and 89.33%; Alcohol content: 26% v/v and 35% v/v; volatile acidity: 1.23 ± 0.08 mg/ml and 1.08 ± 0.18; Total soluble sugar: 2.44 ± 0.21 µg/ml and 2.5 ± 0.11 µg/ml; Total soluble protein: 0.49 ± 0.02 µg/ml and 0.51 ± 0.05 µg/ml; Crude fat: 0.159 ± 0.04 µg/ml and 0.154 ± 0.02 µg/ml) for Sai and Rohi mods respectively. DPPH scavenging activity (%) and phenolic content were found to be higher in Rohi and recorded as 75.15 ± 0.82 and 550 ± 1.563 GAE/l; however, for Sai, the value was recorded as 63.24 ± 0.31 and 535 ± 2.212, respectively. In contrast, flavonoid content (mg catechin/l) was higher in Sai in comparison to Rohi and was found to be 68.23 ± 1.210 and 48.50 ± 2.002, respectively. The mineral content of Rohi and Sai mods showed a difference. Cadmium was not recorded in either of them. However, the values of other minerals (chromium, copper, nickel, and zinc) were recorded in ppm in the range of 4.7196 (zinc) for Rohi to 0.0628 (chromium) for Sai. FTIR analysis was also employed to study the conformational properties of Sai and Rohi mods. It is believed that the present study will give new insights on both the research and industrial scales in the near future.
Synthetic pigments pose toxicity and harmful impacts on humans and environment. In this context, the exploration of microbial sources for producing natural pigments that are useful in various industrial applications is gaining prominence. Microbial pigments give nature its color and are a novel class of chemicals with diversity of biological functions including antibacterial, anti-mycotic, antimalarial, immuno-modulating, and anticancer properties. Microbial pigment sources have been acknowledged to be extraordinarily valuable and are in great demand, attributable to their varied uses in the food and cosmetic industries, textile, and pharmaceutical industries. Prodigiosin has recently been suggested as the most potential choice and promising biomolecule with multifaceted applications, including antimicrobial, immunosuppressive, antimalarial, antineoplastic, and anticancer properties. Prodigiosin has recently received a renewed interest owing to its reported characteristics of having profound biological activities. The focus of this review article is the most important qualities and prospective uses of prodigiosin pigment.
Herbs are rich in polyphenols compounds that are often added to beer to boost its antioxidant ability. To prove herbs can efficiently enhance the antioxidant ability of beer, the total polyphenols and flavonoids content, the distribution of phenolics profile, and antioxidant ability in commercial herbal beers were analyzed in this work. The results showed that most herbal beers had stronger antioxidant ability than conventional beers. The total polyphenols content in most herbal beers (250.8–1063.5 mg/L) was higher than that in conventional beers (179.2–308.4 mg/L), especially free phenolic acids (3.151–26.469 mg/L) such as gallic acid (0.248–2.853 mg/L), chlorogenic acid (0.422–6.420 mg/L), vanillic acid (0.567–10.256 mg/L), and caffeic acid (0.422–10.093 mg/L), which can explain the strong antioxidant ability of herbal beers to a certain extent. Among them, roselle beer (ROL) and jasmine beer (JAS) performed the strongest antioxidant ability. Furthermore, according to the distribution of phenolics profile, it was found that ROL and JAS had highest content of flavonoids including catechol, epicatechin, and quercetin compared with all beers. This was an important reason why ROL and JAS had the strongest antioxidant ability. In a word, the addition of herbs in beer brewing can effectively improve the content of polyphenols and antioxidant ability of beer, especially the herbs rich in flavonoids have the most significant contribution to the antioxidant ability of beer, which is helpful to improve beer quality.
Artificial Intelligence (AI) has the potential to dramatically change the field of healthcare and nutrition by imitating human cognitive processes. This field involves smart machine-based applications, such as Machine Learning (ML), neural networks, and natural language processing to tackle and solve various issues. The current study’s purpose is to highlight specific AI-based applications that are currently being employed in the fields of nutrition and healthcare. The published data from various search engines, such as PubMed/Medline, Google Scholar, Scopus, Web of Science, and Science Direct, were used for collecting the relevant data. The study depicts that there are several AI-based approaches and methods available for by improving diagnosis and treatment, lowering costs, and increasing access to healthcare facilities. Although AI cannot replace the personal touch, empathy, and emotional support provided by healthcare professionals. These approach assistances expanding rapidly are of great use. However, it is crucial to be careful and make sure that moral considerations are given top priority.
Nicotinamide mononucleotide (NMN), as a healthcare product, plays positive effects on the human body. In this study, an engineered Escherichia coli (E. coli) strain was constructed to efficiently produce NMN from glucose and nicotinamide (NAM) through whole-cell biotransformation. First, a highly active nicotinamide phosphoribosyltransferase (NAMPT/NadV) from Vibrio phage KVP40 was screened, which showed the best productivity in catalyzing phosphoribosyl pyrophosphate (PRPP) and NAM to NMN in E. coli. Next, enzymatic activity of E. coli ribose-phosphate diphosphokinase (EcPRS) was improved and its expression level was optimized, which increased the supply of precursor PRPP. The results showed that mutant EcPRSD115S had optimal activity and that the promoter and RBS combination (PJ23119 and RBSIII) optimized for EcPRSD115S further accumulated NMN. With the systematic modification, such as the deletion of pfkA and pncC genes, and the expression of Cgzwf A243T and Cggnd S361F genes from Corynebacterium glutamicum, the biosynthesis of NMN was improved. Finally, NMN production was further increased after the introduction of NAM and NMN transporters. The strain ZY17 achieved NMN production of 1.61 ± 0.02 g/L in the shake flask level (OD600 = 10), and 13.3 ± 0.35 g/L in the 2-L bioreactor with a productivity of 1.11 g/(L.h) and an NAM conversion (α) of 85.3%. In conclusion, we propose a strategy that can effectively enhance the production of NMN in E. coli, which may bring new ideas for the biosynthesis of other nucleotide compounds.
As a vitamin E isomer, δ-tocotrienol has attracted much attention owing to its rich biological activities for human health, especially anticancer activities. Microbial biosynthesis constitutes a promising strategy of δ-tocotrienol production owing to its economic and environmental advantages. In this study, we employed modular engineering strategies to reconstruct and optimize a de novo δ-tocotrienol biosynthesis pathway in Saccharomyces cerevisiae. Subsequently, the rate-limiting steps were identified and eliminated and the key enzymes were assembled in the δ-tocotrienol biosynthesis module to develop a substrate channeling and improve their catalytic efficiency. Furthermore, the shikimate and δ-tocotrienol biosynthesis modules were optimized via combination strategies to further increase δ-tocotrienol production, following which the δ-tocotrienol titer reached 1455.5 μg/L. Finally, overexpression of the endogenous transporter PDR11 and two-phase extraction fermentation were employed for δ-tocotrienol production, yielding up to 3262.2 μg/L δ-tocotrienol. Thus, the findings of this study demonstrate the possibility of efficient δ-tocotrienol biosynthesis in S. cerevisiae.
Elevated O-GlcNAcylation has been shown to be closely correlated with the occurrence and development of cancer, and inhibiting O-GlcNAc transferase (OGT) activity was demonstrated as a potential tumor treatment strategy. However, the development of pharmacological OGT inhibitors still faces challenges, such as low affinity and poor selectivity. Considering to OGT preferences for the sequence of its peptidic substrates, we herein integrated molecular dynamics simulation approaches to give deep insights into the binding behaviors between OGT and its peptidic substrate ZO3S1, and discussed the unfavorable inter-residue contacts inside the binding pocket, especially between H509 of OGT and S15 of the peptide, upon temperature increase. Removing this unfavorable contact from the peptide (ZO3S1 with S15A mutation) was shown to be able to increase its interaction with OGT, which was manifested by the enhanced OGT activity against this peptide. The pseudo-substrate peptide (ZO3S1 with S13A and S15A mutations) inhibited the activity of purified OGT with an IC50 of 192.9 μM and it can also inhibit the total O-GlcNAcylation in cancer cell lines in a concentration-dependent manner. Our results provided useful models and basis for further rational optimization of selective OGT inhibitors based on the dynamic interactions of OGT and its peptidic substrates.
Escherichia coli contains 12 chaperone-usher operons, including 64 genes, used for biosynthesis and assembly of various fimbriae which consume a lot of energy and material. In this study, each of the 12 operons was deleted in an L-threonine-producing E. coli strain TWF001, and the resulting 12 deletion mutants produced more L-threonine than TWF001 after 16 or 24 h cultivation. Therefore, the 12 chaperone-usher operons were deleted in different combinations, resulting in 11 strain mutants which lack at least 2 operons. The cell growth and L-threonine production of these 11 mutants were determined. Among these 11 mutants, TWK021 in which 10 chaperone-usher operons were deleted, showed the highest L-threonine production. TWK021 produced 15.75 g L-threonine from 40 g glucose after 36 h cultivation. The conversion rate of glucose to L-threonine reached 0.394 g/g in TWK021, which is 32.2% higher than the control strain TWF001. These results suggest that the fimbria lacking E. coli TWK021 is a good host for efficient production of L-threonine.
DNA scaffold that enhances the spatial proximity of enzymes and the local concentration of intermediates, is promising tools in optimizing heterologous metabolic pathways for target product biosynthesis. Here, we display the utility of a DNA scaffold system for the production of ectoine in E. coli MWZ003. Three fused enzymes EctA-ZFa, EctB-ZFb, and EctC-ZFc were firstly constructed by fusing enzymes of ectoine synthesis pathway with corresponding zinc finger domains. The copy number of the plasmid-expressing fusions was adapted by substitution of different replicons. Furthermore, a series of modifications were carried out on the DNA scaffold system through optimizing the spacer between enzyme binding sites, the binding direction of fusion enzymes, the repeating unit of DNA scaffolds, the stoichiometric ratio of enzyme binding sites, and the expression level of the rate-limiting enzyme. The optimized DNA scaffold system in the plasmid pFV30 involving use of pMB1 replicon, reverse binding, 11-bp spacer, 4 repeating units, stoichiometric ratio (1:2:2), and enhanced expression of EctB-ZFb increased the ectoine titer and yield, respectively, to 22.79 g/L and 0.65 g/g glucose with increase by 92% compared with that of the control strain. The post-translational strategy based on DNA scaffold was efficient in promoting heterologous synthesis of ectoine, which could also be used in combination with other genetic engineering tools.
Chromite mine soil is highly contaminated with different heavy metals like chromium, cadmium, lead, and arsenic. In the present study, 42 numbers of metal-tolerant bacteria (Cr-14, Cd-9, Pb-11, and As-8) were isolated from three soil samples collected from chromite mine soils of Sukinda, Odisha using nutrient broth medium supplemented with 50 mg/L of each of the above four metals. The isolated metal-tolerant bacteria were screened using increased above mentioned metal ion concentrations (50, 100 ppm, 200, 500, 700, and 1000 mg/L) to know the maximum resistance of the bacteria towards a particular metal. Out of the 42 predominant bacterial isolates, 4 bacteria (CrS2C7, PbS1M1, CdS2M1 and AsS1M2) showing high resistance to respective metal ions were subjected to biochemical characterization followed by 16S rRNA sequencing and identified as Pseudomonas argentinensis (CrS2C7), Bacillus subtilis (PbS1M1), Bacillus thuringiensis (CdS2M1), and Bacillus fungorum (AsS1M2). Further, the Cr+6 biosorption potential of these 4 bacteria (CrS2C7, PbS1M1, AsS1M2, and CdS2M1) were evaluated using 100 mg/L K2Cr2O7 metal solution. Out of the above four isolates, P. argentinensis showed highest biosorption of 73.4% for Cr+6 metal ion solution The present finding of Cr+6 biosorption by P. argentinensis indicates that it can be used as an effective biosorbent for biosorption of Cr+6 from chromate contaminated aqueous environment.
Recently, consumers have demanded safe, nutritious, and healthy food. Due to rapid heat transfer, microwave treatment (MT) preserves nutritional quality and provides safe food. In this investigation, the effect of Microwave heating (MH) treatment was analysed on the bioactive components and quality characteristics of buransh (Rhododendron arboretum) flower squash. The fresh squash (TSS 19.6°Brix) was prepared by adjusting TSS to 3, 4, and 5°Brix by using RO water, and microwave treatment (MT) was given to the prepared squash as per the Box Behnken design (BBD). Process optimization was done using Design Expert 13 software. Microwave Power (MP), time, and total soluble solids (TSS) as process parameters showed significant (p < 0.05) effects on responses, namely pH, TSS, total phenolic content (TPC) and total flavonoid content (TFC). The optimized values at optimum process parameters (300 W, 30 s and 3°Brix) were found to be 3.79 for pH, 2.395°Brix for TSS, 44.28 mg GAE/100 mL for TPC and 132.159 mg QE/100 mL for TFC. The experimental values were found to be in accordance with the predicted values, having error (%) 0.5249, 3.96, 2.75, and 1.55 for pH, TSS, TPC, and TFC respectively. It was concluded that microwave heating can be used for the treatment of buransh (Rhododendron arboretum) flower squash with superior retention of nutritional quality.
Staphylococcus aureus is one of the most common bacteria in the human skin and causes various severe infections. Methicillin-resistant Staphylococcus aureus (MRSA) has raised significant concerns and challenges because they are difficult to kill due to their ability to resist many antibiotics. In the present study, a blue light-emitting diode with a wavelength of 405 nm was designed and applied for inactivating S. aureus. We investigated the dependence of the S. aureus inactivation rate on the irradiation factors, including time, distance, and energy dose. The results indicated that irradiation time and distance were observed to significantly affect the growth of S. aureus. In particular, the energy dose of 322.2 J/cm2 at a distance of 5 cm for 35 min was recommended to inactivate S. aureus growth completely. The study also proposes the DNA self-repair mechanism causing the delaying time during inactivating S. aureus. Therefore, energy dose is a reliable parameter for designing a single-factor experiment to optimize the irradiation time and distance to obtain a suitable range of values. The comparison between red and blue LED also confirmed the ability of the blue LED to inhibit bacterial growth, while red LED enhances the growth of bacteria. Briefly, our research suggested that blue LED at the wavelength of 405 nm has the potential to be applied in clinical hygiene and food processing as an antimicrobial technique to prevent the development of antibiotic resistance.
The conventional methods of nanoparticles synthesis led to the production of highly toxic by-products and the use of toxic chemicals that are highly expensive in nature. Thus, the recent past has witnessed a surge in green synthesis of nanoparticles as a sustainable alternative. The present study outlines the biogenic silver nanoparticles (Ag-NPs) synthesis from an aqueous extract of Chlorella minutissima. The effect of certain parameters such as the reaction mixture’s pH and precursor metal solution to algal extract ratios were explored and optimized. The UV spectrophotometric analysis of Ag-NPs gave surface plasmon response maximally at 426 nm. The developed Ag-NPs were characterized using zeta potential, indicating their high stability (-21.2 mV) with a mean diameter of 73.13 nm. Results from field emission-scanning electron microscopy (FE-SEM) showed that the particles were spherical in shape. Ag-NPs synthesized using Chlorella minutissima extract could significantly inhibit the growth of both Gram-positive and Gram-negative bacterial species. The study highlights that using C. minutissima extract for Ag-NPs synthesis is a convenient and fast process for controlling the growth of Gram-positive as well as Gram-negative bacteria.
Cyclic-di-GMP (c-di-GMP) is a ubiquitous signaling molecule in many microorganisms that orchestrates genetic regulation during the transition between a sessile and motile lifestyle. The intracellular levels of c-di-GMP are stringently monitored by two enzymes, diguanylate cyclases (DGCs) consisting of GGDEF domain and phosphodiesterases (PDEs) with EAL or HD-GYP domain. This study scanned the probiotic strain Escherichia coli Nissle 1917 (EcN) genome for genes encoding for GGDEF and EAL domain-containing proteins. A total of 30 genes coding for proteins with canonical GGDEF, EAL, or both domains were identified. The expression of these genes in EcN during host intestinal cell colonization is yet to be characterized. Herein, the transcript levels of the 30 predicted genes implicated in c-di-GMP metabolism were analyzed in EcN during in vitro colonization of intestinal epithelial cells, Caco-2. The expression of two genes, dgcZ and sfaY, which play a vital role in the initial attachment to the cell surface and stress response, was highly up-regulated in EcN after 2 h of incubation with Caco-2 cells. Further, an affinity pull-down assay identified 53 c-di-GMP binding proteins. The in-depth genetic characterization will provide further insights into the c-di-GMP-mediated regulatory mechanisms during host colonization by the probiotic strains.
D-p-hydroxyphenylglycine (D-HPG) is an important intermediate in the pharmaceutical industry, and it is commonly synthesized by cascading D-hydantoinase (DHase) and D-carbamoylase (DCase). In this study, the stability of DCase was identified as the main problem that limits its application. Therefore, the complexed structure of AkDCase (DCase from the Agrobacterium sp. strain KNK712) with the substrate N-carbamoyl-D-p-hydroxyphenylglycine (CpHPG) (with 2.52 Å resolution) and catalytic mechanism were resolved. Based on the catalytic mechanism and electrostatic stabilization, salt bridge engineering was adopted to improve AkDCase thermostability. The best variant, AkDCaseD30A, increased the Tm by 2.91 °C and half-life (t1/2) at 40 and 60 °C by 18.43 h and 23.21 min, respectively. After AkDCaseD30A was assembled with GsDHase (DHase from Geobacillus stearothermophilus SD-1) in a single Escherichia coli cell, the recombinant strain could produce 29.53 g/L D-HPG within 12 h, with a 97% conversion and a 2.46 g/(L·h) space–time yield (STY). The titer of D-HPG increased by 40.55% compared to the E. coli cell harboring pETduet-1-AkDCase- GsDHase. The recombinant strain could be used for two cycles. Our research provides a basis for the industrial production of D-HPG.
The human gastrointestinal system is home to billions of different microbial species that coexist peacefully with the host. Age, diet, and environment affect gut microbial structure and diversity. Bacteroidetes and Firmicutes are the main gut microbial groups in humans. Human physiology is greatly affected by gut microbial metabolism and host–microbe interactions. Gestational diabetes mellitus (GDM) women have a different gut microbial makeup. However, whether these microbial changes in the stomach may be detected before the diagnosis of GDM remains unknown. This study aims to study the dynamics of gut microbe in healthy and GDM women and analyze the potential biomarkers for early detection of GDM in pregnant women. In this study, gut microbial variations have been characterized and their relationship is evaluated with GDM development. A large multi-taxon microbial shift between GDM and normal cases was evident, indicating that few microbial disruptions from early to mid-pregnancy were related to the later development of GDM. Results of linear discriminant analysis (LDA) show the variable relative abundance of 13 taxonomic groups (at the family level) between the GDM and normal groups. Moreover, 8 taxonomic groups (at the family level) were found to be present in the GDM group only. The results contribute to the gut microbiota’s potential relevance as a biomarker for the early detection of GDM.
Functional peptides are short aminoacidic sequences that could contribute to human health. Functional peptides possess a wide range of bioactivities. Hence, a growing interest in these kinds of molecules, mainly focused on studying their different production strategies and respective economic and technological challenges, is increasing, where enzymatic hydrolysis is one of the leading strategies for their generation. The aim of this study is to find new sources of fungal proteases with antioxidant peptide production potential. A screening of 17 fungal strains with proteolytic activity was achieved, employing a skimmed milk medium. Six fungal strains were selected to study their proteolytic kinetics profile, and two of them were evaluated for their capacity to produce bioactive peptides from calcium caseinate through enzymatic hydrolysis. ABTS, DPPH, and FRAP assays measured the antioxidant activity of the peptides obtained. Blakeslea trispora and Rhizomucor pusillus were the higher proteolytic strains (1–4 EU/mL), and their crude extracts after caseinate hydrolysis generated an inhibition percentage of ABTS radical between 53.6–65.1%, 33.4–52.4% in DPPH and 496.8–690.8 µM Fe2+/g of lyophilized peptides. Both enzymatic extracts are reliable to be used in the production of antioxidant peptides.
In the current study, the phytochemical composition and biological activity (antimicrobial, antioxidant, antigenotoxic, and antimutagenic) of different leaf extracts of green tea (Camellia Sinensis) were investigated. Maximum total polyphenol content (17.12 ± 0.02), total flavonoid content (16.18 ± 0.03), antioxidant activity (DPPH) to scavenge free radicals (95.9%) and highest antimicrobial activity against Escherichia coli and Staphylococcus aureus with a minimum inhibitory concentration value of 3.12 mg/ml and a 26 mm zone of inhibition were recorded for the methanolic extract. The antigenotoxic potential of various extracts of C. sinensis at a concentration of 1000 µg/ml reduced DNA damaged by 37% against Cd-induced genotoxicity as indicated by tail moment (7.9 ± 0.4) during comet assay. Antimutagenic potential of methanolic extracts showed a mutagenic inhibition of 52% against sodium azide mutagen in TA98 strain (+ S9) during the ames test. Gas chromatography–mass spectrometry analysis of methanolic extract showed the presence of a number of compounds The above findings suggest the potential application of C. sinensis methanol extract in food and the health business, and to meet expanding human requirements.
3′-Sialyllactose (3′-SL) promotes the proliferation of the bifidobacterial population and shapes gut microbiota. Thus, it can be used in the infant formula industry. In this study, an engineered Bacillus subtilis 168 modified by previous research can synthesize Neu5Ac, an important precursor for 3′-SL biosynthesis, named 3D6.2. Firstly, neuA and nst from Neisseria meningitidis were introduced into 3D6.2 to realize the de novo biosynthesis of 3′-SL from glucose and lactose. Subsequently, the 3′-SL synthetic pathway was optimized with a push–pull-restrain strategy. The promoters and ribosome binding site sequences of neuA and nst were optimized to increase the expression of the two enzymes. Moreover, the β-galactosidase gene was deleted to reduce the consumption of competitive lactose. As a result, the 3′-SL titer was found to improve from 3.8 to 344.7 mg/L. Furthermore, several protein scaffolds were chosen to improve the catalytic efficiency of NeuA and Nst, and the ratio of these two enzymes in the synthetic scaffolds was optimized. Thus, a final 3′-SL titer of 1252.1 mg/L was obtained. These results demonstrate the potential for improving 3′-SL production in B. subtilis in the future.
Maltooligosyl trehalose synthase (MTSase) and maltooligosyl trehalose hydrolase (MTHase) are used to produce trehalose, a disaccharide of interest to many different industries, from starch. MTSase and MTHase from Arthrobacter ramosus S34 were first produced using separate Escherichia coli BL21(DE3) strains. The activities obtained in a 3-L fermenter under optimized conditions were 1608.3 U mL−1 and 8766.2 U mL−1, respectively. Then, MTSase and MTHase were co-produced in E. coli BL21(DE3) using a co-expression construct. After optimizing induction conditions, the MTSase and MTHase activities produced by the superior strain reached 1827.4 U mL−1 and 2944.9 U mL−1, respectively. When the co-produced enzymes were used to synthesize trehalose from starch, a conversion rate identical to that achieved using separately produced enzymes (about 67%) was obtained. This is the first describing the co-production of the MTSase and MTHase in a 3-L fermentor. The results represented the highest MTSase production level reported to date, and the MTHase activity from co-production was sufficient for trehalose synthesis. Using co-produced enzymes during trehalose synthesis would lower costs without sacrificing yield. Therefore, this study provided a foundation for the industrial synthesis of trehalose using co-produced enzymes.
A Lactobacillus sp. was screened from various cereal sourdoughs and was designated as Lactobacillus plantarum YXY418 based on the 16S rRNA gene analysis. A putative Lactobacillus plantarum maltogenic amylase, LpMA, was discovered based on computer-aided analysis. Then, its encoding gene (lpma) was expressed in E. coli BL21(DE3). The expressed recombinant LpMA (reLpMA) was efficiently purified to 12.2-fold using the one-step nickel-nitrilotriacetic acid (Ni–NTA) affinity chromatography. The final recovery yield and specific activity of the purified reLpMA were 61% and 36.4 U/mg towards soluble starch, respectively. The purified reLpMA exhibited optimal amylolytic activity towards soluble starch at 45 °C and pH 6.0, with a good pH stability ranging from pH 5.0 to 8.0. Besides, the reLpMA also hydrolyzed soluble starch, β-CD and pullulan to maltose with specific activity of 96.4 SU/mL, 78.2 CU/mL and 2.0 PU/mL, respectively. The reLpMA hydrolytic activity was increased in the presence of metal ions especially Ca2+ and Zn2+, which could be applied to different processing processes. Baking test indicated after 7-day storage, the reLpMA at a dosage of 2000 U/300 g could significantly reduce hardness and chewiness by 29.5% and 26.4%, respectively, compared with the control. Adding reLpMA improved bread quality, increased bread volume and decreased hardness during storage, thus extending its shelf life.
Soybean meal (SM) is an important protein source for animal feed, and the β-conglycinin included is a significant allergen that can cause severe allergic reactions such as diarrhea in piglets and other animals. Fermentation can remove anti-nutritional factors and allergenic proteins, however, it is very difficult to find the fermentation endpoint, because the current protein detection methods are difficult to be applied for the rapid detection. In this study, AuNPs-aptamer sensor was used to evaluate the degradation of β-conglycinin during SM fermentation, and found a synergistic effect between alkaline protease and Lactobacillus plantarum DY6 with the optimum process parameters below: solid–liquid ratio 2, L. plantarum DY6 5%, alkaline protease 500 U/g, molasses 3%, 37 °C for 72 h. The fermented SM reached the 80.75% degradation and the final products possessed with a 1.87-fold increase in total free amino acids. This research validates the feasibility of using aptamer sensor to assess the anti-nutritional factors, and provides data for further large-scale production.
Previously, we have developed an Escherichia coli strain MWZ003/pFT28-ectABC-EclysC*-aspDH-ppc3 in which the titer of ectoine reached 30.37 g/L with a yield of 0.13 g/g glucose after 36 h fed-batch fermentation. In this study, this strain was further modified to improve the production of ectoine. Genes pflB encoding the pyruvate formatelyase, poxB encoding the pyruvate oxidase, adhE encoding the alcohol dehydrogenase, and aroG encoding the 3-deoxy-7-phosphoheptulonate synthase were deleted from MWZ003, resulting in the strain MWL007. Comparing with the control MWZ003/pFT28-ectABC-EclysC*-aspDH-ppc3, ectoine production in MWL007/pFT28-ectABC-EclysC*-aspDH-ppc3 increased 21% with a yield of 0.43 g/g glucose. The gene mscS encoding the small conductance mechanosensitive channel MscS was further deleted in MWL007, resulting in the strain MWL009. Comparing with the control MWZ003/pFT28-ectABC-EclysC*-aspDH-ppc3, ectoine production in MWL009/pFT28-ectABC-EclysC*-aspDH-ppc3 increased 28% with a yield of 0.46 g/g glucose. After the fermentation conditions were optimized, ectoine production in MWL009/pFT28-ectABC-EclysC*-aspDH-ppc3 further increased and the yield reached 0.63 g/g glucose. After 60 h fed-batch fermentation, the titer of ectoine in MWL009/pFT28-ectABC-EclysC*-aspDH-ppc3 reached 34.27 g/L with the yield of 0.34 g/g glucose. These results indicate that ectoine production in MWL009/pFT28-ectABC-EclysC*-aspDH-ppc3 can be improved by accumulation of the key precursors.
The study conducted on the antioxidant, antimicrobial, and anticancer potential of methanol and acetone extracts of two species of cyanobacteria (Lyngbya majuscula SB12-13 and Lyngbya martensiana SBD24) isolated from Odisha coast and identified using morphological and molecular approach. The maximum DPPH and ABTS radical scavenging activity was recorded in L. majuscula and the IC50 value was 251.34 ± 0.96 and 282.24 ± 0.87 μg/ml for L. majuscula and L. martensiana, respectively. The FRAP assay was also higher in L. majuscula (0.762 ± 0.0015) as compared to L. martensiana (0.679 ± 0.0009 µg/ml). The TP (69.18 ± 0.79 μg gallic acid equivalents g−1) and TF (38.21 ± 0.61 μg quersentin equivalents g−1) contents were more in methanol extract. The SOD and CAT activities were more prominent in L. majuscula than in L. martensiana. The antimicrobial activity study exhibited highest zone of inhibition in L. majuscula (23 ± 0.74 mM) as compared to L. martensiana (10 ± 0.77 mM). The FT-IR and GC–MS spectra analysis of methanol extract of L. majuscula and L. martensiana confirmed the presence of functional groups and respective phytocompounds. Further, the cytotoxicity results exhibited decreasing cell viability of HepG2 cells in methanol extract of L. majuscula in a dose-dependent trend without cytotoxicity on normal cells. The results obtained indicated differential characteristic among both species of same genus from which L. majuscula was a potent species and might be a good source for the production of promising bioactive compounds and can be used as an alternative for natural compounds for commercial applications.