This review provides a comprehensive overview of the conformational changes between the open and closed states of loop structures within enzyme molecules and their significance in enzyme engineering. The article begins by introducing the developmental background of protein engineering and the structural characteristics of enzymes, with a particular focus on the pivotal role of loops in enzyme activity, substrate specificity, and environmental adaptability. Through instrumental analysis and computational approaches, the molecular mechanisms underlying loop opening and closing are thoroughly examined, and recent practices of loop engineering based on point mutations, directed evolution, and rational design are systematically summarized. The studies indicate that the dynamic changes of loops are central to influencing catalytic efficiency and specificity, and their modification can significantly optimize enzyme performance. Therefore, the in-depth research in this field not only provides novel perspectives for elucidating the dynamic processes of enzymatic catalysis but also paves innovative pathways for optimizing engineering strategies to develop high-performance artificial enzymes and green biomanufacturing technologies.
The health benefits of rare sugars as substitute sweeteners and their enormous commercial potential across various industries have attracted significant attention. However, their low availability and the limited, costly synthesis methods remain major obstacles to their widespread use. Bacterial exopolysaccharides (EPSs), composed of diverse monosaccharides, offer a sustainable source of rare sugars. In this study, exopolysaccharide—producing bacteria were isolated from the rhizosphere of the leguminous plant Arachis pintoi. Two highly productive strains R3 and R11, produced 5.80 g/L and 6.07 g/L of EPS, respectively. Gas Chromatography–Mass Spectrometry (GC–MS) analysis revealed that rhamnose and glucosamine were the predominant sugar monomers in the EPSs produced by these bacteria. Additionally, other rare sugars, including fucose and arabinose in lower concentrations, were also identified. Besides, alkali media modified the monosaccharide profile of the EPS and substantially increased its production, with yields increasing by 101.07% in R3 and 238.89% in R11. Particularly, when the pH increased, EPS from R11 showed an enhancement in ribose, galacturonic acid, and glucosamine synthesis. According to the analysis of 16S rRNA and phylogenetic tree, the promising EPS producers R3 and R11 were identified as Enterobacter sp. and Klebsiella sp. The EPSs produced by these bacteria demonstrated notable antioxidant capacity. The findings of this study indicate that exopolysaccharides (EPSs) have potential as raw materials for the production of rare sugars applicable in the food, nutraceutical, and pharmaceutical industries. Furthermore, the results underscore the effectiveness of alkaline media in enhancing the yield of EPSs. This finding paves the way for a novel research avenue focused on the production of rare sugars for industrial applications using exopolysaccharides (EPSs) synthesized by Enterobacter sp. and Klebsiella sp.
Recently, microbial polysaccharides have gained significant importance as biomaterials, owing to their health benefits and inherent biocompatibility. Accordingly, the present study focuses on characterizing exopolysaccharides (EPS) from Lactiplantibacillus plantarum E1K2R2, investigating their structure, biocompatibility, antioxidant properties, antitumor effects, and digestibility under simulated saliva and gastrointestinal conditions. The EPS was identified as a heteropolysaccharide primarily composed of galactose and glucose monomers, with a molecular weight ranging from 3.15 × 103 to 3.95 × 105 Da. The measured particle size was 498.7 nm, while its negative zeta potential (− 7.20 mV) pointed to an acidic profile. FTIR analysis further supported this finding by identifying characteristic hydroxyl and carboxyl functional groups at 3273.57 cm−1 and 1646.91 cm−1, respectively. 1H NMR spectroscopy demonstrated that monomers in the EPS were linked by α- glycosidic bonds. Functionally, the EPS displayed dose-dependent antioxidant activity within 0.1–1 mg/mL, equivalent to 0.07–0.17 mM/mL ascorbic acid, with ABTS·+ radical scavenging ranging from 33.63 to 65.63%, demonstrating strong free radical–scavenging potential. In cytotoxicity assays, the EPS moderately inhibited HepG2 cell proliferation (6.49–20.53%) while maintaining 95% viability in normal HUVEC cells at 1 mg/mL after 24 h of incubation, indicating selective cytotoxicity toward cancer cells and good biocompatibility with normal cells. The EPS also resists simulated saliva and gastrointestinal conditions and promotes the growth of some potential probiotic lactic acid bacteria, indicating that digestive conditions do not adversely affect the EPS. These findings suggest that EPS has promising applications due to its antioxidant, antitumor, and prebiotic contexts, supported by its high biocompatibility.
Lacto-N-neotetraose (LNnT), a functional oligosaccharide abundant in human milk, holds significant nutritional and biomedical value. This study engineered the industrial workhorse Corynebacterium glutamicum ATCC13032, which lacks native LNnT biosynthesis capability, through systematic metabolic engineering. Initial strain CL001 was constructed by heterologously expressing LgtA and LgtB from Neisseria meningitidis and LacY from Escherichia coli, achieving 0.084 g/L LNnT. Subsequent introduction of E. coli galactokinase GalK (strain CL002) enhanced production to 0.286 g/L. Metabolic flux optimization through zwf gene knockout to suppress the pentose phosphate pathway (strain CL013) further increased titers to 0.477 g/L. Implementation of an ABC transporter system (strain CL014) elevated LNnT production to 1.05 g/L. Whole-cell catalysis of CL014 for LNnT production was optimized after 54 h cultivation at 37 °C. Substrate optimization established ideal concentrations at 60 mM lactose, 60 mM galactose, and 240 mM glucose. Under these conditions, CL014 achieved 9.12 g/L LNnT production, representing a 108-fold improvement over the initial engineered strain. This work demonstrates the potential of engineered C. glutamicum as an efficient platform for human milk oligosaccharide biosynthesis.
Lacto-N-neotetraose is an abundant human milk oligosaccharide with multifaceted physiological functions. In this study, Escherichia coli MG1655 has been used as the chassis strain for lacto-N-neotetraose production. First, the genes lacz, ugd, gcd, nagB, ushA, setA were deleted in MG1655, and the Neisseria meningitidis genes lgtA and lgtB encoding β-1,3-N-acetylglucosaminyltransferase and β-1,4-galactosyltransferase, respectively, were overexpressed, and the resulting strain WY011/pC-A/pR-B could produce 1.102 g/L lacto-N-neotetraose. Next, the rate-limiting gene overexpression, expression vector screening, and ribosome-binding site strength were optimized, and the resulting strain WY011/pR-BEA1R could produce 2.634 g/L lacto-N-neotetraose. Further optimization of fermentation parameters enabled the engineered strain WY011/pR-BEA1R to produce 3.533 g/L lacto-N-neotetraose, and a whole-cell catalysis process developed by using this strain with the mixed substrates of glycerol, lactose, and galactose boosted the titer to 10.845 g/L, which is the highest reported titer in small-scale systems so far. Ultimately, fed-batch fermentation of the strain WY011/pR-BEA1R in a 2-L bioreactor achieved an LNnT titer of 16.202 g/L.
Enzymatic degradation of polyethylene terephthalate (PET) has garnered attention as a new PET decomposition technology because of its progression under mild conditions. However, the practical application of these enzymes is significantly hindered by their high purification costs. To address this, we developed an Escherichia coli strain with surface-engineered capabilities for efficient PET degradation by expressing PET-degrading enzymes. Several components were considered and optimized for the efficient degradation of PET using this microorganism catalyst. When comparing the expression systems, cells displaying PETase on the surface using the arabinose-inducible system showed a 2.9-fold improvement in PET degradation compared to those using the isopropyl β-d-1-thiogalactopyranoside-inducible system. Furthermore, cells with both surface-displayed FAST-PETase and localized MHETase in the periplasm exhibited a 6.6-fold increase in PET degradation efficiency compared to the initial pETDuet system. Notably, MHETase expression shifted the product profile toward predominantly terephthalic acid (TPA) formation. These results demonstrate that incorporating MHETase into a PET degradation system predominantly yields TPA, suggesting potential applications for efficient PET upcycling into valuable chemicals.
Endo-polygalacturonases are valuable for decomposing pectin into functional oligosaccharides, which exhibit various bioactivities such as prebiotic, anticancer, and antibacterial effects. In this work, a novel endo-polygalacturonase from Penicillium arizonense (Kp-pePGB) was heterologously expressed in Komagataella phaffii. The recombinant enzyme showed optimal activity at 65 °C and pH 5.0, along with broad pH stability (pH 3.5–8.0). Nonetheless, its thermal instability limited its industrial utility. To address this, structure-based rational design was employed to engineer stabilizing mutations. The triple mutant E257C/N286C/D207N exhibited a remarkable 11.6-fold increase in residual activity after incubation at 55 °C for 60 min, without compromising catalytic performance. Molecular dynamics simulations suggested that the enhanced thermostability arose from newly introduced disulfide bonds and optimized electrostatic interactions. Importantly, the mutant efficiently generated pectic oligosaccharides, which significantly promoted the growth of beneficial probiotics including Pediococcus acidilactici, Lactobacillus paracasei, and Lactobacillus plantarum, demonstrating their prebiotic potential. This study not only provides a promising endo-polygalacturonase for the production of prebiotic oligosaccharides under thermally demanding conditions, but also offers an effective engineering strategy for improving the thermostability of other enzymes in the polygalacturonase family.
The genus Sanghuangporus, renowned in China for its traditional use due to its antitumor, antioxidant, and hypoglycemic properties, includes medicinal species with therapeutic potential. However, the metabolic profile of Sanghuangporus quercicola under different cultivation strategies remains poorly understood. Here, we present the first metabolomic comparison of S. quercicola fruiting bodies and mycelia obtained via substitute cultivation, solid-state fermentation, and submerged fermentation. (I) UHPLC-QTOF-MS-based untargeted metabolomics identified 164 metabolites, including phenolics, terpenoids, steroids, and nitrogen-containing compounds, with 57 showing significant differential accumulation. (II) Multivariate analyses (PCA, PLS-DA) and KEGG pathway enrichment revealed that solid-state fermentation enhanced bioactive metabolite accumulation, especially phenolics and terpenoids, whereas substitute cultivation yielded greater chemical diversity in fruiting bodies, suggesting potential for targeted therapeutic development. (III) In vitro assays showed that the solid-state fermentation extract exhibited the strongest α-glucosidase inhibition (95.35%) and moderate α-amylase inhibition (30.50%), comparable to those of acarbose, indicating selective modulation of carbohydrate digestion. These findings highlight cultivation-dependent metabolic reprogramming in S. quercicola and its potential as a source of functional anti-diabetic agents, while encouraging future in vivo validation.
Methanol has emerged as promising single-carbon and sustainable feedstocks with no association with food production. Methanol bioconversion by methylotrophic fermentations has been practiced for decades, but their applications are still limited to natural methylotrophs due to the lack of synthetic biology tools and suitable classis. Bacillus species generally has good resistance and safety, and are potential chassis in industry. Moreover, some Bacillus species are natural methylotrophs, which means that Bacillus species employed as methanol bioconversion classis may be feasible. This review systematically analyzes methanol metabolic pathways in natural methylotrophs, focusing on three critical biochemical processes: methanol oxidation, formaldehyde assimilation, and dissimilation pathways. Additionally, the article provides a comprehensive discussion of applications existing challenges in methylotrophic Bacillus engineering, including those related to methanol oxidation, formaldehyde toxicity, cofactor balance, and fermentation equipment, while proposing potential strategies to address these technical bottlenecks in the construction of Bacillus methanol classis. This review seeks to demonstrate the potential of the Bacillus species in next-generation methanol-based applications and provide a practical reference for researchers selecting methylotrophic strains.
Tannic acid (TA), the secondary metabolite of plant, produced to protect themselves from predators. It forms chelat to precipitate out the dietary as well as extracellular protein molecules. Present study depicted the adaptation strategies of tannase producing Stenotrophomonas maltophilia PKA14 (Accession No.: KY921597) to avail nutrient in TA environment. A thick outer layer of bacteria composed of carbohydrate was observed during TA exposure which gradually become thinner with the production of tannase. Initially, non-reducing sugar concentration was high to protect the cell from TA stress. With time, the non-reducing sugar gradually converted to reducing sugar to supply carbon source to entrapped vegetative cell. Regulation of gallic acid, the product of tannase hydrolysis, using Gal cluster gene were also analysed through in silico molecular docking. This observation could be applicable for tannase producing microbiome study, growth optimization of tannase producing industrial culture and enhancement of industrial production of gallic acid.
Complete illustration of tannic acid stress management by tannase producing bacteria, tannic acid degradation and utilization of glucose and gallic acid.
The enzymatic hydrolysis of gellan gum (GG) using gellan lyase presents an efficient and practical method for generating gellan oligosaccharides (GOSs). GOSs have promising applications due to their diverse biological functions, including beneficial properties and immunoreactivity. They are characterized by low molecular weight, high water solubility, and easy absorption and utilization by the body. In addition, GOSs exhibit properties such as plant-induced disease resistance and enhanced immune activity. However, current degradation methods for producing GOSs remain limited. In this study, recombinant Bacillus sp. GL1-derived gellan lyase was successfully expressed in Pichia pastoris under the control of the AOX1 promoter. DO-stat carbon source of waste mixed sugar used as a substitute for glycerol, feeding strategy was adopted for scale-up cultivation in a 7 L fermenter. The highest dry cell weight (DCW) and enzyme activity reached 68.7 g L−1 and 1473.5 U L−1, respectively. Furthermore, a novel co-feeding fermentation approach was developed to enable the one-step production of GOSs by directly supplementing GG into the fermentation medium. During the induction phase, a co-feeding strategy of methanol and 0.5% GG solution (2:3 ratio) was adopted, achieving a gellan lyase activity of 1728.1 U L−1, which increased 17.3%, compared to the enzyme activity without supplementation of GG. Notably, the co-feeding method not only enhanced enzyme induction efficiency but also selectively generated GOSs with a degree of polymerization (DP) of three. This study provides a direct and simple way to produce oligosaccharides, as well as a novel fermentation method for enhancing gellan lyase production as well as GOSs production simultaneously.
The present study describes the production, statistical optimization and characterization of alkaline protease produced by Penicillium oxalicum JML 15, and evaluation of its application as detergent additive. In One Factor At a Time (OFAT) studies, maximum protease activity was observed on day 6 at pH 9 and 30 °C, with glucose and yeast extract as optimum carbon and nitrogen sources, respectively. The presence of Mg2⁺ facilitated enhanced enzyme activity, and 1% casein supported optimum enzyme production. Statistical optimization using Plackett–Burman design identified glucose and incubation period as two significant factors for high protease yield. Response Surface Methodology was used to maximize protease yield up to 2.4-fold (348.33 U/ml) compared to unoptimized (144.72 U/ml). The partially purified protease showed optimal activity at 50 °C and pH 10, with stability retained up to 60 °C and pH 11, indicating excellent thermal–alkaline tolerance desirable for detergent applications. Enzyme activity was significantly increased in the presence of Mn2⁺ and retained about 80% of its activity at 2.0 M NaCl, indicating considerable halotolerance. The protease remained fully active in commercial detergents such as Ariel and Tide, confirming its compatibility and stability in complex detergent formulations. In stain removal assays, the enzyme achieved complete removal of bloodstains within 10 min, demonstrating its efficiency in degrading proteinaceous stains. These characteristics highlight its strong potential as a bioadditive for detergent formulations and its applicability in industries that require alkaline- and salt-tolerant proteases.
The global food sector faces increasing pressure to adopt sustainable, nutritious, and eco-friendly production systems. Algae have emerged as a promising solution due to their superior biomass productivity, minimal land and freshwater requirements, and rich composition of proteins, lipids, and bioactive compounds. This review examines the nutritional potential of diverse algal species, highlighting their macronutrient and micronutrient profiles. Technological advancements in photobioreactors, cultivation, as well as bioprocessing innovations, have significantly improved algal yield, extraction efficiency, and functional integration into food systems. Algae's role in food innovation is further explored through its application in meat analogues, dairy alternatives, fortified snacks, and beverages. Additionally, the article evaluates the health-promoting effects of algae alongside potential risks, underscoring the necessity for rigorous safety assessments.
Enzymatic depolymerization of polyesters has been established as green recycling strategy to reduce plastic pollution. However, applied on industrially relevant mixed plastic waste, it generates a wide mixture of various oligomers and monomers, complicating downstream processing and monomer recycling. Several Pseudomonas taiwanensis strains have been engineered to grow on plastic monomers while producing valuable aromatics. This enables metabolic funneling of diverse monomers, supporting efficient bio-upcycling. Integrating hydrolysis and monomer conversion into one intensified process would increase the competitiveness of biotechnological upcycling. Therefore, a one-pot process was developed in which hydrolysis of poly(butylene adipate-co-terephthalate) (PBAT) via a cutinase from Humicola insolens (HiC) was coupled with cultivation of P. taiwanensis strains metabolizing the resulting monomers, adipic acid (AA), terephthalic acid (TA) and 1,4-butanediol (BDO). For this purpose, the buffer strength and stirring rate for PBAT-hydrolysis were adjusted for compatibility with cultivation of P. taiwanensis. An impact of various process settings on enzymatic hydrolysis was found with the temperature as main parameter, where enzymatic and microbial conversion conflict. Hence, two consecutive steps were carried out within one reactor—a 24-h hydrolysis at 70 °C, followed by inoculation with Pseudomonas after changing the conditions to 30 °C. Growth on PBAT was established this way, but the TA metabolism was strongly inhibited by the hydrolysate compared to pure TA. This is probably due to an inhibitory effect of AA and TA-containing oligomers on TA uptake or metabolism. After 10 days, all PBAT monomers were completely consumed, setting the path for a novel, industrially promising plastic upcycling concept.
Carotenoids, valued for their antioxidant properties and industrial applications, are witnessing rapidly accelerating market demands. Meanwhile, cyanobacteria with rapid growth and simple nutrient needs are extensively employed for the production of bioactive compounds in an environmentally sustainable manner. To address the limited application of cyanobacterial species in large-scale carotenoid production, our study investigated the effects of nutrients (NaNO₃ and KH₂PO₄), as well as salt (NaCl), metal (CuSO4.5H2O), and oxidative (H2O2) stress conditions, on carotenoid biosynthesis in the recently discovered fast-growing Synechococcus sp. PCC 11901. In our study, the RSM model (R2 value of 0.9373) based medium optimization has enhanced carotenoid and chlorophyll a production in PCC 11901 by 1.99- and 1.74-fold, respectively, under mild deficiencies of NaCl, NaNO₃, and KH₂PO₄. Furthermore, exposure to low Cu (40 µM) and moderate H₂O₂ stress (3 mM) elevated carotenoid production by 1.74- and 1.53-fold, respectively, on the eighth day. In comparison, previous optimization studies in cyanobacteria have reported up to a 2.52-fold (9.78 µg ml⁻1) and a 1.52-fold (0.369 µg ml⁻1) increase in carotenoid yields. Additionally, the increased Cu accumulation of 0.76 µg/mg dry biomass under 40 µM Cu stress indicates PCC 11901 potential in metal pollution bioremediation. In addition to the observed radical scavenging potential of the carotenoids, Cu and NaCl stress induced the highest ascorbate peroxidase and catalase antioxidant enzymatic activities in PCC 11901, respectively, compared to control and other stress conditions. The study revealed the potential of fast-growing Synechococcus sp. PCC 11901 in carotenoid production under abiotic stress conditions for environmentally safe and sustainable bio-manufacturing.
This study evaluates the potential of tropical grasses—Elephant Grass (Pennisetum purpureum), Guinea Grass (Panicum maximum), and Water Hyacinth (Eichhornia crassipes)—as low-cost substrates for l-lysine production by Corynebacterium glutamicum (ATCC 13032). Aqueous extracts of all three grasses were fermented with C. glutamicum; however, only elephant grass, which yielded the highest l-lysine levels, was subjected to optimization of fermentation conditions (pH, temperature, nutrient concentration, and aeration) to enhance microbial growth and l-lysine biosynthesis. Batch-to-batch consistency was ensured by standardizing sample collection and processing. Maximum l-lysine production reached 195 mg/L at pH 7.0, with the highest production (168.7 mg/L) at 40 °C. Thermodynamic analysis revealed an endothermic and spontaneous biosynthesis process (ΔH = + 9.15 kJ/mol, negative ΔG). Biochemical analysis showed increases in soluble protein, glucose, antioxidant capacity, total phenols, and flavonoids, especially in Water Hyacinth extracts. Enzyme assays confirmed enhanced α-amylase and protease activities. These results demonstrate the feasibility of using tropical grasses, particularly Water Hyacinth, as sustainable substrates for l-lysine production.
Achieving carbon neutrality has emerged as a critical global goal to address climate change. As the most significant greenhouse gas, CO2 poses environmental challenges while representing a potential carbon resource. Biotransformation technologies offer sustainable, low-energy-consumption solutions for CO2 utilization; however, their development is restricted by low gas solubility and microbial efficiency limitations. Crystalline materials have recently demonstrated great potential to enhance CO2 capture, boost electron transfer, and promote microbial immobilization. These materials can function as supports, catalysts, or functional media in microbial systems to improve transformation efficiency. This review summarizes recent advances in CO2-biotransformation crystalline material-microbe hybrid systems, with an in-depth analysis of material functions, key microbes, and carbon fixation mechanisms, and looks forward to the future development trend of crystalline material-microbe hybrid systems for CO2 bioconversion. This study provides critical insights for the development of highly efficient and industrially scalable CO2 biotransformation platforms.
Rhamnolipids (RL) are glycolipid biosurfactants produced by microbes, commonly used in oil recovery and bioremediation. This study investigates the scale-up of RL production by Pseudomonas aeruginosa RS6 using biodiesel side stream waste glycerol as substrate, transitioning from a 1.5 to a 5 L bioreactor under a controlled constant impeller tip speed (Vtip) of 0.882 m s−1. Enhanced gas–liquid mass transfer was achieved in the 5 L system, reflected by a higher volumetric oxygen transfer coefficient (
Improved gas-liquid mass transfer (
Most essential oils (EOs) have insecticidal properties but their use is limited due to their sensitivity to heat, light, and oxidation. A biological way to overcome this is by their encapsulation in yeast thus stabilizing them and also serve as an insecticide. Spent yeast, an underutilized predominant by-product of the wine and beer industries, can be biovalorized for encapsulation process. The present study was conducted to prepare Eucalyptus oil-based spent yeast (Saccharomyces cerevisiae) microcapsules. The effect of pre-treating yeast cells with NaCl, and varying the mass ratio of yeast to oil on encapsulation efficiency and loading capacity was investigated. Subsequently, the stability of the microcapsules was studied at 4 °C and 28 °C for 60 days. Fluorescent microscopy, Fourier transform-infrared spectroscopy and Transmission electron microscopy were used to analyze the process. NaCl [5% (w/v)]was found to be optimal for plasmolysis, resulting in a loading capacity of 44.78 ± 0.56% and a mass ratio of 2:1:4 (plasmolysed yeast: oil: water) recorded the highest encapsulation efficiency of 82.39 ± 1.33%. The storage studies revealed that microcapsules retained the loaded oil and were stable. Analytical methods confirmed the enhancing effect of plasmolysis and successful encapsulation of eucalyptus oil in yeast cells. Further, as yeast is a preferred food for mosquito larvae, EO-encapsulated yeast can be a promising, eco-friendly mosquito larvicide. Hence, the present work demonstrates proof-of-concept at laboratory scale, the field validation of which is needed to translate it into scalable, real-world bioinsecticidal applications.
1,3-Propanediol is a key monomer for producing polytrimethylene terephthalate. However, most reported microbial production systems rely on costly pure glycerol substrates or require expensive vitamin B12 supplementation, which considerably restricts their economic scalability. To overcome these constraints, a previously isolated Klebsiella pneumoniae strain was engineered to convert crude glycerol to 1,3-propanediol efficiently via adaptive laboratory evolution and rational metabolic engineering. In addition, an adenosine-driven ATP regeneration system was introduced; the engineered strain K. pneumoniae CG6.6 was capable of producing 110 g/L 1,3-propanediol in 48 h in a 5-L bioreactor, achieving a yield of 0.48 g/g without vitamin B12 supplementation. These findings highlight the potential of the engineered K. pneumoniae for sustainable and cost-effective 1,3-propanediol biosynthesis from crude glycerol.
Yeast culture, a product of anaerobic solid-state fermentation of plant-based substrates inoculated with yeast strains and dehydrated, is widely used in animal production. Studies show its diverse benefits, including improved feed palatability, increased feed intake, enhanced gastrointestinal and immune health, and better productivity and reproduction in various animal species. However, the specific bioactive components responsible for these effects remain unclear. In the context of postbiotics, yeast culture from solid-state fermentation contains a mix of cellular lysates, metabolites, and growth substrates, fitting the postbiotic definition. These components can be classified into six main groups: proteins, polysaccharides, organic acids, enzymes, vitamins, and nucleotides. Research gaps exist in understanding how these components interact with animal physiology. This review consolidates data on yeast culture postbiotics, explores their metabolic effects in animals, and proposes strategies for targeted accumulation of these compounds to drive innovation in yeast culture applications to more effectively improve animal production performance such as weight gain rate, milk production, and health levels such as improving intestinal barrier function. These improvements reduce the feed cost per unit product by improving the overall breeding efficiency.
Cyanobacteria are abundant autotrophic prokaryotes that can survive in changing environmental conditions by modifying their transcriptomes through different regulatory pathways. The Rnc and Mrnc proteins crucially regulate gene expression by functioning in RNA processing at the post-transcriptional level under both standard and environmental stress conditions. Despite the critical roles of Rnc and Mrnc in cyanobacteria, their structural attributes have not been extensively studied till date, either computationally or experimentally. Therefore, in the current study, we investigated the structure of Rnc and Mrnc proteins in Nostoc PCC 7524, a well-studied organism for the production of bio-fertilisers and metabolites. Further, the structural stability and compactness of both the proteins were established through in silico docking and molecular dynamics simulations to understand their biological significance and role in adaptation to stressful environmental conditions. Conclusively, the obtained stable Rnc and Mrnc complex from Nostoc sp. PCC 7524 suggests its potential unexplored role in gene regulation and cellular stress adaptation.
Low-density polyethylene (LDPE) has become a vital component of daily human existence. However, excessive usage resulted in build-up in the environment, producing environmental pollution, since there were no safe and appropriate disposal techniques available. Biodegradation is the most effective way to reduce synthetic plastic waste pollution in a sustainable manner. This study focused on the bacterial degradation of LDPE films using a novel isolate of Brevibacillus brevis strain CSK obtained from soil samples collected from a 20–30 years-old plastic waste dump at Yercaud foothills in Salem District, Tamil Nadu, India. The bacterial isolate B. brevis strain CSK degraded LDPE film after screening and exhibited has been condensed to focus mainly on the degradation rate (9.77% ± 0.42 in 30 days), key analytical methods (UV-Vis, ATR-FTIR, FE-SEM), and implications for eco-friendly biodegradation. The ATR-FTIR analysis of LDPE degraded film distinctive peaks corresponding to functional groups, with observable peak shifts in the treated films. FE-SEM analysis revealed biofilm formation, and topographical changes such as cracks, pits, cavities, and roughness in treated LDPE films. The research emphasizes the bacterial isolate’s potential for secure and efficient microbial remediation of LDPE films.
2’-Fucosyllactose (2’-FL), the most abundant human milk oligosaccharides (HMOs), plays crucial biological roles in regulation of intestinal microbiota, inhibition of pathogen adhesion, and immune enhancement. It holds considerable market potential in the pharmaceutical and food industries. Compared to chemical route, microbial synthesis using engineered strains, such as Escherichia coli, offers a more economical and efficient production route and currently represents the industrial standard for 2’-FL manufacturing. Notably, α-1,2-fucosyltransferase (FucT2) serves as the key rate-limiting enzyme in the 2’-FL biosynthetic pathway. Therefore, enhancing FucT2 activity is essential for improving 2’-FL titer. In this study, we explored and characterized an efficient FucT2 from Helicobacter himalayensis (H.him-FucT2), which demonstrated high catalytic activity in the synthesis of 2’-FL. Comparative analysis confirmed that H.him-FucT2 exhibits superior activity to FucT2 from Helicobacter pylori. Upon expression in an engineered Escherichia coli capable of de novo 2’-FL biosynthesis, the initial titer of 2’-FL reached 0.22 g/L. To further enhance the titer of 2’-FL, we adopted a dual-module bypass gene knockout strategy to redirect metabolic flux. Specifically, metabolic bypass genes (pfkA, pfkB, nudD, nudK, lacZ, and wcaJ) were systematically knocked out using the CRISPR/Cas9 system. Subsequently, through the composite application of global regulation and fermentation condition optimization, the titer of 2’-FL was further increased by 13.7 times, which reached 11.02 g/L with fed-batch fermentation.
Cyanide, a toxic compound from plant metabolism, might be accumulated in the plant-based food fermentation. It is important to degrade cyanide in these food fermentations. This study identified a potential cyanide-degradation enzyme gene (nitC) in Bacillus amyloliquefaciens CCTCC M 20242168 with cyanide-degrading activity in Baijiu fermentation. The nitC gene was expressed in Escherichia coli BL21 (DE3), and the purified enzyme successfully degraded cyanide into ammonia and formic acid. The purified nitrilase showed the highest activity (23.7 U/g) at the optimal pH of 6.5 and temperature of 35 °C. Substrate specificity analysis revealed high catalytic activity toward both cyanide and benzonitrile. Nitrilase NitC also exhibited high enzymatic activity under acidic conditions and its observed tolerance towards 1%–10% ethanol, suggesting its potential application in cyanide bioremediation, particularly under acidic conditions.