The concern for better life quality has been encouraging the bioprocess industries to develop technological strategies to obtain new biomolecules. Galactooligosaccharides (GOS) are an important class of food-grade oligosaccharides, being classified as non-digestible, and which present prebiotic potential, promoting better conditions of health and well-being. The main benefits include the selective stimulation of beneficial microorganisms, the decrease in the formation of toxic compounds, the increase in the absorption of minerals, improvement of an immune response, and a reduction in the severity of obesity and diabetes. This review approaches the recent methodologies and strategies to obtain GOS, their health benefits, purification, and technological properties for industrial application. Improvements in the process are continuously being investigated, with the technique of enzyme immobilization representing a potentially promising strategy. Sustainable GOS productions have been reported by the use of agro-industrial residues, such as cheese whey. Despite these advances, the main concern of the process consists in the low yield, which implies high investments in the purification of the bioproducts. Technological and nutritional approaches to the GOS application in different industrial sectors are also reported.

Plant pathogens damage crops and threaten global food security. Plants have evolved complex defense networks against pathogens, using crosstalk among various signaling pathways. Key regulators conferring plant immunity through signaling pathways include protein-coding genes and non-coding RNAs (ncRNAs). The discovery of ncRNAs in plant transcriptomes was first considered "transcriptional noise". Recent reviews have highlighted the importance of non-coding RNAs. However, understanding interactions among different types of noncoding RNAs requires additional research. This review attempts to consider how long-ncRNAs, small-ncRNAs and circular RNAs interact in response to pathogenic diseases within different plant species. Developments within genomics and bioinformatics could lead to the further discovery of plant ncRNAs, knowledge of their biological roles, as well as an understanding of their importance in exploiting the recent molecular-based technologies for crop protection.

Photosynthesis is responsible for the primary productivity and maintenance of life on Earth, boosting biological activity and contributing to the maintenance of the environment. In the past, traditional crop improvement was considered sufficient to meet food demands, but the growing demand for food coupled with climate change has modified this scenario over the past decades. However, advances in this area have not focused on photosynthesis per se but rather on fixed carbon partitioning. In short, other approaches must be used to meet an increasing agricultural demand. Thus, several paths may be followed, from modifications in leaf shape and canopy architecture, improving metabolic pathways related to CO2 fixation, the inclusion of metabolic mechanisms from other species, and improvements in energy uptake by plants. Given the recognized importance of photosynthesis, as the basis of the primary productivity on Earth, we here present an overview of the latest advances in attempts to improve plant photosynthetic performance. We focused on points considered key to the enhancement of photosynthesis, including leaf shape development, RuBisCO reengineering, Calvin-Benson cycle optimization, light use efficiency, the introduction of the C4 cycle in C3 plants and the inclusion of other CO2 concentrating mechanisms (CCMs). We further provide compelling evidence that there is still room for further improvements. Finally, we conclude this review by presenting future perspectives and possible new directions on this subject.

The trypsin being universal enzyme forming family of proteases catalyzes the hydrolysis of proteins into amino acids and regenerates the serine hydroxyl an active site. The trypsin enzyme from D. saccharalis, uses sericin as its preferred substrate. Presence of catalytic triad (serine, aspartic acid and histidine) at the substrate binding site of this enzyme is very important for the catalytic activity. In the current study, the interacting mechanism between the substrate sericin protein and enzyme trypsin protein were explored by integrating various computational approaches including physico-chemical properties, biophysical properties, dynamics, gene ontology, molecular docking, protein - protein interactions, binding free energy calculation and structural motifs were studied. The evolutionary study performed by MEGA X showed that trypsin protein sequence (ALE15212.1) is closely related to cocoonase protein sequence (ADG26770.1) from Antheraea pernyi. 3-D models of trypsin and sericin proteins were predicted using I-TASSER and further validated by PROCHECK, and ProSAweb softwares. The predicted trypsin structure model was assigned E.C. no. 3.4.21.4 which refers hydrolytic mechanism. Gene Ontology predicted by QuickGO showed that trypsin has serine hydrolase activity (GO: 00017171), and part of proteolysis (GO: 0006508) as well as protein metabolic process (GO:0019538) actvity. Molecular docking studies between trypsin and sericin proteins were conducted by the HADDOCK 2.4 having best docked protein complex with Z-score - 1.9. 2D and 3D protein-protein interaction was performed with LIGPLOT+ and HAWKDOCK, PDBsum, respectively. The amino acid residues interacting across proteins interface are sericin_chain A representing "Ser133, Tyr214, Thr188, Thr243, Ser225, Ser151, Ser156, His294, Arg293, Gly296″ and trypsin_chain B "Lys120, Tyr246, Asn119, Glu239, Ser62, Tyr194, Ile197, Ser171, Tyr169, Gly170″. Based on our results trypsin shows similarity with cocoonase and presumably trypsin can be used as an alternative source in cocoon degumming.

Heavy metal contamination is a global issue, with cadmium (Cd2+) and its treatment becoming major environmental challenge that could be solved by microbial restoration, an eco-friendly technique. Serratia marcescens KMR-3 exhibits high tolerance and removal rate of Cd2+ (≤500 mg/L). Here, we aimed to explore mechanisms underlying tolerance to and removal of Cd2+ by KMR-3. Scanning electron microscopy, X-ray photoelectron spectroscopy, and Fourier transform infrared spectrometry were conducted to analyze characteristics of the KMR-3 biofilm and Cd2+ combined forms. The results revealed varying degrees of cell adhesion, membrane thickening, and shrinkage on the surface of the bacteria. The binding elements, electronic binding energy, and functional groups on the surface of the bacteria exhibited changes. Furthermore, the biofilm amount following treatment with Cd2+ was 1.5-3 times higher than that in the controls, treatment with Cd2+ substantially enhanced biofilm generation and increased Cd2+ adsorption. Cd2+ adsorption by its own secondary metabolite prodigiosin produced by KMR-3 was enhanced by 19.5 % compared with that observed without prodigiosin. Through transcriptome sequencing and RT-qPCR, we observed that Znu protein-chelating system regulated gene expression (znuA, znuB, and znuC), and the efflux mechanism of the P-type ATPase regulated the expression of genes (zntA, zntB, and zntR), which were significantly enhanced. Through the combined action of various strategies, KMR-3 demonstrated a high tolerance and removal ability of Cd2+, providing a theoretical basis to treat Cd2+ pollution.

Computational fluid dynamics (CFD) was used to investigate cascade photobioreactors (cascade PBRs) with two different bottom configurations-flat and wavy-to establish the effect that fluid-flow regimes exert on the photosynthetic productivity of Chlorella sorokiniana. In the flat-bottom PBR, areal biomass productivities decreased from 6.8 to 4.2 g·m-2·d-1 when the flow rate of a culture per unit of lane width was increased from 33 to 132 L·m-1·min-1. We found that this decrease in the areal productivity was the result of a decrease in the volumetric photon flux densities (volumetric PFDs), which was caused by an increase in the depth of the culture in the lane. Through CFD calculation and long-exposure photography, the flow of the culture in the wavy-bottom PBR was characterized in an upper straightforward section and underneath the swirling section. Under identical conditions of flow rate and volumetric PFD (66 L·m-1·min-1 and 50 μmol·m-3·s-1, respectively), the cell growth accelerated in the wavy-bottom PBR with areal productivity that reached 6.5 g·m-2·d-1-productivity was 5.1 g·m-2·d-1 in the flat-bottom PBR. The swirling flow in the wave troughs held the culture for longer periods in the illuminated lane, and the resultant extended period of mixing improved the photosynthetic productivity.

Protein L (PpL) is a universal binding ligand that can be used for the detection and purification of antibodies and antibody fragments. Due to the unique interaction with immunoglobulin light chains, it differs from other affinity ligands, like protein A or G. However, due to its current higher market price, PpL is still scarce in applications. In this study, we investigated the recombinant production and purification of PpL and characterized the product in detail. We present a comprehensive roadmap for the production of the versatile protein PpL in E. coli.

Isopropanol has a good potential as a new fuel substitution. In the model biosynthesis pathway of isopropanol synthesis, acetoacetyl-CoA is converted to acetoacetate by acetoacetyl-CoA transferases, which requires an acetate molecule as a substrate. Herein, a novel isopropanol synthesis pathway based on mammalian ketone metabolic pathway was developed. In this pathway, acetoacetyl-CoA is condensed with acetyl-CoA to generate 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) by HMG-CoA synthase, and then catalyzed by HMG-CoA lyase to generate acetoacetate. This process is acetate-independent. Under the same experimental system using glycerol as carbon source, the E. coli strain MG::ISOP1 containing the novel pathway produced 11.7 times more isopropanol than the strain MG::ISOP0 containing the model pathway. The pta-ackA knockout mutant strain MG∆pta-ackA::ISOP1, which reduced the conversion of acetyl-CoA to acetate, further increased the production from 76 mg/L to 360 mg/L. In another strategy, knocking out atoDA to block the acetoacetate degradation pathway in strain MG∆atoDA::ISOP1 increased the production to 680 mg/L. By knocking out both of pta-ackA and atoDA, strain MGΔpta-ackAΔatoDA::ISOP1 produced 964 mg/L of isopropanol, which was 12.7 times that of MG::ISOP1. This study indicated that the novel pathway is competent for isopropanol synthesis, and provides a new perspective for biosynthesis of isopropanol.

Currently, organic field effect transistors (OFETs) are widely used in the field of biodetection because of their inherent gain amplification function and good biocompatibility. However, the vast majority of OFET biosensors have disadvantages including a long preparation cycle, complicated processing and expensive materials. Thus, this paper proposes a simple and inexpensive preparation method for label-free detection of immunoglobulin G (IgG). In this work an active dielectric bilayer field effect transistor with 3D-printed silver source and drain electrodes was used. The active layer was modified by physically adsorbing an anti-IgG antibody on P3HT as a recognition element and then sealed with bovine serum albumin (BSA) to prevent nonspecific adsorption. The carrier mobility of the bilayer dielectric layer active field effect transistor reached 4.6 × 10-2 cm2/Vs, and the dynamic detection range for IgG spanned 6 orders of magnitude with a detection limit of 2.9 Picomolar (pM). Controlled experiments demonstrated that the developed sensor has high selectivity for IgG. Overall, this easy-to-operate and low-cost OFET biosensor has broad commercialization prospects and lays the foundation for the early clinical detection of disease-causing biomolecules.

Streptococcus agalactiae (Group B Streptococcus, GBS) is primarily known as a major neonatal pathogen. In adults, these bacteria often colonize the gastrointestinal and urogenital tracts. Treatment of infections using antibiotics is often complicated by recurrences caused by multi-resistant streptococci. Endolysin EN534 from prophage A2 of human isolate Streptococcus agalactiae KMB-534 has a modular structure consisting of two terminal catalytic domains, amidase_3 and CHAP, and one central binding domain, LysM. The EN534 gene was cloned into an expression vector, and the corresponding recombinant protein EN534-C was expressed in Escherichia coli in a soluble form and isolated by affinity chromatography. The lytic activity of this endolysin was tested on cell wall substrates from different GBS serotypes, B. subtilis, L. jensenii, and E. coli. The enzyme lysed streptococci, but not beneficial vaginal lactobacilli. The isolated protein is stable in a temperature range of 20-37 °C. Calcium ions enhanced the activity of the enzyme in the pH range from 5.0 to 8.0. The exolytic activity of EN534-C was observed by time-lapse fluorescence microscopy on a S. agalactiae CCM 6187 substrate. Recombinant endolysin EN534-C may have the potential to become an antimicrobial agent for the treatment of S. agalactiae infections.

To investigate the role of the sugar transporter MAL31 on pullulan biosynthesis, the coding gene mal31 was respectively disrupted and overexpressed in the parental strain A. pullulans CCTCC M 2012259 to construct mutants of A. pullulans Δmal31 and A. pullulans Mal31. Batch pullulan production significantly decreased by 69.1 % in A. pullulans Δmal31 but increased by 15.9 % in A. pullulans Mal31, as compared to the parental strain. We performed kinetics analysis, assays of key enzymes, determination of intracellular UDPG, NADH, and ATP contents, and measurement of transcriptional levels of genes associated with pullulan biosynthesis and excretion. The results confirmed that the mal31 disruption decreased the glucose consumption rate, decreased the formation rate and titer of pullulan, but increased the intracellular UDPG supply for β-glucan accumulation. In contrast, the mal31 overexpression increased the transcriptional levels of genes associated with pullulan biosynthesis, and accelerated the rates of glucose consumption and pullulan formation, thereby increased pullulan production. Our findings revealed that MAL31 is involved in the transport of precursors for pullulan biosynthesis. This study provides an accurate operating site for genetic modification of A. pullulans for improving pullulan production and also presents a feasible technique route for the overproduction of other polysaccharides.

Pattern detection and string matching are fundamental problems in computer science and the accelerated expansion of bioinformatics and computational biology have made them a core topic for both disciplines. The requirement for computational tools for genomic analyses, such as sequence alignment, is very important, although, in most cases the resources and computational power required are enormous. The presented Multiple Genome Analytics Framework combines data structures and algorithms, specifically built for text mining and (repeated) pattern detection, that can help to efficiently address several computational biology and bioinformatics problems, concurrently, with minimal resources. A single execution of advanced algorithms, with space and time complexity O(nlogn), is enough to acquire knowledge on all repeated patterns that exist in multiple genome sequences and this information can be used as input by meta-algorithms for further meta-analyses. For the proof of concept and technology of the proposed Framework scalability, agility and efficiency, a publicly available dataset of more than 300,000 SARS-CoV-2 genome sequences from the National Center for Biotechnology Information has been used for the detection of all repeated patterns. These results have been used by newly introduced algorithms to provide answers to questions such as common patterns among all variants, sequence alignment, palindromes and tandem repeats detection, different organism genome comparisons, polymerase chain reaction primers detection, etc.

During this decade, selenium nanoparticles have been found to play a crucial role in helping plants endure several stress conditions, which thereby helps enhance the production of crops in such harsh environments. Globally, high salinity is considered a long-term stress in the crop fields which affects the growth and production of many crops, including mustard-one of the most important oil crops. Here, the activities of spherical-shaped selenium nanoparticles with an average particle size of 55.81 nm, synthesized and functionalized by phytochemicals of fresh grape aqueous extract, were evaluated in the salinity stress (200 mM NaCl) tolerance of mustard plants grown hydroponically in modified Hoagland's solution. These bioactive nanoparticles (30 mg L-1) have exhibited significant activity in alleviating the salt stress complications in mustard, enhancing the activities of antioxidant enzymes (SOD 41.20 %, CAT 64.10 %, APX 63.06 %, and POX 70.43 %), phenolic content (98.88 %), flavonoid content (86.90 %), and free radical scavenging activity (61.89 %). The seed germination percentage, root and shoot length, fresh and dry weight per plant, water content percentage, chlorophyll content, carbohydrate content, and protein content were significantly improved by 39.66 %, 75 %, 60.64 %, 41.2 %, 22.11 %, 1.02 %, 81.92 %, 24.65 % and 79.14 % respectively by the nano selenium application during NaCl stress compared to the control group growing under salt stress without nanoparticles. Gas chromatography-mass spectrometry chromatogram analysis inferred the interaction between the nano-selenium and mustard plants under salt stress. Besides, the in-silico analysis revealed the active molecular interactions between selenium and 20 different proteins of mustard, including glutathione peroxidase, an important antioxidant enzyme.

Using lignocellulosic biomass is immensely beneficial for the economical production of biochemicals. However, utilizing mixed sugars from lignocellulosic biomass is challenging because of bacterial preference for specific sugar such as glucose. Although previous studies have attempted to overcome this challenge, no studies have been reported on isobutanol production from mixed sugars in the Escherichia coli strain. To overcome catabolite repression of xylose and produce isobutanol using mixed sugars, we applied the combination of three strategies: (1) deletion of the gene for the glucose-specific transporter of the phosphotransferase system (ptsG); (2) overexpression of glucose kinase (glk) and glucose facilitator protein (glf); and (3) overexpression of the xylose regulator (xylR). xylR gene overexpression resulted in 100% of glucose and 82.5% of xylose consumption in the glucose-xylose mixture (1:1). Moreover, isobutanol production increased by 192% in the 1:1 medium, equivalent to the amount of isobutanol produced using only glucose. These results indicate the effectiveness of xylR overexpression in isobutanol production. Our findings demonstrated various strategies to overcome catabolite repression for a specific product, isobutanol. The present study suggests that the selected strategy in E. coli could overcome the major challenge using lignocellulosic biomass to produce isobutanol.

Streptomyces corchorusii TKR8, Streptomyces corchorusii JAS2 and Streptomyces misionensis TBS5 were previously obtained from rice fields and have been studied as a biocontrol agent against the causal agent of Bacterial Panicle Blight (BPB) disease on rice, Burkholderia glumae, and rice plant growth promoter. This study evaluated the potential of plant growth-promoting Streptomyces (PGPS) to control B. glumae and promote rice plants' growth under greenhouse conditions. PGPS were further characterized based on their phenotypic and biochemical differences. Multilocus sequence analysis (MLSA) by amplifying gyrB, rpoB and trpB using PCR was conducted to identify the PGPS further. The antimicrobial activity of PGPS against B. glumae was investigated using a survival assay and microscopic analysis. Result indicates that JAS2 (61.2 %) utilized the highest number of carbohydrates tested, followed by TKR8 (53.1 %) and TBS5 (40.8 %) as analyzed using API 50 CH. Based on MLSA analysis of the concatenated partial sequences (1520 bp) from three housekeeping genes, the neighbor-joining tree identified JAS2 and TKR8 as S. corchorusii. Meanwhile, TBS5 as S. misionensis. Antimicrobial activity of PGPS against B. glumae has found that the supernatant of Streptomyces reduced the survival viability of B. glumae up to 50.7-70.3 %. SEM images showed that substantial morphological changes happened in cell membranes of B. glumae after the Streptomyces treatment. The highest vigor index of inoculated seedlings was determined when rice seeds were treated with a spore suspension of 1 × 107 spore/mL (for JAS2 and TKR8) and 1 × 106 spore/mL (for TBS5). Under greenhouse conditions, Streptomyces-treated plants showed improvement in rice plants' growth and grain yield and reduced the BPB disease severity. Results suggest that the S. corchorusii TKR8, S. corchorusii JAS2 and S. misionensis TBS5 should be promoted as biocontrol agents against B. glumae and bioformulations for rice crops.

By applying periodic DFT computations, the possible employment of BC3 nanotube (BC3NT) as a drug delivery system (DDS) for Thiotepa (TPA) anticancer medicine has been examined. Quantum mechanics computations by the B3LYP-6-31 +G(d,p) method with dispersion correction including to be used for calculation the details of electronic, geometric, and energetic features of the interactions between TPA drug and BC3NT. The appropriate orientation for the TPA interacting with BC3NT has been assessed and adsorption energies (ΔEad) have been computed. The band distance of S and B atom in complex C2 is about 1.89 Å, also the value of ΔEad of - 29.83 kcal/mol in most stable compounds. By applying frontier molecular orbital analysis, it has been assessed that during stimulation, TPA medicine performs as HOMO and delivers the charge towards the LUMO, i.e., BC3NT. In the aqueous phase, the λmax of BC3NT-AP complex is blue-shifted by 36 nm. Drug delivery to the specific cells after protonation has been studied, due to the point that cancer cells have lower pH than others. The value of computed solvation energy (ΔEsolvation) shows the solubility BC3NT@TPA system in the water phase. The effects of the present investigation verified the ability of BC3NT as a drug delivery agent for TPA in the treatment of cancer.

Lack of appropriate process models, reliable online sensors, and process variability in bioprocess systems are poising challenges in real-time monitoring and control of critical process parameters (CPPs). This present investigation deals with the development of a non-invasive soft sensor by utilizing metabolic heat rate as input signal for online estimation of specific growth rate (μest) during the induction phase of glycoengineered Pichia pastoris for human interferon-alpha 2b (huIFNα2b) production. Feedforward strategy employing a predetermined exponential feeding of methanol during the induction phase was dealt at defined setpoint values (μSP). Standard PID controller with predetermined gain values regulated methanol feeding in accordance with the deviation from the μSP value. An adaptive PID (gain scheduling) significantly minimized the deviation of μ from its μSP value, reduced the amplitude of oscillation and achieved long-term controller stability. Robust control of methanol feeding by adaptive PID resulted in a 1.5 and 2.2-fold increase in productivity of huIFNα2b compared to standard PID and feedforward controls respectively. Moreover, adaptive PID control facilitated narrow range control of μ for longer durations (> 20 h) with a low average tracking error (< 6%) enumerating its scope of application in therapeutic protein production in near future.

An effective prototype of photobioreactor namely hollow fiber membrane bioreactor is employed to produce algal biomass for biofuel production. The effect of two-stage cultivation system on the biomass productivity and lipid production is studied using freshwater green microalga Tetradesmus obliquus MT188616.1. The hybrid system combines exponential biomass production in hollow fiber membrane photobioreactor (HFMPBr) and a coordinated high-lipid stimulation phase in nitrogen-deprived medium by the shake flask method. This work is proposed to examine the usefulness of HFMPBr module to enhance the microalgal growth rate through the effective mass transfer by standardizing the culture medium re-circulation flow rates (5 - 45 mL min-1) through the hollow fiber membranes. Cultivation is carried out at continuous mode in HFMPBr containing polysulfone fabricated membranes at fixed light intensity of 50 µmol m-2 s-1 and temperature at 28 °C. Biomass productivity and specific growth rate obtained are 0.44 g L-1d-1 and 1.61 µ d-1 with a lipid yield of 0.1 g L- 1. The key operating parameter i.e., liquid flow rate is optimized based on biomass production. The highest biomass concentration is produced at the flow rate of 45 mL min-1. The results showed that HFMPBr module is a better choice for the algal-cultivation to obtain higher biomass yield.

Previously, we reported, based on an untargeted metabolomics, carnitine derivatives are part of a mechanism to overcome impaired mitochondrial functioning triggered by an acyl-group overflow in CHO cells. In this study, we analyzed the cell-specific rates of 24 selected metabolites using two metrics: correlation coefficients and root-mean-square deviations (RMSDs) between glucose-fed versus glucose/lactic acid-fed cultures. The time-course profiles of acetylcarnitine, adipoylcarnitine, glutarylcarnitine, glutamate, and succinate exhibited significant negative correlations between the two culture conditions. Based on RMSDs, seven carnitine derivatives, 3-hydroxy-methyl-glutarate, mevalonate, pyridoxamine-5-phosphate, succinate, and glycine were substantially different. The analyses from the two metrics reveal a distinctive rearrangement of rates from the following metabolic pathways: (i) high secretion rates of carnitines as part of the acyl-group removal, (ii) low secretion rates of succinate, related to the tricarboxylic acid cycle and the electron-transport chain, (iii) low secretion rates of pyridoxamine-5-phosphate - a co-factor for amino acid catabolism, transaminations, and transsulfuration, and (iv) increases in the consumption rates of glutamate and glycine, both used to produce glutathione. The rewiring in rates observed upon feeding lactic acid is best explained by the activation of pathways supporting homeostasis of acyl-groups and antioxidant synthesis, which are required for continuous proper functioning of oxidative phosphorylation.

Modulation of expression levels of endogenous or recombinant genes can be of great interest for diverse applications, such as the study of genotype-phenotype relationships for a gene of interest, or fine-tuning of transcription to determine physiologically relevant effects of gene expression levels. During the last decades, several synthetic biology tools were established to control gene expression in mammalian cells such as Chinese hamster ovary (CHO) cells, one of the most important cell systems for basic research as well as the production of biopharmaceuticals. Here we describe the use of triplex forming oligos (TFOs), short RNA or ssDNA molecules that can bind to the major grove of their target duplex with great specificity, to control transgene expression in CHO cells. For proof of concept, a panel of TFOs with a size of 10-20 nts were designed with the help of the on-line tool Triplexator targeting the viral cytomegalovirus (CMV) promoter/enhancer region controlling the downstream reporter gene hCD4. The effect of TFOs was tested as ssDNA oligos pre-annealed to the promoter/enhancer region in vitro as well as upon endogenous transcription of the TFO as an RNA molecule binding to their target duplex in vivo. Results showed that not only binding of the TFO, but the exact location of triplex formation within the promoter/enhancer is paramount for transcription inhibition. After relieving a binding conflict by introducing a point mutation within the CMV promoter, longer TFOs (26-30 nts) could be designed and analysed. Selected TFOs achieved a reduction in recombinant hCD4 expression of up to 85% in CHO-K1 cells.

Simultaneous coexpression of multiple proteins is essential for biotechnology and synthetic biology. Currently, the most popular polyprotein coexpression system utilizes the foot-and-mouth disease virus (FMDV) 2A peptide that mediates translational ribosome-skipping events. However, due to unfavorable consumer acceptance of transgenic products containing animal-virus sequences, novel non-viral 2A-like peptides from purple sea urchin (Strongylcentrotus purpuratus) and California sea slug (Aplysia californica) were investigated for polyprotein coexpression in this study. We demonstrated that these non-viral 2A sequences functioned similarly to their viral counterpart in polyprotein processing, in both plant and mammalian cells, and were successfully used to express a functional recombinant antibody. The new non-viral 2A-like sequences offer an alternative tool for engineering multigenic traits or production of protein complexes as biomedicine via coexpression of protein subunits.

Microalgae biomass has been considered as one of the potential feedstocks in biofuel production. Yet, biomass harvesting poses a challenge to the overall production cost due to its low cell density. Flocculation has been marked as one of the promising processes in microalgae harvesting technology. In this study, the first screening of two anionic (A-230, and A-330E) and five cationic polymers (C-810E, C-810EL, C-810EB, C-810ELH, and C-810EMB) followed by gravity settling with the mixed microalgae concentration of 2.24 gTSS/L revealed that anionic polymers are less effective. Whereas all cationic polymers achieved above 90% harvesting efficiency. Therefore, the maximum mass recovery of 98.7% with 86.8 gTSS/L sediment content was achieved by adjusting pH to 6-0.6 mL/L (115.178 mg/gbiomass) of C-810E followed by 15-min settling. The cationic polymer addition followed by settling would enable cost-effective downstream processing of microalgal biomass.

Microalgae are highly photosynthetic unicellular organism that have increased demand in the recent days owing to the presence of valuable cellular metabolites. They are ubiquitous in terrestrial and aquatic habitats, rich in species diversity and are capable of generating significant biomass by efficiently using CO2, light and other nutrients like nitrogen, phosphate etc., The microalgal biomass has upsurged in economic potential and is used as both food and feed in many countries across the world, accounting for more than 75 % of annual microalgal biomass production in the past decades. The microalgal cells are sustainable resource that synthesize various secondary metabolites such as carotenoids, polysaccharides, polyphenols, essential amino acids, sterols, and polyunsaturated fatty acids (PUFA). Microalgae and its derived compounds possess significant pharmacological and biological effects such as antioxidant, anti-inflammatory, anti-cancer, immunomodulatory and anti-obesity. Because of their potential health promoting properties, the utilization of microalgae and its derived substances in food, pharmaceutical and cosmetic industries has skyrocketed in recent years. In this context, the current review discusses about the benefits of microalgae and its bioactive compounds against several neurodegenerative disorders like Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), and amyotrophic lateral sclerosis (ALS).

The economical production of value-added chemicals from renewable biomass is a promising aspect of producing a sustainable economy. Itaconic acid (IA) is a high value-added compound that is expected to be an alternative to petroleum-based chemicals. In this study, we developed a metabolic engineering strategy for the large-scale production of IA from glucose using the fission yeast Schizosaccharomyces pombe. Heterologous expression of the cis-aconitic acid decarboxylase (CAD) gene from Aspergillus terreus, which encodes cis-aconitate decarboxylase in the cytosol, led to the production of 0.132 g/L of IA. We demonstrated that mitochondrial localization of CAD enhanced the production of IA. To prevent the leakage of carbon flux from the TCA cycle, we generated a strain in which the endogenous malate exporter, citrate lyase, and citrate transporter genes were disrupted. A titer of 1.110 g/L of IA was obtained from a culture of this strain started with 50 g/L of glucose. By culturing the multiple mutant strain at increased cell density, we succeeded in enhancing the IA production to 1.555 g/L. The metabolic engineering strategies presented in this study have the potential to improve the titer of the biosynthesis of derivatives of intermediates of the TCA cycle.

The growth of resistance to multiple herbicides in grass weeds is a major threat to global cereal production and in the UK, is epitomized by the loss of control of blackgrass (Alopecurus myosuroides), causing losses in winter wheat production equating to 5% of national consumption. With an urgent need to develop new black-grass management tools, we have developed a lateral flow assay (LFA) that can predict resistance to multiple herbicides within 10 min.

Recombinant Escherichia coli grown in large-scale fermenters are used extensively to produce plasmids and biopharmaceuticals. One method commonly used to control culture growth is predefined glucose feeding, often an exponential feeding profile. Predefined feeding profiles cannot adjust automatically to metabolic state changes, such as the metabolic burden associated with recombinant protein expression or high-cell density associated stresses. As the culture oxygen consumption rates indicates a culture's metabolic state, there exist several methods to estimate the oxygen uptake rate (OUR). These common OUR methods have limited application since these approaches either disrupt the oxygen supply, rely on empirical relationships, or are unable to account for latency and filtering effects. In this study, an oxygen transfer rate (OTR) estimator was developed to aid OUR prediction. This non-disruptive OTR estimator uses the dissolved oxygen and the off-gas oxygen concentration, in parallel. This new OTR estimator captures small variations in OTR due to physical and chemical manipulations of the fermenter, such as in stir speed variation, glucose feeding rate change, and recombinant protein expression. Due its sensitivity, this non-disruptive real-time OTR estimator could be integrated with feed control algorithms to maintain the metabolic state of a culture to a desired setpoint.

Stevia rebaudiana is one of the vastly acclaimed commercial plant in the world and belongs to Asteraceae family. The exclusive advantage of Stevia over artificial sweeteners is impeccable and targets its potentiality to the presence of diterpene glycosides. Moreover, the flaunting sweetness of steviol glycosides with associated medicinal benefits, turns the plant to be one of the most economic assets, globally. As compared to vegetative propagation through stem-cuttings, plant tissue culture is the most suitable approach in obtaining true-to-type plants of superior quality. During last few decades, significant in vitro propagation methods have been developed and still the research is ongoing. The present review discusses the tissue culture perspectives of S. rebaudiana, primarily focusing on the mineral nutrition, growth regulators and other accessory factors, motioning the optimum growth and development of the plant. Another crucial aspect is the generation of sweeter varieties in order to reduce the bitter-off taste, which is noticed after the consumption of the leaves. The in vitro cultures pose an efficient alternative system for production of steviol glycosides, with higher rebaudioside(s) content. Moreover, the review also covers the recent approaches pertaining to scale-up studies and genome editing perspectives.

(S)-2-chlorophenylglycine ((S)-CPG) is a key chiral intermediate for the synthesis of clopidogrel. Herein, a novel, efficient and environmentally friendly chemo-enzymatic route for the preparation of optically pure (S)-CPG was developed. A straightforward chemical synthesis of the corresponding prochiral keto acid substrate (2-chlorophenyl)glyoxylic acid (CPGA) was developed with 91.7% yield, which was enantioselectively aminated by leucine dehydrogenase (LeuDH) to (S)-CPG. Moreover, protein engineering of LeuDH was performed via directed evolution and semi-rational design. A beneficial variant EsLeuDH-F362L with enlarged substrate-binding pocket and increased hydrogen bond between K77 and substrate CPGA was constructed, which exhibited 2.1-fold enhanced specific activity but decreased thermal stability. Coupled with a glucose dehydrogenase from Bacillus megaterium (BmGDH) for NADH regeneration, EsLeuDH-F362L completely converted up to 0.5 M CPGA to (S)-CPG in 8 h at 40 °C.

Biohydrogen production using renewable sources has been regarded as one of the most sustainable ways to develop low-cost and green production technology. In order to achieve this objective, herein biohydrogen production has been conducted using the combination of untreated secondary sewage sludge (Sss), algal biomass hydrolyzate (Abh), graphene oxide (GO) and bacterial consortia that forms a granular system. Thus, naturally formed granular system produced cumulative H2 of 1520 mL/L in 168 h with the maximum production rate of 13.4 mL/L/h in 96 h at initial pH 7.0, and optimum temperature of 37 °C. It is noticed that the combination of Abh, Sss and GO governed medium showed 42.05 % higher cumulative H2 production along with 22.71 % higher production rate as compared to Abh and Sss based H2 production medium. The strategy presented herein may find potential applications for the low-cost biohydrogen production using waste biomasses including Sss and Abh.

The oleaginous yeast Lipomyces starkeyi is expected to be a new lipid source since this microorganism is capable of accumulating more than 85% lipid per dry cell weight. For effective utilization of oleaginous yeast, mutants with improved lipid production compared to the wild-type have been screened by methods such as single-cell sorting and Percoll density gradient centrifugation. Because these methods need to reculture all mutated oleaginous yeasts together in a flask, it is difficult to evaluate the growth of each individual mutant. Thus, screening for the slow-growing mutants with high-throughput has never been performed by conventional methods. In this study, we developed a high-throughput method using gel microdroplets (GMD). With this method, the growth and lipid production of L. starkeyi can be evaluated simultaneously. L. starkeyi grew in GMD and the size of these microcolonies was evaluated by scattered light. Finally, a mutant with a 10-fold delay in growth compared to the wild-type was obtained. Analysis of genetic information in this mutant could reveal valuable information about critical genes involved in the growth of these microorganisms, which could then be utilized further.

Postbiotics is a novel term proposed to describe as a set of bioactive compounds obtained from beneficial microorganisms. In this work, postbiotics from four lactic acid bacteria (LAB) including Leuconostoc mesenteroides subsp. cremoris, Pediococcus acidilactici, Lactococcus lactis subsp. lactis and Streptococcus thermophilus were prepared in MRS broth. The antimicrobial properties and organic acids content of postbiotics were also investigated. Postbiotics were used to tentatively reduce the production of biogenic amines by foodborne pathogens (i.e., Salmonella paratyphi A and Escherichia coli) on lysine decarboxylase broth (LDB). Experimental data showed that acetic, propionic, and butyric acids were in the range of 387.51-709.21 mg/L, 0.00-1.28 mg/L, and 0.00-20.98 mg/L, respectively. The inhibition zone of postbiotics on E. coli and S. paratyphi A were 11.67, and 12.33 mm, respectively. Two different levels of postbiotics (25%, and 50%) were used in LDB to measure the diamines (cadaverine and putrescine), polyamines (agmatine, spermidine, and spermine, ammonia), and other biogenic amine formation by pathogens. E. coli produced cadaverine and putrescine with concentrations of 1072.21 and 1114.18 mg/L, respectively. The postbiotics reduced cadaverine formation by 67% in E. coli, and cadaverine production was mostly suppressed by postbiotics from P. acidilactici in E. coli (97%) and L. lactis subsp. lactis in S. paratyphi A (90%). Putrescine production by E. coli was reduced by 94% with postbiotics of P. acidilactici at a concentration of 25%, whereas putrescine production by S. paratyphi A has been decreased by 61% in the presence of postbiotics from L. lactis subsp. Lactis with a 25% concentration. The results revealed that an increase in postbiotics concentration (from 25% to 50%) in LDB may lead to synergistic effects, resulting from the production of biogenic amines by microbial pathogens. It was importantly concluded that postbiotics of LAB may degrade biogenic amines or prevent their formation by foodborne pathogens.

Poly(ethylene terephthalate) (PET) is one of the main synthetic plastics produced worldwide. The extensive use of this polymer causes several problems due to its low degradability. In this scenario, biocatalysts dawn as an alternative to enhance PET recycling. The enzymatic hydrolysis of PET results in a mixture of terephthalic acid (TPA), ethylene glycol (EG), mono-(2-hydroxyethyl) terephthalate (MHET) and bis-(2-hydroxyethyl) terephthalate (BHET) as main products. This work developed a new methodology to quantify the hydrolytic activity of biocatalysts, using BHET as a model substrate. The protocol can be used in screening enzymes for PET depolymerization reactions, amongst other applications. The very good fitting (R2 = 0.993) between experimental data and the mathematical model confirmed the feasibility of the Michaelis-Menten equation to analyze the effect of BHET concentration (8-200 mmol L-1) on initial hydrolysis rate catalyzed by Humicola insolens cutinase (HiC). In addition to evaluating the effects of enzyme and substrate concentration on the enzymatic hydrolysis of BHET, a novel and straightforward method for MHET synthesis was developed using an enzyme load of 0.025 gprotein gBHET-1 and BHET concentration of 60 mmol L-1 at 40 °C. MHET was synthesized with high selectivity (97 %) and yield (82 %). The synthesized MHET properties were studied using differential scanning calorimetry (DSC), thermogravimetry (TGA), and proton nuclear magnetic resonance (1H NMR), observing the high purity of the final product (86.7 %). As MHET is not available commercially, this synthesis using substrate and enzyme from open suppliers adds new perspectives to monitoring PET hydrolysis reactions.

For biotechnology applications, a novel nanobiocomposite was synthesized based on modification of graphene oxide (GO) by extracted silk fibroin (SF), natural polymer pectin (Pec) and zinc chromite (ZnCr2O4) nanoparticles (NPs). The structure and properties of hybrid nanobiocomposite GO-Pec/SF/ZnCr2O4 such as thermal stability, less toxicity, biocompatibility, antibacterial, and biodegradable were proved by using field emission scanning electron microscope (FE-SEM), Fourier-transformed infrared (FT-IR), Energy dispersive X-ray spectroscopy (EDS), thermal gravimetric analysis (TGA), and X-Ray diffraction (XRD). According to the biological features of substances, the GO-Pec/SF/ZnCr2O4 nanobiocomposite shows perfect results in MTT (83.71 %) and Hemolysis (16.52 %) assays. accordingly, mentioned properties of this nanobiocomposite can be used as a scaffold for medical applications.

l-Rhamnose isomerase (l-RhI) catalyzes rare sugar isomerization between aldoses and ketoses. In an attempt to alter the substrate specificity of Thermoanaerobacterium saccharolyticus NTOU1 l-RhI (TsRhI), residue Ile102 was changed to other polar or charged amino acid residues by site-directed mutagenesis. The results of activity-screening using different substrates indicate that I102N, I102Q, and I102R TsRhIs can increase the preference against d-allose in comparison with the wild-type enzyme. The catalytic efficiencies of the purified I102N, I102Q, and I102R TsRhIs against d-allose are 148 %, 277 %, and 191 %, respectively, of that of wild-type enzyme, while those against l-rhamnose are 100 %, 167 % and 87 %, respectively. Mutant I102N, I102Q, and I102R TsRhIs were noted to have the altered substrate specificity, and I102Q TsRhI has the highest catalytic efficiency against d-allose presumably through the formation of an additional hydrogen bond with d-allose. The purified wild-type and mutant TsRhIs were further used to produce d-allose from 100 g/L d-fructose in the presence of d-allulose 3-epimerase, and the yields can reach as high as 22 % d-allulose and 12 % d-allose upon equilibrium. I102Q TsRhI takes only around half of the time to reach the same 12 % d-allose yield, suggesting that this mutant enzyme has a potential to be applied in d-allose production.

Caproic acid is the precursor of ethyl caproate, the main representative flavor substance of strong-flavor baijiu (SFB). Caproic acid-producing bacteria are considered to be the most important type of acid-producing microorganisms in the pit mud of the SFB ecosystem. In this study, the Rummeliibacillus suwonensis 3B-1 with a high yield of caproic acid (4.064 g/L) was screened from SFB pit mud. The genome of the R. suwonensis 3B-1 was sequenced, the total size was found to be 4117,671 bp and a calculated GC content of 35.86%. The caproic acid biosynthesis pathway was identified and analyzed, and it showed that 3B-1 could not only use ethanol, but it could also use glucose and other carbon sources as substrates to produce caproic acid. According to the genome analysis and with an optimized medium, the optimal conditions for caproic acid production were yeast powder at 3 g/L, sodium acetate at 15 g/L, and 1% biotin at 8 mL/100 mL. The yield of caproic acid reached 4.627 g/L, an increase of 13.9%, which was higher than that of general caproic acid bacteria. This is the first report of the synthesis of caproic acid by R. suwonensis. This strain could be used to produce caproic acid, an artificial pit mud preparation, and/or an enhanced inoculum in the production of SFB.

As a valuable platform chemical, 2,3-Butanediol (2,3-BDO) has a variety of industrial applications, and its microbial production is particularly attractive as an alternative to petroleum-based production. In this study, the regulation of intracellular carbon flux and NADH/NAD+ was used to increase the 2,3-BDO production of Enterobacter aerogenes. The genes encoding lactate dehydrogenase (ldh) and pyruvate formate lyase (pfl) were disrupted using the λ-Red recombination method and CRISPR-Cas9 to reduce the production of several byproducts and the consumption of NADH. Knockout of ldh or pfl increased intracellular NADH/NAD+ by 111 % and 113 %, respectively. Moreover, two important genes in the 2,3-BDO biosynthesis pathway, acetolactate synthase (budB) and acetoin reductase (budC), were overexpressed in E. aerogenes to further amply the metabolic flux toward 2,3-BDO production. And the overexpression of budB or budC increased intracellular NADH/NAD+ by 46 % and 57 %, respectively. In shake-flask cultivation with sucrose as carbon source, the 2,3-BDO titer of the IAM1183-LPBC was 3.55 times that of the wild type. In the 5-L fermenter, the maximal 2,3-BDO production produced by the IAM1183-LPBC was 2.88 times that of the original strain. This work offers new ideas for promoting the biosynthesis of 2,3-BDO for industrial applications.