Potential Biotechnological Applications Research Articles

Phytofabrication of silver nanoparticles using callus extracts of natural tetraploid Trifolium pratense L. and its bioactivities

One of the main subjects of plant biotechnology is plant tissue culture and in recent years is considered a possible approach model for green and eco-friendly biosynthesis of nanoparticles. This study aimed to present calli produced from the natural tetraploid Trifolium pratense L. containing high amounts of phenolic compounds and glycosidic bioactive macromolecules and the biosynthesis of silver nanoparticles from calli. Combinatorial optimization of silver nanoparticles was achieved for the first time in this study, thanks to the stabilizing and reducing properties of hypocotyl, apical meristem, and epicotyl derived callus extracts of the natural tetraploid T. pratense L. biosynthesized nanoparticles from three different callus extracts. Callus extracts were used to create different experiments with AgNO3 at various concentrations (0.16, 0.5, 0.84, 1.18, 1.52 and 1.96 mg L-1), different temperatures (40, 50, 60, 70, 80, 90, 100°C), and different pH levels (5, 7, 10) to carry out the biosynthesis of AgNPs. Biologically synthesized AgNPs were easily monitored by color change in ultraviolet and UV-Vis spectroscopy proved to be a fast and simple method. Also, TEM, XRD, and FTIR analyses were done to characterize and confirm the formation of crystalline nanoparticles. It was determined that antibacterial activity inhibition was achieved by using the Agar-well diffusion method for antibacterial activity measurements on Gram-positive Staphylococcus aureus ATCC 25923 and Gram-negative Escherichia coli CECT 4972 bacteria. Biosynthesized AgNPs were observed in the wavelength range of 400-500 nm in the UV-VIS spectrum. TEM analysis demonstrated the size and shape of biosynthesized silver nanoparticles under different conditions. It was observed that the smaller silver nanoparticles were spherical and the larger silver nanoparticles were triangular, elliptical, and spherical shape. The XRD analysis proved the presence of Ag0 in nanoparticles and showed crystal structure for silver nanoparticles. By FTIR analysis, O-H hydroxyl groups of functional groups on the AgNP surface, H-linked OH stretching, C-H stretching, -CH stretching of -CH2 and -CH3 functional groups, C-N and carboxylate, aliphatic phosphate and primary amine stretching were expressed. Biosynthesized silver nanoparticles showed antibacterial activity against Gram-positive S. aureus ATCC 25923 bacteria, AgNP hypocotyl (1.7mm), AgNP-epicotyl (1.1mm) against Gram-negative E. coli CECT 4972 bacteria. Among the hypocotyl, apical meristem, and epicotyl callus cultures, the highest antioxidant activity was observed in the AgNPs obtained from hypocotyl-concentration experiments, with a DPPH radical activity of 52% and an ABTS radical activity of 68%. In conclusion, these findings underscore the potential of biotechnological strategies in green nanotechnology, which can be offered for developing metal nanoparticles with potential biomedicine and biotechnology applications.

Elucidating the Role of Optical Activity of Polymers in Protein-Polymer Interactions.

Proteins are biomolecules with potential applications in agriculture, food sciences, pharmaceutics, biotechnology, and drug delivery. Interactions of hydrophilic and biocompatible polymers with proteins may impart proteolytic stability, improving the therapeutic effects of biomolecules and also acting as excipients for the prolonged storage of proteins under harsh conditions. The interactions of hydrophilic and stealth polymers such as poly(ethylene glycol), poly(trehalose), and zwitterionic polymers with various proteins are well studied. This study evaluates the molecular interactions of hydrophilic and optically active poly(vitamin B5 analogous methacrylamide) (poly(B5AMA)) with model proteins by fluorescence spectroscopy, nuclear magnetic resonance (NMR) spectroscopy, and circular dichroism (CD) spectroscopy analysis. The optically active hydrophilic polymers prepared using chiral monomers of R-(+)- and S-(-)-B5AMA by the photo-iniferter reversible addition fragmentation chain transfer (RAFT) polymerization showed concentration-dependent weak interactions of the polymers with bovine serum albumin and lysozyme proteins. Poly(B5AMA) also exhibited a concentration-dependent protein stabilizing effect at elevated temperatures, and no effect of the stereoisomers of polymers on protein thermal stability was observed. NMR analysis, however, showed poly(B5AMA) stereoisomer-dependent changes in the secondary structure of proteins.

Freshwater Sponges as a Neglected Reservoir of Bacterial Biodiversity.

Freshwater sponges (Spongillida: Demospongiae), including more than 240 described species, are globally distributed in continental waters (except for Antarctica), where they cover both natural and artificial surfaces. However, fragmentary studies have targeted their microbiome, making it difficult to test hypotheses about sponge-microbe specificity and metabolic relationships, along with the environmental factors playing key roles in structuring the associated microbial communities. To date, particular attention has been paid to sponges (family Lubomirskiidae) that are endemic to Lake Baikal. Few other freshwater sponge species (e.g., Ephydatia spp., Eunapius spp., and Spongilla lacustris), from lakes and rivers spanning from Europe to South and North America, have been targeted for microbiological studies. Representatives of the phyla Proteobacteria, Bacteroidetes, and Actinobacteria largely predominated, and high differences were reported between the microbiome of freshwater and marine sponges. Several bacterial strains isolated from freshwater sponges can produce bioactive compounds, mainly showing antibiotic activities, with potential application in biotechnology. Understanding the roles played by sponge microbiomes in freshwater ecosystems is still in its infancy and has yet to be clarified to disentangle the ecological and evolutionary significance of these largely under-investigated microbial communities. This review was aimed at providing the main available information on the composition and biotechnological potential of prokaryotic communities associated with healthy freshwater sponges, as a neglected component of the global sponge microbiome, to stimulate researchers interested in the field.

Spectroelectrochemistry for determination of the redox potential in heme enzymes: Dye-decolorizing peroxidases

Dye-decolorizing peroxidases (DyPs) are heme-containing enzymes that are structurally unrelated to other peroxidases. Some DyPs show high potential for applications in biotechnology, which critically depends on the stability and redox potential (E°') of the enzyme. Here we provide a comparative analysis of UV–Vis- and surface-enhanced resonance Raman-based spectroelectrochemical methods for determination of the E°' of DyPs from two different organisms, and their variants generated targeting E°' upshift. We show that substituting the highly conserved Arginine in the distal side of the heme pocket by hydrophobic amino acid residues impacts the heme architecture and redox potential of DyPs from the two organisms in a very distinct manner. We demonstrate the advantages and drawbacks of the used spectroelectrochemical approaches, which is relevant for other heme proteins that contain multiple heme centers or spin populations.

Functional characterization and structural insights of three cutinases from the ascomycete Fusarium verticillioides

Cutinases are serine esterases that belong to the α/β hydrolases superfamily. The natural substrates for these enzymes are cutin and suberin, components of the plant cuticle, the first barrier in the defense system against pathogen invasion. It is well-reported that plant pathogens produce cutinases to facilitate infection. Fusarium verticillioides, one important corn pathogens, is an ascomycete upon which its cutinases are poorly explored. Consequently, the objective of this study was to perform the biochemical characterization of three precursor cutinases (FvCut1, FvCut2, and FvCut3) from F. verticillioides and to obtain structural insights about them. The cutinases were produced in Escherichia coli and purified. FvCut1, FvCut2, and FvCut3 presented optimal temperatures of 20, 40, and 35 °C, and optimal pH of 9, 7, and 8, respectively. Some chemicals stimulated the enzymatic activity. The kinetic parameters revealed that FvCut1 has higher catalytic efficiency (Kcat/Km) in the p-nitrophenyl-butyrate (p-NPB) substrate. Nevertheless, the enzymes were not able to hydrolyze polyethylene terephthalate (PET). Furthermore, the three-dimensional models of these enzymes showed structural differences among them, mainly FvCut1, which presented a narrower opening cleft to access the catalytic site. Therefore, our study contributes to exploring the diversity of fungal cutinases and their potential biotechnological applications.

Bacterial community structure, adaptations and prevalence of antimicrobial resistance in bacteria from Antarctica: A review

Antarctica serves as an ideal prototype to study past climatic condition as the ice core records dates back to 100,000 years. Microbiological research on ice core has opened a doorway to our understanding on biodiversity and its contribution to the frozen ecosystem. In this review, the ice core bacterial diversity of East and West Antarctica has been documented. Cold-adapted bacteria hold immense potential in biotechnological applications as they possess functional advantages over mesophilic bacteria. Some of the neoteric applications of bioproducts of Antarctic bacteria have been stated in this review. The current climate change due to global warming exerts a humongous effect on the ecosystem of the polar region. The region being susceptible to any fluctuation in temperature, the exacerbated melting of Antarctic glaciated mass contributes to the rise of global sea level, with loss of ice harbouring unknown microbial entities. This review mainly focuses on the bacterial diversity of East and West Antarctic counterparts, the prevalence of antibiotic-resistant genes, the adaption strategies conferred by bacteria thriving in Antarctic habitats and their potential biotechnological applications.

Diatom-Bacteria Interactions in the Marine Environment: Complexity, Heterogeneity, and Potential for Biotechnological Applications.

Diatom-bacteria interactions evolved during more than 200 million years of coexistence in the same environment. In this time frame, they established complex and heterogeneous cohorts and consortia, creating networks of multiple cell-to-cell mutualistic or antagonistic interactions for nutrient exchanges, communication, and defence. The most diffused type of interaction between diatoms and bacteria is based on a win-win relationship in which bacteria benefit from the organic matter and nutrients released by diatoms, while these last rely on bacteria for the supply of nutrients they are not able to produce, such as vitamins and nitrogen. Despite the importance of diatom-bacteria interactions in the evolutionary history of diatoms, especially in structuring the marine food web and controlling algal blooms, the molecular mechanisms underlying them remain poorly studied. This review aims to present a comprehensive report on diatom-bacteria interactions, illustrating the different interplays described until now and the chemical cues involved in the communication and exchange between the two groups of organisms. We also discuss the potential biotechnological applications of molecules and processes involved in those fascinating marine microbial networks and provide information on novel approaches to unveiling the molecular mechanisms underlying diatom-bacteria interactions.

Crystal structure and interconversion of monomers and domain-swapped dimers of the walnut tree phytocystatin

Biotechnological applications of phytocystatins have garnered significant interest due to their potential applications in crop protection and improve crop resistance to abiotic stress factors. Cof1 and Wal1 are phytocystatins derived from Coffea arabica and Juglans regia, respectively. These plants hold significant economic value due to coffee's global demand and the walnut tree's production of valuable timber and widely consumed walnuts with culinary and nutritional benefits. The study involved the heterologous expression in E. coli Lemo 21(DE3), purification by immobilized metal ion affinity and size exclusion chromatography, and biophysical characterization of both phytocystatins, focusing on isolating and interconverting their monomers and dimers. The crystal structure of the domain-swapped dimer of Wal1 was determined revealing two domain-swapped dimers in the asymmetric unit, an arrangement reminiscent of the human cystatin C structure. Alphafold models of monomers and Alphafold-Multimer models of domain-swapped dimers of Cof1 and Wal1 were analyzed in the context of the crystal structure. The methodology and data presented here contribute to a deeper understanding of the oligomerization mechanisms of phytocystatins and their potential biotechnological applications in agriculture.

Insights into Genomic Features and Potential Biotechnological Applications of Bacillus halotolerans Strain HGR5.

Algeria is one of the wealthiest countries in terms of hydrothermal sources, with more than two hundred hot springs. However, diverse and little-described microbial communities colonize these habitats, making them an intriguing research subject. This work reports the isolation of bacteria from two hot springs water samples in northeastern Algeria, evaluating their enzymatic activities and effect on plant pathogens. Out of the obtained 72 bacterial isolates and based on the 16S rRNA gene sequence analysis, the strain HGR5 belonging to Bacillus halotolerans had the most interesting activity profile. Interestingly, HGR5 was substantially active against Fusarium graminearum, Phytophthora infestans, and Alternaria alternata. Furthermore, this strain presented a high ability to degrade casein, Tween 80, starch, chitin, cellulose, and xylan. The genome sequence of HGR5 allowed taxonomic validation and screening of specific genetic traits, determining its antagonistic and enzymatic activities. Genome mining revealed that strain HGR5 encloses several secondary metabolite biosynthetic gene clusters (SM-BGCs) involved in metabolite production with antimicrobial properties. Thus, antimicrobial metabolites included bacillaene, fengycin, laterocidine, bacilysin, subtilosin, bacillibactin, surfactin, myxovirescin, dumulmycin, and elansolid A1. HGR5 strain genome was also mined for CAZymes associated with antifungal activity. Finally, the HGR5 strain exhibited the capacity to degrade polycaprolactone (PCL), a model substrate for polyester biodegradation. Overall, these results suggest that this strain may be a promising novel biocontrol agent with interesting plastic-degradation capability, opening the possibilities of its use in various biotechnological applications.

ABZYMES: A Comprehensive Review of Applications and Advances

Abzymes, or antibody enzymes, are antibodies that have enzymatic activity. They can catalyze chemical reactions, similar to traditional enzymes. Abzymes have potential applications in medicine and biotechnology, as they can be engineered to target specific molecules, making them promising for therapies and diagnostics. They often called catmab (from catalytic monoclonal antibody).They can be artificial or naturally occurring, even in autoimmune diseases. They function by stabilizing reaction intermediates, making them valuable tools in biotechnology

Production of fungal phytases in solid state fermentation and potential biotechnological applications.

Phytases are important enzymes used for eliminating the anti-nutritional properties of phytic acid in food and feed ingredients. Phytic acid is major form of organic phosphorus stored during seed setting. Monogastric animals cannot utilize this phytate-phosphorus due to lack of necessary enzymes. Therefore, phytic acid excretion is responsible for mineral deficiency and phosphorus pollution. Phytases have been reported from diverse microorganisms, however, fungal phytases are preferred due to their unique properties. Aspergillus species are the predominant producers of phytases and have been explored widely as compared to other fungi. Solid-state fermentation has been studied as an economical process for the production of phytases to utilize various agro-industrial residues. Mixed substrate fermentation has also been reported for the production of phytases. Physical and chemical parameters including pH, temperature, and concentrations of media components have significantly affected the production of phytases in solid state fermentation. Fungi produced high levels of phytases in solid state fermentation utilizing economical substrates. Optimization of culture conditions using different approaches has significantly improved the production of phytases. Fungal phytases are histidine acid phosphatases exhibiting broad substrate specificity, are relatively thermostable and protease-resistant. These phytases have been found effective in dephytinization of food and feed samples with concomitant liberation of minerals, sugars and soluble proteins. Additionally, they have improved the growth of plants by increasing the availability of phosphorus and other minerals. Furthermore, phytases from fungi have played an important roles in bread making, semi-synthesis of peroxidase, biofuel production, production ofmyo-inositol phosphates and management of environmental pollution. This review article describes the production of fungal phytases in solid state fermentation and their biotechnological applications.

Genome sequencing and metabolic network reconstruction of a novel sulfur-oxidizing bacterium Acidithiobacillus Ameehan.

Sulfur-oxidizing bacteria play a crucial role in various processes, including mine bioleaching, biodesulfurization, and treatment of sulfur-containing wastewater. Nevertheless, the pathway involved in sulfur oxidation is highly intricate, making it complete comprehension a formidable and protracted undertaking. The mechanisms of sulfur oxidation within the Acidithiobacillus genus, along with the process of energy production, remain areas that necessitate further research and elucidation. In this study, a novel strain of sulfur-oxidizing bacterium, Acidithiobacillus Ameehan, was isolated. Several physiological characteristics of the strain Ameehan were verified and its complete genome sequence was presented in the study. Besides, the first genome-scale metabolic network model (AMEE_WP1377) was reconstructed for Acidithiobacillus Ameehan to gain a comprehensive understanding of the metabolic capacity of the strain.The characteristics of Acidithiobacillus Ameehan included morphological size and an optimal growth temperature range of 37-45°C, as well as an optimal growth pH range of pH 2.0-8.0. The microbe was found to be capable of growth when sulfur and K2O6S4 were supplied as the energy source and electron donor for CO2 fixation. Conversely, it could not utilize Na2S2O3, FeS2, and FeSO4·7H2O as the energy source or electron donor for CO2 fixation, nor could it grow using glucose or yeast extract as a carbon source. Genome annotation revealed that the strain Ameehan possessed a series of sulfur oxidizing genes that enabled it to oxidize elemental sulfur or various reduced inorganic sulfur compounds (RISCs). In addition, the bacterium also possessed carbon fixing genes involved in the incomplete Calvin-Benson-Bassham (CBB) cycle. However, the bacterium lacked the ability to oxidize iron and fix nitrogen. By implementing a constraint-based flux analysis to predict cellular growth in the presence of 71 carbon sources, 88.7% agreement with experimental Biolog data was observed. Five sulfur oxidation pathways were discovered through model simulations. The optimal sulfur oxidation pathway had the highest ATP production rate of 14.81 mmol/gDW/h, NADH/NADPH production rate of 5.76 mmol/gDW/h, consumed 1.575 mmol/gDW/h of CO2, and 1.5 mmol/gDW/h of sulfur. Our findings provide a comprehensive outlook on the most effective cellular metabolic pathways implicated in sulfur oxidation within Acidithiobacillus Ameehan. It suggests that the OMP (outer-membrane proteins) and SQR enzymes (sulfide: quinone oxidoreductase) have a significant impact on the energy production efficiency of sulfur oxidation, which could have potential biotechnological applications.

Microbial diversity and community structure dynamics in acid mine drainage: Acidic fire with dissolved heavy metals

Acid mine drainage (AMD) is one of the leading causes of environmental pollution and is linked to public health and ecological consequences. Microbes-mineral interaction generates AMD, but microorganisms can also remedy AMD pollution. Exploring the microbial response to AMD effluents may reveal survival strategies in extreme ecosystems. Three distinct sites across a mine (inside the mine, the entrance of the mine, and outside) were selected to study their heavy metal concentrations due to significant variations in pH and physicochemical characteristics, and high-throughput sequencing was carried out to investigate the microbial diversity. The metal and ion concentrations followed the order SO42−, Fe, Cu, Zn, Mg, Pb, Co, Cr, and Ni from highest to lowest, respectively. Maximum sequences were allocated to Proteobacteria and Firmicutes. Among archaea, the abundance of Thaumarchaeota and Euryarchaeota was higher outside of mine. Most of the genera (23.12 %) were unclassified and unknown. The average OTUs (operational taxonomic units) were significantly higher outside the mine; however, diversity indices were not significantly different across the mine sites. Hierarchical clustering of selective genera and nMDS ordination of OTUs displayed greater segregation resolution inside and outside of mine, whereas the entrance samples clustered with greater similarity. Heterogeneous selection might be the main driver of community composition outside the mine, whereas stochastic processes became prominent inside the mine. However, the ANOSIM test shows a relatively even distribution of community composition within and between the groups. Microbial phyla showed both positive and negative correlations with physicochemical factors. A greater number of biomarkers were reported outside of the mine. Predictive functional investigation revealed the existence of putative degradative, metabolic, and biosynthetic pathways. This study presents a rare dataset in our understanding of microbial diversity and distribution as shaped by the ecological gradient and potential novelty in phylogenetic/taxonomic diversity in AMD, with potential biotechnological applications.

Development of an efficient protein expression system in the thermophilic fungus Myceliophthora thermophila

BackgroundThermophilic fungus Myceliophthora thermophila has been widely used in industrial applications due to its ability to produce various enzymes. However, the lack of an efficient protein expression system has limited its biotechnological applications.ResultsIn this study, using a laccase gene reporting system, we developed an efficient protein expression system in M. thermophila through the selection of strong constitutive promoters, 5’UTRs and signal peptides. The expression of the laccase was confirmed by enzyme activity assays. The results showed that the Mtpdc promoter (Ppdc) was able to drive high-level expression of the target protein in M. thermophila. Manipulation of the 5’UTR also has significant effects on protein expression and secretion. The best 5’UTR (NCA-7d) was identified. The transformant containing the laccase gene under the Mtpdc promoter, NCA-7d 5’UTR and its own signal peptide with the highest laccase activity (1708 U/L) was obtained. In addition, the expression system was stable and could be used for the production of various proteins, including homologous proteins like MtCbh-1, MtGh5-1, MtLPMO9B, and MtEpl1, as well as a glucoamylase from Trichoderma reesei.ConclusionsAn efficient protein expression system was established in M. thermophila for the production of various proteins. This study provides a valuable tool for protein production in M. thermophila and expands its potential for biotechnological applications.

CuI Coordination Polymer‐Catalyzed Azide–Alkyne Cycloaddition Click Reaction at Room Temperature

AbstractFramework based compounds for eco‐friendly catalysis is of utmost industrial interest. CuI‐catalyzed “click” reactions are a class of highly efficient and widely used chemical reactions in organic synthesis for potential applications in biotechnology, medicine, polymer, and material sciences. The importance of click chemistry also acknowledge by 2022 Noble Prize in chemistry. One of the most well‐known click reactions is the copper‐catalyzed azide‐alkyne cycloaddition (CuAAC), which is a type of Huisgen 1,3‐dipolar cycloaddition reaction. Herein, three stable copper(I)‐coordination polymers (CPs), [Cu(4‐ABPT)Cl], CP‐1; [Cu(4‐ABPT)Br], CP‐2; [Cu2(4‐ABPT)0.5I2], CP‐3; (4‐ABPT=3,5‐di(pyridine‐4‐yl)‐4H‐1,2,4‐triazol‐4‐amine) have been chosen for CuAAC reaction, due to presence of both Lewis acidic (copper ion), and basic (pyridyl and amine) sites. The counter anion plays a crucial role in their structural motif as well as catalytic efficiency. These easily synthesizable and scalable catalysts exhibit the trend of CP‐1>CP‐2>CP‐3 in catalysis for the selective conversion of phenylacetylene, benzyl bromide, and sodium azide into 1,4‐cycloaddition product within 2 h at ambient temperature and solvent free reaction conditions. CuAAC reaction using catalyst CP‐1 exhibits near quantitative conversion within 2 h at room temperature in solvent free conditions. Herein, we demonstrated the bulk‐scale synthesis of CuI CPs for efficient azide‐alkyne cycloaddition click reaction under mild reaction conditions.

Delving into the lifestyle of Sundarban Wetland resident, biofilm producing, halotolerant Salinicoccus roseus: a comparative genomics-based intervention

BackgroundMicrobial community played an essential role in ecosystem processes, be it mangrove wetland or other intertidal ecologies. Several enzymatic activities like hydrolases are effective ecological indicators of soil microbial function. So far, little is known on halophilic bacterial contribution and function on a genomic viewpoint of Indian Sundarban Wetland. Considering the above mentioned issues, the aims of this study was to understand the life style, metabolic functionalities and genomic features of the isolated bacterium, Salinicoccus roseus strain RF1H. A comparative genome-based study of S. roseus has not been reported yet. Henceforth, we have considered the inclusion of the intra-species genome comparison of S. roseus to gain insight into the high degree of variation in the genome of strain RF1H among others.ResultsSalinicoccus roseus strain RF1H is a pink-red pigmented, Gram-positive and non-motile cocci. The bacterium exhibited high salt tolerance (up to 15% NaCl), antibiotic resistance, biofilm formation and secretion of extracellular hydrolytic enzymes. The circular genome was approximately 2.62978 Mb in size, encoding 574 predicted genes with GC content 49.5%. Presence of genomic elements (prophages, transposable elements, CRISPR-Cas system) represented bacterial virulence and multidrug-resistance. Furthermore, genes associated with salt tolerance, temperature adaptation and DNA repair system were distributed in 17 genomic islands. Genes related to hydrocarbon degradation manifested metabolic capability of the bacterium for potential biotechnological applications. A comparative pangenome analysis revealed two-component response regulator, modified C4-dicarboxylate transport system and osmotic stress regulated ATP-binding proteins. Presence of genes encoding arginine decarboxylase (ADC) enzyme being involved in biofilm formation was reported from the genome. In silico study revealed the protein is thermostable and made up with ~ 415 amino acids, and hydrophilic in nature. Three motifs appeared to be evolutionary conserved in all Salinicoccus sequences.ConclusionThe first report of whole genome analysis of Salinicoccus roseus strain RF1H provided information of metabolic functionalities, biofilm formation, resistance mechanism and adaptation strategies to thrive in climate-change induced vulnerable spot like Sundarban. Comparative genome analysis highlighted the unique genome content that contributed the strain’s adaptability. The biomolecules produced during metabolism are important sources of compounds with potential beneficial applications in pharmaceuticals.

Sustainable Immobilization of β-Glucosidase onto Silver Ions and AgNPs-Loaded Acrylic Fabric with Enhanced Stability and Reusability.

Modified polymer design has attracted significant attention for enzyme immobilization, offering promising applications. In this study, amine-terminated polymers were synthesized by incorporating functional groups into polyacrylonitrile using hexamethylenediamine. This work highlights the successful enzyme immobilization strategy using modified polymers, offering improved stability and expanded operational conditions for potential biotechnological applications. The resulting amino groups were utilized to capture silver ions, which were subsequently converted to silver nanoparticles (AgNPs). The obtained materials, AgNPs@TA-HMDA (acrylic textiles coated silver nanoparticles AgNPs) and Ag(I)@TA-HMDA (acrylic textiles coated with Ag ion) were employed as supports for β-glucosidase enzyme immobilization. The highest immobilization yields (IY%) were achieved with AgNPs@TA-HMDA at 92%, followed by Ag(I)@TA-HMDA at 79.8%, resulting in activity yields (AY%) of 81% and 73%, respectively. Characterization techniques such as FTIR, FE-SEM, EDX, TG/DTG, DSC, and zeta potential were employed to investigate the structural composition, surface morphologies, elemental composition, thermal properties, and surface charge of the support materials. After 15 reuses, the preservation percentages decreased to 76% for AgNPs@TA-HMDA/β-Glu and 65% for Ag(I)@TA-HMDA/β-Glu. Storage stability revealed that the decrease in activity for the immobilized enzymes was smaller than the free enzyme. The optimal pH for the immobilized enzymes was broader (pH 5.5 to 6.5) compared to the free enzyme (pH 5.0), and the optimal temperature for the immobilized enzymes was 60 °C, slightly higher than the free enzyme's optimal temperature of 50 °C. The kinetic analysis showed a slight increase in Michaelis constant (Km) values for the immobilized enzymes and a decrease in maximum velocity (Vmax), turnover number (Kcat), and specificity constant (Kcat/Km) values compared to the free enzyme. Through extensive characterization, we gained valuable insights into the structural composition and properties of the modified polymer supports. This research significantly contributes to the development of efficient biotechnological processes by advancing the field of enzyme immobilization and offering valuable knowledge for its potential applications.

Growth, morphology, and formation of cinnabarin in Pycnoporus cinnabarinus in relation to different irradiation spectra

BackgroundThe demand for natural pigments in general, and for fungi-derived pigments in particular, is constantly rising. Wood-decomposing fungi represent a promising source for natural pigments and they are usually easy to cultivate in pure culture. One of them, i.e., Pycnoporus cinnabarinus, offers a highly interesting spectrum of bioactivity, partly due to the formation of the orange–red pigment cinnabarin. However, apart from a few studies addressing its diverse potential biotechnological applications, there is still a large gap of knowledge concerning the influence of light on the formation of cinnabarin. The aim of this work was to investigate the effect of different irradiations on the cinnabarin content, the growth, and the morphology of three different P. cinnabarinus strains. We used highly standardized irradiation conditions and cultivation techniques in combination with newly developed methods for the extraction and direct quantification of cinnabarin.ResultsRed, green, blue, and UV-A irradiation (mean irradiance Ee = 1.5 ± 0.18 W m−2) had considerable effects on the growth and colony appearance of all three P. cinnabarinus strains tested. The cinnabarin content determined was, thus, dependent on the irradiation wavelength applied, allowing strain-specific thresholds to be defined. Irradiation with wavelengths below this strain-specific threshold corresponded to a lower cinnabarin content, at least at the intensity applied. The orange–red pigment appeared by light microscopy as incrusted extracellular plaques present on the hyphal walls. Highly efficient vegetative propagation occurred by arthroconidia, and we observed the tendency that this asexual reproduction was (i) most frequent in the dark but (ii) never occurred under UV-A exposure.Conclusion This study highlights a differential photo-dependence of growth, morphology, and cinnabarin formation in P. cinnabarinus. This confirms that it is advisable to consider the wavelength of the light used in future biotechnological productions of natural pigments.Graphical abstract

Biofabrication of Mg-doped ZnO nanostructures for hemolysis and antibacterial properties.

The aim of this work was to synthesize 0.02 and 0.06Mg-doped ZnO nanoparticles (NPs) using the aqueous extract of Plectranthus barbatus leaf. The structural integrity of the hexagonal phase was emphasized by X-ray diffraction analysis. The average crystallite size (D) of 0.02 and 0.06Mg-doped ZnO NPs was found to be 23.83 and 26.95nm, respectively. The scanning electron microscope images revealed a surface morphology of irregular nano-shapes of about 83nm diameter with an elongated one-dimensional structure. The hemolysis activity demonstrated the safe nature of the synthesized materials at low doses. Antibacterial activity against S. aureus and E. coli, which assessed using the disc diffusion method, indicated that the prepared NPs could inhibit S. aureus but not E. coli. These findings suggest that the synthesized NPs could be explored for potential applications in biotechnology and medicine.

Computational analysis of carboxylesterase genes and proteins in non-pathogenic food bacterium Alicyclobacillus acidocaldarius: insights from proteogenomics.

Over recent years, Alicyclobacillus acidocaldarius, a Gram-positive nonpathogenic rod-shaped thermo-acid-tolerant bacterium, has posed numerous challenges for the fruit juice industry. However, the bacterium's unique characteristics, particularly its nonpathogenic and thermophilic capabilities, offer significant opportunities for genetic exploration by biotechnologists. This study presents the computational proteogenomics report on the carboxylesterase (CE) enzyme in A. acidocaldarius, shedding light on structural and evolutional of CEs from this bacterium. Our analysis revealed that the average molecular weight of CEs in A. acidocaldarius was 41kDa, with an isoelectric point around 5. The amino acid composition favored negative amino acids over positive ones. The aliphatic index and hydropathicity were approximately 88 and - 0.15, respectively. While the protein sequence showed no disulfide bonds in the CEs' structure, the presence of Cys amino acids was observed in the structure of CEs. Phylogenetic analysis presented more than 99% similarity between CEs, indicating their close evolutionary relationship. By applying homology modeling, the 3-dimensional structural models of the carboxylesterase were constructed, which with the help of structural conservation and solvent accessibility analysis highlighted key residues and regions responsible for enzyme stability and conformation. The specific patterns presented the total solvent accessibility of less than 25 (Å2) was in considerable position as well as Gly residues were noticeably have high accessibility to solvent in all structures. Ala was the more frequent amino acids in the conserved-SASA of carboxylesterases. Furthermore, unsupervised agglomerative hierarchical clustering based on solvent accessibility feature successfully clustered and even distinguished this enzyme from proteases from the same genome. These findings contribute to a deeper understanding of the nonpathogenic A. acidocaldarius carboxylesterase and its potential applications in biotechnology. Additionally, structural analysis of CEs would help to address potential solutions in fruit juice industry with utilization of computational structural biology.