Chemically modified nylons as supports for enzyme immobilization. Polyisonitrile-nylon.
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ABSTRACT:
SUBMITTER: Goldstein L
PROVIDER: S-EPMC1168419 | biostudies-other | 1974 Dec
REPOSITORIES: biostudies-other
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Project description:BackgroundChemistry and particularly enzymology at surfaces is a topic of rapidly growing interest, both in terms of its role in biological systems and its application in biocatalysis. Existing protein immobilization approaches, including noncovalent or covalent attachments to solid supports, have difficulties in controlling protein orientation, reducing nonspecific absorption and preventing protein denaturation. New strategies for enzyme immobilization are needed that allow the precise control over orientation and position and thereby provide optimized activity.Methodology/principal findingsA method is presented for utilizing peptide ligands to immobilize enzymes on surfaces with improved enzyme activity and stability. The appropriate peptide ligands have been rapidly selected from high-density arrays and when desirable, the peptide sequences were further optimized by single-point variant screening to enhance both the affinity and activity of the bound enzyme. For proof of concept, the peptides that bound to ?-galactosidase and optimized its activity were covalently attached to surfaces for the purpose of capturing target enzymes. Compared to conventional methods, enzymes immobilized on peptide-modified surfaces exhibited higher specific activity and stability, as well as controlled protein orientation.Conclusions/significanceA simple method for immobilizing enzymes through specific interactions with peptides anchored on surfaces has been developed. This approach will be applicable to the immobilization of a wide variety of enzymes on surfaces with optimized orientation, location and performance, and provides a potential mechanism for the patterned self-assembly of multiple enzymes on surfaces.
| S-EPMC3073003 | biostudies-literature
Project description:The high porosity, interconnected pore structure, and high surface area-to-volume ratio make the hydrophilic nonwoven nanofiber membranes (NV-NF-Ms) promising nanostructured supports for enzyme immobilization in different biotechnological applications. In this work, NV-NF-Ms with excellent mechanical and chemical properties were designed and fabricated by electrospinning in one step without using additives or complicated crosslinking processes after electrospinning. To do so, two types of ultrahigh-molecular-weight linear copolymers with very different mechanical properties were used. Methyl methacrylate-co-hydroxyethyl methacrylate (p(MMA)-co-p(HEMA)) and methyl acrylate-co-hydroxyethyl acrylate (p(MA)-co-p(HEA)) were designed and synthesized by reverse atom transfer radical polymerization (reverse-ATRP) and copper-mediated living radical polymerization (Cu0-MC-LRP), respectively. The copolymers were characterized by nuclear magnetic resonance (1H-NMR) spectroscopy and by triple detection gel permeation chromatography (GPC). The polarity, topology, and molecular weight of the copolymers were perfectly adjusted. The polymeric blend formed by (MMA)1002-co-(HEMA)1002 (M w = 230,855 ± 7418 Da; M n = 115,748 ± 35,567 Da; PDI = 2.00) and (MA)11709-co-(HEA)7806 (M w = 1.972 × 106 ± 33,729 Da; M n = 1.395 × 106 ± 35,019 Da; PDI = 1.41) was used to manufacture (without additives or chemical crosslinking processes) hydroxylated nonwoven nanofiber membranes (NV-NF-Ms-OH; 300 nm in fiber diameter) with excellent mechanical and chemical properties. The morphology of NV-NF-Ms-OH was studied by scanning electron microscopy (SEM). The suitability for enzyme binding was proven by designing a palette of different surface functionalization to enable both reversible and irreversible enzyme immobilization. NV-NF-Ms-OH were successfully functionalized with vinyl sulfone (281 ± 20 μmol/g), carboxyl (560 ± 50 μmol/g), and amine groups (281 ± 20 μmol/g) and applied for the immobilization of two enzymes of biotechnological interest. Galactose oxidase was immobilized on vinyl sulfone-activated materials and carboxyl-activated materials, while laccase was immobilized onto amine-activated materials. These preliminary results are a promising basis for the application of nonwoven membranes in enzyme technology.
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Project description:Enzyme immobilization is a powerful strategy for enzyme stabilization and recyclability. Materials covered with multipoint molecules are very attractive for this goal, since the number of active moieties to attach the enzyme increases with respect to monofunctional linkers. This work evaluates different dendrimers supported on silica to immobilize a protease enzyme, Alcalase. Five different dendrimers were employed: two carbosilane (CBS) dendrimers of different generations (SiO2-G0Si-NH2 and SiO2-G1Si-NH2), a CBS dendrimer with a polyphenoxo core (SiO2-G1O3-NH2), and two commercial polyamidoamine (PAMAM) dendrimers of different generations (SiO2-G0PAMAM-NH2 and SiO2-G1PAMAM-NH2). The results were compared with a silica support modified with a monofunctional molecule (2-aminoethanethiol). The effect of the dendrimer generation, the immobilization conditions (immobilization time, Alcalase/SiO2 ratio, and presence of Ca2+ ions), and the digestion conditions (temperature, time, amount of support, and stirring speed) on Alcalase activity has been evaluated. Enzyme immobilization and its activity were highly affected by the kind of dendrimer and its generation, observing the most favorable behavior with SiO2-G0PAMAM-NH2. The enzyme immobilized on this support was used in two consecutive digestions and, unlike CBS supports, it did not retain peptides released in the digestion.
| S-EPMC9783553 | biostudies-literature
Project description:A highly effective method was developed to functionalize ceramic supports (Al2O3 powders and membranes) using newly synthesized spacer molecules. The functionalized materials were subsequently utilized for Candida antarctica lipase B enzyme immobilization. The objective is to systematically evaluate the impact of various spacer molecules grafted onto the alumina materials will affect both the immobilization of the enzymes and specific material surface properties, critical to enzymatic reactors performance. The enzyme loading was significantly improved for the supports modified with shorter spacer molecules, which possessed higher grafting effectiveness on the order of 90%. The specific enzyme activity was found to be much higher for samples functionalized with longer modifiers yielding excellent enantioselectivity >97%. However, the enantiomeric ratio of the immobilized lipase was slightly lower in the case of shorter spacer molecules.
| S-EPMC7794798 | biostudies-literature
Project description:In this research, it has been demonstrated that amino-modified microspheres (A-LMS) based on bio-waste derived material, such as kraft lignin, have good prospects in usage as a support for enzyme immobilization, since active biocatalyst systems were prepared by immobilizing β-galactosidase from A. oryzae and laccase from M. thermophila expressed in A. oryzae (Novozym® 51003) onto A-LMS. Two types of A-LMS were investigated, with different emulsifier concentrations (5 wt% and 10 wt%), and microspheres produced using 5 wt% of emulsifier (A-LMS_5) showed adequate pore shape, size and distribution for enzyme attachment. The type of interactions formed between enzymes (β-galactosidase and laccase) and A-LMS_5 microspheres demonstrated that β-galactosidase is predominantly attached via electrostatic interactions while attachment of laccase is equally governed by electrostatic and hydrophobic interactions. Furthermore, the A-LMS_5-β-galactosidase exhibited specificity towards recognized prebiotics (galacto-oligosaccharides (GOS)) synthesis with 1.5-times higher GOS production than glucose production, while for environmental pollutant lindane degradation, the immobilized laccase preparation exhibited high activity with a minimum remaining lindane concentration of 22.4% after 6 days. Thus, this novel enzyme immobilization support A-LMS_5 has potential for use in green biotechnologies.
| S-EPMC9054402 | biostudies-literature
Project description:Biochar has received widespread attention as an eco-friendly and efficient material for immobilization of toxic heavy metals in aqueous environments. In the present study, three types of coconut fiber-derived biochars were obtained by pyrolyzing at three temperatures, i.e., 300, 500, and 700 °C. In addition, nine types of biochars were prepared by chemical modification with ammonia, hydrogen peroxide, and nitric acid, respectively, which were used to investigate changes in physico-chemical properties by inter alia, Fourier transformation infrared spectrophotometry (FTIR), scanning electron microscope (SEM), and BET specific surface area analysis. Batch sorption experiments were carried out to determine the sorption capacity of the biochars for lead (Pb) in aqueous solutions. Results showed that the cation exchange capacity of biochar pyrolyzed at 300 °C and modified with nitric acid increased threefold compared to the control. Loosely corrugated carbon surface and uneven carbon surface of the biochar pyrolyzed at 300 °C were produced during ammonia and nitric acid modifications. Removal rate of Pb by the coconut biochar pyrolyzed at 300 °C and modified with ammonia was increased from 71.8 to 99.6 % compared to the untreated biochar in aqueous solutions containing 100 mg L-1 Pb. However, chemical modification did not enhance adsorption of Pb of the biochars pyrolyzed at higher temperatures (e.g., 500 or 700 °C), indicating that resistance of biochars to chemical treatment increased with pyrolysis temperature.
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Project description:Poly(ethylene terephthalate) (PET) and nylon find their main applications in working clothes, domestic furniture and as indoor decoration (curtains and carpets). The increasing attention on healthy lifestyle, together with protection and safety, gained a strong interest in today's society. In this context, reducing the flammability of textiles has been tackled by designing flame retardants (FRs) able to suppress or delay the flame propagation. Commercially available FRs for textiles often consist of brominated, chlorinated and organo-phosphorus compounds, which are considered a great concern for human health and for the environment. In this study, Deoxyribose Nucleic Acid (DNA) was investigated as a green and eco-friendly alternative to halogen-containing FRs. DNA is in fact able to provide flame retardant properties due to its intrinsically intumescent building blocks (deoxyribose, phosphoric-polyphosphoric acid, and nitrogen-containing bases). In a first step, anchor groups (i.e., carboxyl groups) for subsequent coupling of DNA were introduced to PET and nylon-6 fabrics via limited surface hydrolysis with Humicola insolens cutinase (HiC). Released monomer/oligomers were measured via HPLC (1 mM of BHET for PET and 0.07 mM of caprolactam from nylon after 72 h). In a next step, DNA immobilization on the activated polymers was studied by using three different coupling systems, namely: EDC/NHS, dopamine, and tyrosine. DNA coupling was confirmed via FT-IR that showed typical bands at 1,220, 970, and 840 cm-1. The tyrosine/DNA coupling on nylon fabrics resulted to be the most effective as certified by the lowest burning rate and total burning time (35 s, 150 mm, and 4.3 mm*s-1 for the blank and 3.5 s, 17.5 mm, and 5 mm* s-1 for nylon/tyrosine/DNA) which was also confirmed by FT-IR and ESEM/EDS measurements. Thermogravimetric analysis (TGA) further confirmed that tyrosine/DNA coated nylon showed a lower thermal degradation between 450 and 625°C when compared to the untreated samples.
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Project description:In recent years, 3D printing has emerged in the field of chemical engineering as a powerful manufacturing technique to rapidly design and produce tailor-made reaction equipment. In fact, reactors with complex internal geometries can be easily fabricated, optimized and interchanged in order to respond to precise process needs, such as improved mixing and increased surface area. These advantages make them interesting especially for catalytic applications, since customized structured bed reactors can be easily produced. 3D printing applications are not limited to reactor design, it is also possible to realize functional low cost alternatives to analytical equipment that can be used to increase the level of process understanding while keeping the investment costs low. In this work, in-house designed ceramic structured inserts printed via vat photopolymerization (VPP) are presented and characterized. The flow behavior inside these inserts was determined with residence time distribution (RTD) experiments enabled by in-house designed and 3D printed inline photometric flow cells. As a proof of concept, these structured inserts were fitted in an HPLC column to serve as solid inorganic supports for the immobilization of the enzyme Phenolic acid Decarboxylase (bsPAD), which catalyzes the decarboxylation of cinnamic acids. The conversion of coumaric acid to vinylphenol was chosen as a model system to prove the implementation of these engineered inserts in a continuous biocatalytic application with high product yield and process stability. The setup was further automated in order to quickly identify the optimum operating conditions via a Design of Experiments (DoE) approach. The use of a systematic optimization, together with the adaptability of 3D printed equipment to the process requirements, render the presented approach highly promising for a more feasible implementation of biocatalysts in continuous industrial processes. Graphical abstract.Supplementary informationThe online version contains supplementary material available at 10.1007/s41981-021-00163-4.
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Project description:The morphological development and thermal properties of different polyamides with long-chain branches without forming a network structure were characterized by differential scanning calorimetry, polarized optical microscopy, and nonisothermal crystallization kinetics. The crystallization characteristics were analyzed using the nonisothermal kinetic equation proposed by Seo. Polarized optical microscopy and the Avrami exponent show the effect of the structural changes on the molecular ordering during the crystallization and early morphological development. The Avrami exponent, n, determined from the analysis of the nonisothermal crystallization kinetics, indicates a reduced heterogeneous nucleation for the modified polyamides. Structural changes (branching) of the polyamides impede crystallization, as indicated by the shift of the crystallization peaks to lower temperatures.
| S-EPMC7676331 | biostudies-literature
Project description:Embryonic salivary gland mesenchyme (eSGM) secretes various growth factors (bioactives) that support the proper growth and differentiation of salivary gland epithelium. Therefore, eSGM cells can be used as feeder cells for in vitro-cultured artificial salivary gland if their survival and bioactivity are properly maintained. As eSGM is encapsulated in a hyaluronan (HA)-rich developmental milieu, we hypothesized that mimicking this environment in vitro via surface immobilization of HA might enhance survival and bioactivity of eSGM. In this study, various HA derivatives, conjugated with catechol (HA-CA), thiol (HA-SH), or amine (HA-EDA) moieties, respectively, were screened for their efficacy of culturing eSGM-derived feeder cells in vitro. Among these HA derivatives, HA-CA showed the highest surface coating efficiency and growth enhancement effect on the embryonic submandibular gland. In addition, the HA-CA coating enhanced the production of growth factors EGF and FGF7, but not FGF10. These effects were maintained when eSGM cells isolated from the embryonic salivary gland were re-seeded to develop the feeder layer cells. CD44s (a major HA receptor) in eSGM cells were clustered at the cell membrane, and enhanced EGF expression was detected only in CD44 cluster-positive cells, suggesting that membrane clustering of CD44 is the key mechanism for the increased expression of EGF.
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