Lactose hydrolysis in packed-and fluidized-bed reactors using a recombinant β-galactosidase immobilized on magnetic core-shell capsules.

The objective of this study was to develop a bioprocess for lactose hydrolysis in diverse dairy matrices, specifically skim milk and cheese whey, utilizing column reactors employing a core-shell enzymatic system featuring β-galactosidase fused to a Cellulose Binding Domain (CBD) tag (β-galactosidase-CBD). The effectiveness of reactor configurations, including ball columns and toothed columns operating in packed and fluidized-bed modes, was evaluated for catalyzing lactose hydrolysis in both skim milk and cheese whey. In a closed system, these reactors achieved lactose hydrolysis rates of approximately 50% within 5 h under all evaluated conditions.

Magnetic core-shell cellulose system for the oriented immobilization of a recombinant β-galactosidase with a protein tag.

The objective of this study was to immobilize a recombinant β-galactosidase (Gal) tagged with a cellulose-binding domain (CBD) onto a magnetic core-shell (CS) cellulose system. After 30 min of reaction, 4 U/capsule were immobilized (CS@Gal), resulting in levels of yield and efficiency exceeding 80 %. The optimal temperature for β-galactosidase-CBD activity increased from 40 to 50 °C following oriented immobilization.

A systematic review about affinity tags for one-step purification and immobilization of recombinant proteins: integrated bioprocesses aiming both economic and environmental sustainability.

The present study reviewed and discussed the promising affinity tags for one-step purification and immobilization of recombinant proteins. The approach used to structure this systematic review was The Preferred Reporting Items for Systematic Review and Meta-analysis (PRISMA) methodology. The Scopus and Web of Science databases were used to perform the bibliographic survey by which 267 articles were selected.

Application of cellulosic materials as supports for single-step purification and immobilization of a recombinant β-galactosidase via cellulose-binding domain.

This study aimed to develop single-step purification and immobilization processes on cellulosic supports of β-galactosidase from Kluyveromyces sp. combined with the Cellulose-Binding Domain (CBD) tag. After 15 min of immobilization, with an enzymatic load of 150 U/g, expressed activity values reached 106.

One-step purification of a recombinant beta-galactosidase using magnetic cellulose as a support: Rapid immobilization and high thermal stability.

For the first time, this work reported the one-step purification and targeted immobilization process of a β-galactosidase (Gal) with the Cellulose Binding Domain (CBD) tag, by binding it to different magnetic cellulose supports. The process efficiency after β-galactosidase-CBD immobilization on magnetic cellulose-based supports showed values of approximately 90% for all evaluated enzymatic loads. Compared with free Gal, derivatives showed affinity values between β-galactosidase and the substrate 1.

Synthesis of magnetic nanoparticles functionalized with histidine and nickel to immobilize His-tagged enzymes using β-galactosidase as a model.

The aim of this study was to synthesize iron magnetic nanoparticles functionalized with histidine and nickel (FeO-His-Ni) to be used as support materials for oriented immobilization of His-tagged recombinant enzymes of high molecular weight, using β-galactosidase as a model. The texture, morphology, magnetism, thermal stability, pH and temperature reaction conditions, and the kinetic parameters of the biocatalyst obtained were assessed. In addition, the operational stability of the biocatalyst in the lactose hydrolysis of cheese whey and skim milk by batch processes was also assessed.

Production of beta-galactosidase fused to a cellulose-binding domain for application in sustainable industrial processes.

This study aimed to produce and characterize a recombinant Kluyveromyces sp. β-galactosidase fused to a cellulose-binding domain (CBD) for industrial application. In expression assays, the highest enzymatic activities occurred after 48 h induction on Escherichia coli C41(DE3) strain at 20 °C in Terrific Broth (TB) culture medium, using isopropyl β-d-1-thiogalactopyranoside (IPTG) 0.

Effect of by-products from the dairy industry as alternative inducers of recombinant β-galactosidase expression.

Objective: The aim of the present study was to evaluate the efficiency of lactose derived from cheese whey and cheese whey permeate as inducer of recombinant Kluyveromyces sp. β-galactosidase enzyme produced in Escherichia coli. Two E.

Magnetic cellulose: Versatile support for enzyme immobilization - A review.

Enzymes are proteins specialized in catalyzing biological reactions. However, factors such as cost and operational limitations could limit their applications in the industrial sector. An alternative to these limiting factors is enzyme immobilization, which enables reuse and increases biocatalyst stability.

Microbial β-Galactosidases of industrial importance: Computational studies on the effects of point mutations on the lactose hydrolysis reaction.

Hydrolysis efficiency of β-galactosidases is affected due to a strong inhibition by galactose, hampering the complete lactose hydrolysis. One alternative to reduce this inhibition is to perform mutations in the enzyme's active site. The aim of this study was to evaluate the effect of point mutations on the active site of different microbial β-galactosidases, using computational techniques.

Immobilization of β-Galactosidases on Magnetic Nanocellulose: Textural, Morphological, Magnetic, and Catalytic Properties.

We describe a process for obtaining nanocrystalline cellulose (NC) by either acidic (H-NC) or alkaline treatment (OH-NC) of microcrystalline cellulose, which was subsequently bonded to magnetic nanoparticles (H-NC-MNP and OH-NC-MNP) and used as support for the immobilization of Aspergillus oryzae (H-NC-MNP-Ao and OH-NC-MNP-Ao) and Kluyveromyces lactis (H-NC-MNP-Kl and OH-NC-MNP-Kl) β-galactosidases. The mean size of magnetic nanocellulose particles was approximately 75 nm. All derivatives reached saturation magnetizations of 7-18 emu/g, with a coercivity of approximately 4 kOe.

Modification of Immobead 150 support for protein immobilization: Effects on the properties of immobilized Aspergillus oryzae β-galactosidase.

We studied the modification of Immobead 150 support by either introducing aldehyde groups using glutaraldehyde (Immobead-Glu) or carboxyl groups through acid solution (Immobead-Ac) for enzyme immobilization by covalent attachment or ion exchange, respectively. These two types of immobilization were compared with the use of epoxy groups that are now provided on a commercial support. We used Aspergillus oryzae β-galactosidase (Gal) as a model protein, immobilizing it on unmodified (epoxy groups, Immobead-Epx) and modified supports.

Chelation by collagen in the immobilization of Aspergillus oryzae β-galactosidase: A potential biocatalyst to hydrolyze lactose by batch processes.

This work is the first study of the immobilization of Aspergillus oryzae β-galactosidase (Gal) on powdered collagen (Col) that had formed a chelate with aluminum (Col-Al-Gal). Other collagen treatments, including those with acetic acid, glutaraldehyde, and a combination of aluminum and glutaraldehyde (Col-Al-Glu-Gal), were also tested. High-yield (superior to 80%) and high-efficiency (superior to 99%) immobilization was obtained for the derivatives Col-Al-Gal and Col-Al-Glu-Gal, even at high protein loads (500-1,000 mg g of support).

Lactose Hydrolysis in Milk and Dairy Whey Using Microbial β-Galactosidases.

This work aimed at evaluating the influence of enzyme concentration, temperature, and reaction time in the lactose hydrolysis process in milk, cheese whey, and whey permeate, using two commercial β-galactosidases of microbial origins. We used Aspergillus oryzae (at temperatures of 10 and 55°C) and Kluyveromyces lactis (at temperatures of 10 and 37°C) β-galactosidases, both in 3, 6, and 9 U/mL concentrations. In the temperature of 10°C, the K.

Purification, immobilization, and characterization of a specific lipase from Staphylococcus warneri EX17 by enzyme fractionating via adsorption on different hydrophobic supports.

Staphylococcus warneri strain EX17 produces three lipases with different molecular weights of 28, 30, and 45 kDa. The 45 kDa fraction (SWL-45) has been purified from crude protein extracts by one chromatographic step based on the selective adsorption of this lipase by interfacial activation on different hydrophobic supports at low ionic strength. The adsorption of SWL-45 on octyl-Sepharose increased the enzyme activity by 60%, but the other lipases were also adsorbed on this support.

Optimization of lipase production by Staphylococcus warneri EX17 using the polydimethylsiloxanes artificial oxygen carriers.

In this research, the combined effects of polydimethylsiloxane (PDMS) and different conditions of oxygen volumetric mass transfer coefficient (k(L)a) on lipase production by Staphylococcus warneri EX17 were studied and optimized in bioreactor cultures. Raw glycerol from biodiesel synthesis was used as the sole carbon source. Full-factorial central composite design and the response surface methodology were employed for the experimental design and analysis of the results.

Single-step purification of different lipases from Staphylococcus warneri.

Three different lipases from the extract crude of Staphylococcus warneri have been purified by specific lipase-lipase interactions using different lipases (TLL, RML, PFL, BTL2) covalently attached to a solid support as adsorption matrix. BTL2 immobilized on glyoxyl-DTT adsorbed selectivity only a 30 kDa lipase from the crude, which was desorbed by adding 0.1% triton X-100.

Improved reactivation of immobilized-stabilized lipase from Thermomyces lanuginosus by its coating with highly hydrophilic polymers.

Immobilized-stabilized aminated lipase from Thermomyces lanuginosus (TLL-A) is not easily reactivated after inactivation by incubation in the presence of organic solvents or chaotropic reagents. To improve the recovered activity of this biocatalyst, immobilized TLL-A has been submitted to different modifications. The best results were obtained when the enzyme was coated with a very hydrophilic and inert polymer: dextran modified with glycine (Dx-Gly).

Improved enzyme stability in lipase-catalyzed synthesis of fatty acid ethyl ester from soybean oil.

In this work, we describe the optimization of the ethanolysis of soybean oil by the enzyme Lipozyme TL-IM in the lipase-catalyzed biodiesel synthesis and the improvement of the enzyme stability over repeated batches. The studied process variables were: reaction temperature, substrate molar ratio, enzyme content, and volume of added water. Fractional factorial design was used to analyze the variables so as to select those with higher influence on the reaction and then perform a central composite design to find the optimal reaction conditions.