Interaction of Ca2+ and Fe3+ in co-precipitation process induced by Virgibacillus dokdonensis and its application.

Biomineralization has garnered significant attention in the field of wastewater treatment due to its notable cost reduction compared to conventional methods. The reinjection water from oilfields containing an exceedingly high concentration of calcium and ferric ions will pose a major hazard in production. However, the utilization of biomineralization for precipitating these ions has been scarcely investigated due to limited tolerance among halophiles towards such extreme conditions. In this study, free and immobilized halophiles Virgibacillus dokdonensis were used to precipitate these ions and the effects were compared, at the same time, biomineralization mechanisms and mineral characteristics were further explored. The results show that bacterial concentration and carbonic anhydrase activity were higher when additionally adding ferric ion based on calcium ion; the content of protein, polysaccharides, deoxyribonucleic acid and humic substances in the extracellular polymers also increased compared to control. Calcium ions were biomineralized into calcite and vaterite with multiple morphology. Due to iron doping, the crystallinity and thermal stability of calcium carbonate decreased, the content of OC = O, NC = O and CO-PO3 increased, the stable carbon isotope values became much more negative, and ß-sheet in minerals disappeared. Higher calcium concentrations facilitated ferric ion precipitation, while ferric ions hindered calcium precipitation. The immobilized bacteria performed better in ferric ion removal, with a precipitation ratio exceeding 90%. Free bacteria performed better in calcium removal, and the precipitation ratio reached a maximum of 56%. This research maybe provides some reference for the co-removal of calcium and ferric ions from the oilfield wastewater.

Predicting the efficiency of arsenic immobilization in soils by biochar using machine learning.

Arsenic (As) pollution in soils is a pervasive environmental issue. Biochar immobilization offers a promising solution for addressing soil As contamination. The efficiency of biochar in immobilizing As in soils primarily hinges on the characteristics of both the soil and the biochar. However, the influence of a specific property on As immobilization varies among different studies, and the development and application of arsenic passivation materials based on biochar often rely on empirical knowledge. To enhance immobilization efficiency and reduce labor and time costs, a machine learning (ML) model was employed to predict As immobilization efficiency before biochar application. In this study, we collected a dataset comprising 182 data points on As immobilization efficiency from 17 publications to construct three ML models. The results demonstrated that the random forest (RF) model outperformed gradient boost regression tree and support vector regression models in predictive performance. Relative importance analysis and partial dependence plots based on the RF model were conducted to identify the most crucial factors influencing As immobilization. These findings highlighted the significant roles of biochar application time and biochar pH in As immobilization efficiency in soils. Furthermore, the study revealed that Fe-modified biochar exhibited a substantial improvement in As immobilization. These insights can facilitate targeted biochar property design and optimization of biochar application conditions to enhance As immobilization efficiency.

Simultaneous immobilization of multiple heavy metal(loid)s in contaminated water and alkaline soil inoculated Fe/Mn oxidizing bacterium.

Two strains of Fe/Mn oxidizing bacteria tolerant to high concentrations of multiple heavy metal(loid)s and efficient decontamination for them were screened. The surface of the bio-Fe/Mn oxides produced by the oxidation of Fe(II) and Mn(II) by Pseudomonas taiwanensis (marked as P4) and Pseudomonas plecoglossicida (marked as G1) contains rich reactive oxygen functional groups, which play critical roles in the removal efficiency and immobilization of heavy metal(loid)s in co-contamination system. The isolated strains P4 and G1 can grow well in the following environments: pH 5-9, NaCl 0-4%, and temperature 20-30°C. The removal efficiencies of Fe, Pb, As, Zn, Cd, Cu, and Mn are effective after inoculation of the strains P4 and G1 in the simulated water system (the initial concentrations of heavy metal(loid) were 1 mg/L), approximately reaching 96%, 92%, 85%, 67%, 70%, 54% and 15%, respectively. The exchangeable and carbonate bound As, Cd, Pb and Cu are more inclined to convert to the Fe-Mn oxide bound fractions in P4 and G1 treated soil, thereby reducing the phytoavailability and bioaccessible of heavy metal(loid)s. This research provides alternatives method to treat water and soil containing high concentrations of multi-heavy metal(loid)s.

Immobilization of laccase on magnetic PEGDA-CS inverse opal hydrogel for enhancement of bisphenol A degradation in aqueous solution.

Endocrine disruptors such as bisphenol A (BPA) adversely affect the environment and human health. Laccases are used for the efficient biodegradation of various persistent organic pollutants in an environmentally safe manner. However, the direct application of free laccases is generally hindered by short enzyme lifetimes, non-reusability, and the high cost of a single use. In this study, laccases were immobilized on a novel magnetic three-dimensional poly(ethylene glycol) diacrylate (PEGDA)-chitosan (CS) inverse opal hydrogel (LAC@MPEGDA@CS@IOH). The immobilized laccase showed significant improvement in the BPA degradation performance and superior storage stability compared with the free laccase. 91.1% of 100 mg/L BPA was removed by the LAC@MPEGDA@CS@IOH in 3 hr, whereas only 50.6% of BPA was removed by the same amount of the free laccase. Compared with the laccase, the outstanding BPA degradation efficiency of the LAC@MPEGDA@CS@IOH was maintained over a wider range of pH values and temperatures. Moreover, its relative activity of was maintained at 70.4% after 10 cycles, and the system performed well in actual water matrices. This efficient method for preparing immobilized laccases is simple and green, and it can be used to further develop ecofriendly biocatalysts to remove organic pollutants from wastewater.

Recombinant Esterase (BaCEm) Immobilized on Polyethyleneimine-Impregnated Mesoporous Silica SBA-15 Exhibits Outstanding Catalytic Performance.

A recombinant esterase, BaCEm, derived from Bacillus aryabhattai and heterologously expressed in Escherichia coli, was successfully immobilized on polyethyleneimine-impregnated mesoporous silica SBA-15. This immobilization utilized glutaraldehyde as a crosslinker. Optimal conditions were established with a PEI/SBA-15 ratio of 25% (w/w), a pH of 7.5, and a glutaraldehyde concentration of 0.5% (w/w), resulting in a loading capacity of 76.4 mg/g, a recovery activity of 43.5%, and a specific activity of 7917 U/g for BaCEm. The immobilized BaCEm demonstrated high enantioselectivity, with an "E" value of 203.92, in the resolution assay of (R,S)-ethyl indoline-2-carboxylate. Notably, the immobilized enzyme, compared to its free counterpart, exhibited enhanced thermostability, maintaining 95.4% of its activity after 3 h at 30 °C. It also showed significant tolerance to organic solvents, retaining 48.4% and 28.7% residual activity in 10% v/v acetonitrile and acetone, respectively. Moreover, its storage stability was confirmed, with 68.5% residual activity preserved after 30 days at 4 °C. Remarkably, the immobilized BaCEm retained 58.1% of its activity after 10 reuse cycles, underscoring the potential of polyethyleneimine-impregnated mesoporous silica SBA-15 as an effective support for enzyme immobilization, promising for industrial applications.

Designing tailor-made steric matters to improve the immobilized ficin specificity for small versus large proteins.

The development of strategies that can permit to adjust the size specificity of immobilized proteases by the generation of steric hindrances may enlarge its applicability. Using as a model ficin immobilized on glyoxyl agarose, two strategies were assayed to generate tailor made steric hindrances. First, ficin has been coimmobilized on supports coated with large proteins (hemoglobin or bovine serum albumin (BSA)). While coimmobilization of ficin with BSA presented no effect on the activity versus any of the assayed substrates, coimmobilization with hemoglobin permitted to improve the immobilized ficin specificity for casein versus hemoglobin, but still significant activity versus hemoglobin remained. Second, aldehyde-dextran has been employed to modify the immobilized ficin, trying to generate steric hindrances to avoid the entry of large proteins (hemoglobin) while enabling the entry of small ones (casein). This also increased the size specificity of ficin, but still did not suppress the activity versus hemoglobin. The combination of both strategies and the use of 37ºC during the proteolysis enabled to almost fully nullify the hydrolytic activity versus hemoglobin while preserving a high percentage of the activity versus casein. The modifications improved enzyme stability and the biocatalyst could be reused for 5 cycles without alteration of its properties.

High efficiency removal of antibiotic resistance gene with designer zinc-finger protein.

The use of agricultural biomass-based fertilizers, and the release of feces into the environment leads to last-lasting pollution of antibiotic resistance genes that cannot be removed from waters via traditional methods, resulting in significant health threats. To solve this issue, an antibiotic resistance gene removal method was proposed and tested that used sequence-specific DNA-binding designer zinc finger proteins, which target an 18-bp DNA sequence for specific antibiotic resistance gene binding and removal. Targeting the sulfonamide-resistant sul1 gene, sul1-binding zinc-finger protein was designed, overexpressed, and purified. This protein showed specific binding with sul1 over tetA that do not have the targeted sequence. This protein was further immobilized on agarose-based resins to prepare a sul1-removal column. When loaded with 10 mg protein, this column can remove over 99 % sul1 in water, suggesting high efficiency. This work presents a new method attempting to eliminate environmental and health threats posed by antibiotic resistance genes.

Efficient production of sophorose from glucose and its potentially industrial application in cellulase production.

Sophorose is the most effective inducer for cellulase production by Trichoderma reesei. Currently, the biosynthesis of sophorose is very inefficient, resulting in that unavailable for cellulase production in industry. In this study, CoGH1A, a multifunctional thermophilic glycoside hydrolase, was employed for sophorose production. Under the optimized conditions, the sophorose yield was 37.86 g/L with a productivity of 9.47 g/L/h which is by far the highest productivity. Meanwhile, the Fe3O4-CS-THP-CoGH1A nanoparticles were constructed to realize the recycling of CoGH1A. After 5 cycles of catalysis, Fe3O4-CS-THP-CoGH1A retained about 83.90 % enzyme activity. Finally, the mixtures of glucose and disaccharides (MGDC) obtained after being catalyzed by CoGH1A was used for cellulase production. As a result, the cellulase productivity achieved 188.38 FPU/L/h in 120 h. These results indicated that sophorose could be efficiently produced from glucose via transglycosylation by CoGH1A, making it possible to be industrially used as the inducer to improving the cellulase productivity.

Encapsulation of Candida antarctica lipase B in metal-organic framework under ultrasound and using it to one-pot synthesis of 1,3,4,5-tetrasubstituted pyrazoles.

Encapsulating the enzyme in metal-organic frameworks (MOFs) is a convenient method to prepare MOF-enzyme biocomposite. In this study, Candida antarctica lipase B (CAL-B) was chosen to immobilize in Cu-BTC MOF under ultrasound irradiation. CAL-B was immobilized in Cu-BTC under ultrasound at 21 kHz and 11.4 W/cm2 and incubation. 98% of CAL-B was immobilized in Cu-BTC with 99 U/mg activity (threefold more active than the free CAL-B). The prepared biocomposite was characterized using FT-IR, XRD, TGA, SEM, EDX, and BET. The thermal and solvent stability of CAL-B@Cu-BTC was investigated. It was found that at a temperature of 55 â„ƒ, CAL-B@Cu-BTC maintains its activity even after 2 h of incubation. Furthermore, in the presence of 20% and 50% concentrations of MeCN, THF, and DMF, CAL-B@Cu-BTC was found to have an activity of over 80%. A prepared biocatalyst was used to synthesize 1,3,4,5-tetrasubstituted pyrazole derivatives (50-75%) in a one-pot vessel, by adding phenyl hydrazine hydrochlorides, benzaldehydes, and dimethyl acetylenedicarboxylate.

Effect of Mn(II) photochemical oxidation on Cd immobilization in hematite.

Hematite, a commonly stable iron oxide in the environment, which can not only adsorb Cd in the environment, but also catalyze the photochemical oxidation of Mn(II) in the environment. However, the impact of Mn(II) on the structure of hematite and the adsorption of Cd during the surface oxidation of hematite remains unknown. In this study, we investigated the surface and structural changes of hematite after the photochemical oxidation of Mn(II), as well as the geochemical behavior of Cd during this process. The results demonstrate that Mn(II) was oxidized to Mn(III/IV) on the hematite surface, with some Mn(III) being incorporated into the hematite structure. Simulations using XRD data showed that higher Mn(II) concentrations resulted in increased levels of Mn doping, leading to significant variations in the hematite unit cell. This was further confirmed through FTIR and Raman spectroscopy characterization. The oxidation of Mn(II) on the hematite surface resulted in a shift in surface charge from positive to negative, enhancing the adsorption capacity of Cd. However, when Mn(II) exceeded 0.4 mM, the immobilization of Cd within the system decreased. This was attributed to the competitive adsorption of Mn(II) and a reduction in the relative abundance of Mn(IV) oxides.

Cleaning and Durability of Upper Extremity Orthotics: A Patient's Perspective.

Introduction We aimed to evaluate orthotic hygiene, preference for immobilization material, and frequency of unplanned orthotic adjustments and replacements. Methods All patients with fiberglass casts, thermoplastic splints, or prefabricated braces who presented at a large private academic institution between January 2020 and July 2023 were provided an 11-item survey assessing the length of immobilization, frequency of orthotic changes, orthotic hygiene, preference of immobilization, and whether patients recall instructions regarding orthotic care. Results A total of 385 surveys were collected, consisting of 96 (24.9%) casts, 202 (52.5%) thermoplastic splints, and 87 (22.6%) prefabricated braces. Patients were most frequently immobilized for two to six weeks. Of those, 106 (27.5%) patients required an unplanned adjustment or replacement. Almost half (182 patients, 47.3%) attempted to clean their orthotics, which was significantly greater among thermoplastic splints. A total of 229 (59.5%) respondents reported either not receiving or were unsure if they received instruction on proper orthotic hygiene. Conclusion Orthotic care and hygiene instructions are often overlooked or not retained by patients. Nearly one-third of patients required an unplanned adjustment or replacement, which was most frequent with thermoplastic orthotics.

Enhancing cauliflower growth under cadmium stress: synergistic effects of Cd-tolerant Klebsiella strains and jasmonic acid foliar application.

The pollution of heavy metals (HMs) is a major environmental concern for agricultural farming communities due to water scarcity, which forces farmers to use wastewater for irrigation purposes in Pakistan. Vegetables grown around the cities are irrigated with domestic and industrial wastewater from areas near mining, paint, and ceramic industries that pollute edible parts of crops with various HMs. Cadmium (Cd) is an extremely toxic metal in arable soil that enters the food chain and damages the native biota, ultimately causing a reduction in plant growth and development. However, the use of microbes and growth regulators enhances plant growth and development as well as HM immobilization into the cell wall and hinders their entry into the food chain. Thus, the integrated use of bacterial consortium along with exogenously applied jasmonic acid (JA) mitigates the adverse effect of metal stress, ultimately reducing the metal mobility into roots by soil. Therefore, the current study was conducted to check the impact of Cd-tolerant bacteria and JA on the growth, nutrient status, and uptake of Cd in the cauliflower (Brassica oleracea). Our results demonstrated that increasing concentrations of Cd negatively affect growth, physiological, and biochemical attributes, while the use of a bacterial consortium (SS7 + SS8) with JA (40 µmol L-1) significantly improved chlorophyll contents, stem fresh and dry biomass (19.7, 12.7, and 17.3%), root length and root fresh and dry weights (28.8, 15.2, and 23.0%), and curd fresh and dry weights and curd diameter (18.7, 12.6, and 15.1%). However, the maximum reduction in soil Cd, roots, and curd uptake was observed by 8, 11, and 9.3%, respectively, under integrated treatment as compared to the control. Moreover, integrating bacterial consortium and JA improves superoxide dismutase (SOD) (16.79%), peroxidase dismutase (POD) (26.96%), peroxidase (POX) (26.13%), and catalase (CAT) (26.86%). The plant nitrogen, phosphorus, and potassium contents were significantly increased in soil, roots, and curd up to 8, 11, and 9.3%, respectively. Hence, a consortium of Klebsiella strains in combination with JA is a potential phytostabilizer and it reduces the uptake of Cd from soil to roots to alleviate the adverse impact on cauliflower's growth and productivity.

Treatment Reproducibility in Brain Stereotactic Radiotherapy Using a Shim Mask Versus a Standard Mask.

Introduction This study aimed to evaluate the setup accuracy of the new shim mask with mouth bite compared to the standard full brain mask in stereotactic radiosurgery (SRS) and radiotherapy (SRT) treatments for brain metastases or tumors. Method A combined retrospective and prospective design was employed, involving 40 patients treated at our center. Patients previously treated using standard head masks formed the retrospective cohort, while those treated with the Shim mask and mouth bite formed the prospective cohort. Daily cone-beam computed tomography (CBCT) scans were obtained before each treatment session to ensure patient setup accuracy. Key metrics included absolute shifts in translational and rotational directions, the number of repeat CBCTs, and the time interval between CBCTs. Results The Shim mask significantly reduced the mean setup errors in the lateral translation (p=0.022) from 0.17 cm (SD=0.10) to 0.10 cm (SD=0.10), and in X-axis rotation (p=0.030) from 0.79° (SD=0.43) to 0.47° (SD=0.47). By considering cutoff points of 1 mm in translational and 1° in rotational directions, the Shim mask was significantly more accurate in the lateral direction (p=0.004). Moreover, while 70% of patients in the standard group required repeat CBCT scans, none in the Shim group did, resulting in an average time saving of 10.4 minutes per patient. Conclusion The Shim mask with mouth bite offers enhanced immobilization accuracy in SRT/SRS treatments, leading to time and potential cost savings by reducing the need for repeat CBCT scans. This underscores the importance of adopting innovative immobilization techniques to optimize patient outcomes.

Immobilization of recombinant serine protease from Virgobacillus natechei FarDT on amino graphene-chitosan biocompatible nanohybrid for enhancing pH and thermal stability.

The serine protease gene was heterologously expressed in Escherichia coli BL21 (DE3) using the PET 28a vector. The purified enzyme was immobilized on a nanohybrid of amino graphene and chitosan. The characterization of synthesized nanohybrids and immobilized enzymes was confirmed by Fourier transform infrared (FTIR), X-ray diffraction (XRD), dynamic light scattering (DLS), and field emission scanning electron microscopy (FE-SEM). Immobilization increased the temperature optimum from 60 to 70 °C for both free and immobilized enzymes, while the optimal pH of the enzymes did not change post-immobilization (pH 8). The immobilized biocatalyst significantly enhanced thermal stability, as well as enzyme stability at significant pH ranges. After 30 days of storage, the immobilized enzymes exhibited approximately 83 % of their relative activity, while the free protease retained only 56 % of its initial activity. Stabilization also altered the kinetic parameters (increasing Km, decreasing Kcat/Km, and Vmax) and thermodynamic parameters (increasing enzyme half-life and activation energy). The study's outcomes represent a significant advancement in the realm of enzyme synthesis and its stabilization using several combined technologies, including enzyme production with recombinant DNA technology based on gene synthesis, and its stabilization using a hybrid substrate synthesized from nanomaterials. Based on these findings, the immobilized recombinant enzyme has high potential for industrial use as an efficient and stable biocatalyst.

Hyperactivation of crosslinked lipases in elastic hydroxyapatite microgel and their properties.

Effective enzyme stabilization through immobilization is essential for the functional usage of enzymatic reactions. We propose a new method for synthesizing elastic hydroxyapatite microgel (E-HAp-M) materials and immobilizing lipase using this mesoporous mineral via the ship-in-a-bottle-neck strategy. The physicochemical parameters of E-HAp-M were thoroughly studied, revealing that E-HAp-M provides efficient space for enzyme immobilization. As a model enzyme, lipase (LP) was entrapped and then cross-linked enzyme structure, preventing leaching from mesopores, resulting in highly active and stable LP/E-HAp-M composites. By comparing LP activity under different temperature and pH conditions, it was observed that the cross-linked LP exhibited improved thermal stability and pH resistance compared to the free enzyme. In addition, they demonstrated a 156% increase in catalytic activity compared with free LP in hydrolysis reactions at room temperature. The immobilized LP maintained 45% of its initial activity after 10 cycles of recycling and remained stable for over 160 days. This report presents the first demonstration of a stabilized cross-linked LP in E-HAp-M, suggesting its potential application in enzyme-catalyzed processes within biocatalysis technology.

Unlocking soil revival: the role of sulfate-reducing bacteria in mitigating heavy metal contamination.

Soil contamination with heavy metals from industrial and mining activities poses significant environmental and public health risks, necessitating effective remediation strategies. This review examines the utilization of sulfate-reducing bacteria (SRB) for bioremediation of heavy metal-contaminated soils. Specifically, it focuses on SRB metabolic pathways for heavy metal immobilization, interactions with other microorganisms, and integration with complementary remediation techniques such as soil amendments and phytoremediation. We explore the mechanisms of SRB action, their synergistic relationships within soil ecosystems, and the effectiveness of combined remediation approaches. Our findings indicate that SRB can effectively immobilize heavy metals by converting sulfate to sulfide, forming stable metal sulfides, thereby reducing the bioavailability and toxicity of heavy metals. Nevertheless, challenges persist, including the need to optimize environmental conditions for SRB activity, address their sensitivity to acidic conditions and high heavy metal concentrations, and mitigate the risk of secondary pollution from excessive carbon sources. This study underscores the necessity for innovative and sustainable SRB-based bioremediation strategies that integrate multiple techniques to address the complex issue of heavy metal soil contamination. Such advancements are crucial for promoting green mining practices and environmental restoration.

In-situ stabilized lipase in calcium carbonate microparticles for activation in solvent-free transesterification for biodiesel production.

Biodiesel serves as a crucial biofuel alternative to petroleum-based diesel fuels, achieved through enzymatic transesterification of oil substrates. This study aims to investigate stabilized lipase (LP) within calcium carbonate (CaCO3) microparticles as a catalyst for solvent-free transesterification in biodiesel synthesis. The specific hydrolysis activity of the in-situ immobilized LP was 66% of that of free LP. However, the specific transesterification activity of immobilized LP in the solvent-free phase for biodiesel production was 2.29 times higher than that of free LP. These results suggest that the interfacial activation of LP molecules is facilitated by the inorganic CaCO3 environment. The immobilized LP demonstrated higher biodiesel production levels with superior stability compared to free LP, particularly regarding methanol molar ratio and temperature. To the best of our knowledge, there are no previous reports on the in-situ immobilization of LP in a CaCO3 carrier without any crosslinker as an interfacial-activated biocatalyst for biodiesel production.

Protein Immobilization on Bacterial Cellulose for Biomedical Application.

New carriers for protein immobilization are objects of interest in various fields of biomedicine. Immobilization is a technique used to stabilize and provide physical support for biological micro- and macromolecules and whole cells. Special efforts have been made to develop new materials for protein immobilization that are non-toxic to both the body and the environment, inexpensive, readily available, and easy to modify. Currently, biodegradable and non-toxic polymers, including cellulose, are widely used for protein immobilization. Bacterial cellulose (BC) is a natural polymer with excellent biocompatibility, purity, high porosity, high water uptake capacity, non-immunogenicity, and ease of production and modification. BC is composed of glucose units and does not contain lignin or hemicellulose, which is an advantage allowing the avoidance of the chemical purification step before use. Recently, BC-protein composites have been developed as wound dressings, tissue engineering scaffolds, three-dimensional (3D) cell culture systems, drug delivery systems, and enzyme immobilization matrices. Proteins or peptides are often added to polymeric scaffolds to improve their biocompatibility and biological, physical-chemical, and mechanical properties. To broaden BC applications, various ex situ and in situ modifications of native BC are used to improve its properties for a specific application. In vivo studies showed that several BC-protein composites exhibited excellent biocompatibility, demonstrated prolonged treatment time, and increased the survival of animals. Today, there are several patents and commercial BC-based composites for wounds and vascular grafts. Therefore, further research on BC-protein composites has great prospects. This review focuses on the major advances in protein immobilization on BC for biomedical applications.

Immobilization of Lipase from Thermomyces Lanuginosus and Its Glycerolysis Ability in Diacylglycerol Preparation.

In the glycerolysis process for diacylglycerol (DAG) preparation, free lipases suffer from poor stability and the inability to be reused. To address this, a cost-effective immobilized lipase preparation was developed by cross-linking macroporous resin with poly (ethylene glycol) diglycidyl ether (PEGDGE) followed by lipase adsorption. The selected immobilization conditions were identified as pH 7.0, 35 °C, cross-linking agent concentration 2.0%, cross-linking time 4 h, lipase amount 5 mg/g of support, and adsorption time 4 h. Enzymatic properties of the immobilized lipase were analyzed, revealing enhanced pH stability, thermal stability, storage stability, and operational stability post-immobilization. The conditions for immobilized enzyme-catalyzed glycerolysis to produce DAG were selected, demonstrating the broad applicability of the immobilized lipase. The immobilized lipase catalyzed glycerolysis reactions using various oils as substrates, with DAG content in the products ranging between 35 and 45%, demonstrating broad applicability. Additionally, the changes during the repeated use of the immobilized lipase were characterized, showing that mechanical damage, lipase leakage, and alterations in the secondary structure of the lipase protein contributed to the decline in catalytic activity over time. These findings provide valuable insights for the industrial application of lipase.

Size-dependent ability of AtaA to immobilize cells in Acinetobacter sp. Tol 5.

Microbial cells serve as efficient and environmentally friendly biocatalysts, but their stability and reusability in practical applications must often be improved through immobilization. Acinetobacter sp. Tol 5 shows high adhesiveness to materials due to its large cell surface protein AtaA, which consists of 3630 amino acids (aa). Previously, we developed a method for immobilizing bacteria using AtaA. Herein, we investigated the cell immobilization ability of in-frame deletion (IFD) mutants of AtaA with different sizes in Tol 5. Mini-AtaA, which consists of 775 aa and is functional in Escherichia coli, was produced and present on the cell surface; however, mini-AtaA showed no immobilization ability in Tol 5. A cell immobilization assay was performed with cells expressing 16 IFD mutants of AtaA with different sizes, revealing that a length of at least 1417 aa was required for the sufficient immobilization of Tol 5 cells; thus, the minimum length needed to achieve the adhesive function of AtaA varies among bacterial species. The constructed mutant library of AtaA ranging from 3630 to 775 aa will allow researchers to quickly and easily explore the optimal size of AtaA, even for bacteria newly introduced to AtaA.