Support For Enzyme Immobilization Research Articles

Catalytic Activity of Immobilized Chymotrypsin on Hybrid Silica-Magnetic Biocompatible Particles and Its Application in Peptide Synthesis.

In this work, novel silica hybrid magnetic particles with biocompatible surface were designed as a support for enzyme immobilization, and the immobilized chymotrypsin (CT) performance was clarified as a model biocatalyst. CT is used in food technology for drink clarification and protein hydrolysis. The enzyme was directly immobilized onto polydopamine-grafted magnetic silica particles (MNP@SiO2@PDA-CT) via the Schiff base reaction. Immobilized enzyme had broadened for both pH and temperature profiles compared with the native CT. The MNP@SiO2@PDA-CT system also improved in thermostability compared with the native enzyme. The immobilized CT was operated in a continuous enzyme reactor (CER) for the hydrolysis of different proteins (i.e., cytochrome c (Cyt c), human serum albumin (HSA), human immunoglobulin G (HIgG), and lysozyme (Lys)). The peptide synthesis rate was shown to decrease as a function of increasing flow rate. The catalytic activity of the CER remained stable for 6.0 h in a continuous operation period; thus, the presented method may increase applicability in the food technology area. The immobilized CT in the CER showed a good hydrolysis performance for all the tested model proteins. The peptides hydrolyzed from the tested proteins were analyzed with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-ToF-MS). Results show that the MNP@SiO2@PDA-CT system also permits for the applicability in the area of proteomic research.

Facile preparation of aminosilane-alginate hybrid beads for enzyme immobilization: Kinetics and equilibrium studies

A simple, facile and potential platform for enzyme immobilization using alginate-based beads has been demonstrated by simultaneous gelation and modification of alginate using calcium chloride (CaCl2) and 3-aminopropyltriethoxysilane (APTES). In this method, sodium alginate solution containing enzyme was simply dripped into a crosslinker solution containing CaCl2 and APTES, leading to the formation of APTES-alginate hybrid beads (AP-beads). The optical observation, FT-IR analysis and amino group measurements provided evidence that APTES was successfully adsorbed to the alginate chain via electrostatic interaction. On the assumption that the binding of Ca2+ ion to polymannuronate residues of alginate via bidentate bridging coordination is competitive with APTES, the equilibrium isotherm and kinetics for the adsorption of APTES to AP-beads was found to follow extended Langmuir isotherm in binary system. Formate dehydrogenase (FDH) as a model enzyme was successfully immobilized in AP-beads and the immobilization yield of ca. 100% could be achieved under optimal conditions of CaCl2 and APTES concentrations in crosslinker solution. Furthermore, the AP-beads were reused efficiently for 9 cycles without loss of FDH activity. The above results demonstrated that AP-beads were effective support for enzyme immobilization.

Peptide-Mediated Immobilization on Magnetoferritin for Enzyme Recycling.

Ferritin possess favorable properties because its exterior and interior surface can be applied to generate functional nanomaterials, which make them possible for enzyme immobilization and recycling. Here, we report the noncovalent immobilization of a genetically modified β-glucosidase onto the outer surface of synthetic magnetoferritin through the electrostatic interaction of a heterodimeric coiled-coil protein formed by coils containing lysine residues (K-coils) and coils containing glutamic acid (E-coils). The immobilized enzyme was characterized, and its enzymatic properties were evaluated. Furthermore, reusability of immobilized enzyme was demonstrated in aqueous solution under an applied magnetic field. The results showed that magnetoferritin was successfully prepared and it was an excellent support for enzyme immobilization. After three times usages, the retention rates were 93.75%, 82.5%, and 56.25%, respectively, demonstrating that immobilized enzyme possessed good retention efficiency and could be used as potential carrier for other biomolecules. The strategy of enzyme immobilization developed in this work can be applied, in general, to many other target molecules.

High-performance enzymatic biofuel cell based on three-dimensional graphene

Enzymatic biofuel cells are a subclass of biofuel cells, which employ enzymes to generate energy from renewable sources. In this study, 3-dimensional graphene (3DG)/glucose oxidase (GOx) bio-nanocomposite was fabricated in order to improve enzyme immobilisation and enzyme lifetime with an enhanced electron transfer rate. These enhancements are due to the unique physical properties of 3DG, e.g. high porosity, large surface area, and excellent electrical conductivity. A power density of 164 μW cm−2 at 0.4 V was achieved from this enzymatic biofuel cell (EBFC) with an acceptable performance compared to that of the other glucose biofuel cells (GBFCs). The 3DG enhances the enzyme lifetime, decreases enzyme leaking and, due to its good conductivity, facilitates the electron harvest and transfer from the enzyme active site to the electrode. This suggests that 3DG could be used as effective support for enzyme immobilisation on the surface of the electrode in EBFC applications and related areas such as biosensors, bioreactors and implantable biofuel cells.

Carbon-based electrode loaded with Y-doped SrTiO3 perovskite as support for enzyme immobilization in biosensors

Many studies have been presented Y-doped SrTiO3 as a potential alternative material for electrical, electrochemical and chemical applications due to its mixed (ionic and electronic) conduction and its morphological characteristics. In this work, 8% mol yttrium doped SrTiO3 (YST08) was used to assemble a Horseradish peroxidase (HRP) biosensor aiming at hydrogen peroxide (H2O2) detection. The perovskite was characterized by X-ray diffraction (XRD), Brunauer–Emmett–Teller N2 physisorption (BET), scanning electron microscopy (SEM) and energy-dispersive X-ray spectroscopy (EDS); and, after HRP immobilization, by zeta potential, atomic force microscopy (AFM), cyclic voltammetry, Fourier-transform infrared spectroscopy (FTIR), and chronoamperometry. The structure presented a low specific surface area due to particle agglomeration as a result of the elevated temperature of calcination. This characteristic was ratified by SEM analysis, which also demonstrated regular and well-rounded grains. Zeta potential results suggest successful HRP immobilization on YST08 surface because of the resemblance of zeta potential of immobilized sample and free HRP. FTIR spectra and AFM micrographs confirm a homogenous HRP adsorption due to functional groups identification and surface topography changes. YST08 + HRP electrode proved to be efficient in sensing H2O2 in different scan rates and background electrolytes under electrochemical evaluation. Results also showed that the process is surface-controlled, exhibiting a low reduction potential for H2O2 around −0.31 V at pH = 7. The YST08 matrix was effective to the diffusion mechanisms presenting a detection limit of 14.97 μM; and increases the enzyme long-term activity, retaining 14.97% of specific capacity after 180 days.

Urease adsorption immobilization on ionic liquid-like macroporous polymeric support

In this work, we report the synthesis and application of polymeric macroporous materials functionalized with ionic liquid (IL)-like moieties to serve as a support for enzyme immobilization. The method was based on bottom-up approach, where poly(styrene–divinylbenzene) (PS–DVB) nanoparticles were used as building blocks to form porous structures. Surface functionalization was done by introducing 1-butyl imidazole into the PS–DVB support to form IL-like imidazolium, which was consequently used for enzyme adsorption immobilization on the porous surface. To demonstrate activity of immobilized enzyme, hydrolysis of urea catalyzed by Jack bean urease was used as a model reaction. The enzymatic activity of the urease to convert urea solution into carbon dioxide (hydrocarbonates in water solution) and ammonia under acidic conditions were monitored both by measuring changes in pH and by a color change in cresol red pH indicator. The immobilized urease was able to provide hydrolysis of urea solution for 30 days while maintaining its activity over 100% compared to free enzyme solution. The activity of freshly immobilized enzyme was increased up to 285% compared to free urease solution under acidic conditions, which is caused by an acidic shift of activity–pH bell-shaped curve. Prepared porous material with immobilized urease was able to undergo four consequent cycles over the period of 4 days with only 15% decrease in activity. Overall, the results indicated that the polymeric support is well suitable to combine the advantages of macroporous material with IL-like surface moieties for enzyme immobilization and its consequent application in bio-catalytic reactions.

Enhanced catalytic performance of lipase covalently bonded on ionic liquids modified magnetic alginate composites

We prepared ionic liquids (ILs) modified magnetic alginate nanoparticles and used these as supports for lipase immobilization. The novel supports were characterized using Fourier transform infrared (FTIR) spectroscopy, nuclear magnetic resonance (1HNMR), vibrating sample magnetometry (VSM), thermogravimetry (TG), transmission electron microscopy (TEM) and water contact angle (WCA) measurements. The immobilized lipase (PPL-IL-MSA) exhibited high activity, 2.1-fold higher than that compared to free lipase and 1.59-fold higher compared to immobilized lipase without IL (PPL-MSA). In addition, the pH and temperature application range of PPL-IL-MSA were both found to be broader than that of free lipase and PPL-MSA. The thermal stability, denaturation stability, and reusing stability of PPL-IL-MSA were also higher than those of other samples. After 10 times of reuse, the residual activity of PPL-IL-MSA was 89.7% higher than that of PPL-MSA (84.4%). Furthermore, the kinetic constant Km of PPL-IL-MSA was 13.7 mg/mL lower than that of free lipase (21.2 mg/mL) and PPL-MSA (18.4 mg/mL). Circular dichroism (CD) was used to study the secondary structure of enzymes in order to explain the mechanism of the performance improvement of PPL-IL-MSA. This work involving the development of a new supports for enzyme immobilization may serve as a reference for further studies in this field.

Influence of reaction parameters in the polymerization between genipin and chitosan for enzyme immobilization

Abstract In this work, we studied the effect of some variables in the reaction between genipin and chitosan, to prepare a support for enzyme immobilization. Reaction temperature, pH, time and genipin concentration were evaluated for support activation and β-galactosidase from Aspergillus oryzae was used as the enzyme model. The immobilized enzyme was characterized by immobilization properties and kinetic parameters and by thermal and operational stability. It was found that support activation at alkaline pH (9.0), at 60 °C for 1 h, resulted in an activity recovery of 37.5%. The immobilized enzyme had a stabilization factor of 2.0 at 50 °C compared with the free enzyme. In batch process, the immobilized preparation kept 100% of its relative initial activity during 40 batches. This research shows, for the first time, a complete study about the parameters that affect the support activation process and the possible structure formed between genipin and chitosan.

Spray-dried immobilized lipase from Geobacillus sp. strain ARM in sago.

Sago starch is traditionally used as food especially in Southeast Asia. Generally, sago is safe for consumption, biodegradable, easily available and inexpensive. Therefore, this research was done to expand the potential of sago by using it as a support for enzyme immobilization. In this study, ARM lipase, which was isolated from Geobacillus sp. strain ARM, was overexpressed in Escherichia coli system and then purified using affinity chromatography. The specific activity of the pure enzyme was 650 U/mg, increased 7 folds from the cell lysate. The purified enzyme was immobilized in gelatinized sago and spray-dried by entrapment technique in order to enhance the enzyme operational stability for handling at high temperature and also for storage. The morphology of the gelatinized sago and immobilized enzyme was studied by scanning electron microscopy. The results showed that the spray-dried gelatinized sago was shrunken and became irregular in structure as compared to untreated sago powder. The surface areas and porosities of spray-dried gelatinized sago with and without the enzyme were analyzed using BET and BJH method and have shown an increase in surface area and decrease in pore size. The immobilized ARM lipase showed good performance at 60–80 °C, with a half-life of 4 h and in a pH range 6–9. The immobilized enzyme could be stored at 10 °C with the half-life for 9 months. Collectively, the spray-dried immobilized lipase shows promising capability for industrial uses, especially in food processing.

Proteolytic ceramic capillary membranes for the production of peptides under flow

In this study, we investigate the effect of membrane surface functionalization on the immobilization of the protease subtilisin A and its performance in the production of peptides from the model protein casein under flow. The surface of tubular ceramic membranes was silanized to yield carboxylated and aminated supports for enzyme immobilization via non-covalent and carbodiimide activated binding. The protease density correlated with electrostatic interactions between the positively charged enzyme and the supports, with the highest enzyme density reached on negatively charged, carboxylated membranes (0.019 molecules/nm², noncovalent approach). Enzyme leaching was reduced by covalent binding of protease to carboxylated supports (5% leached) and slightly improved by binding to aminated membranes (46%) over non-covalent binding to unfunctionalized reference capillaries (66%). Regarding carbodiimide activated immobilization, protease on unfunctionalized and aminated supports exhibited a significantly larger specific activity (0.99 μmol/min/mg) than enzymes on carboxylated surfaces (0.15 μmol/min/mg), which suggests preferred enzyme orientation. In protein hydrolysis, these differences in surface-enzyme interactions were reflected by variations in peptide composition and degree of hydrolysis. Accordingly, we demonstrate that surface functionalization critically determines the surface properties of protease support materials for the production of peptides under flow and allows tailoring the performance of proteolytic capillary membranes.

Organic-inorganic nanocomposites fabricated via functional ionic liquid as the bridging agent for Laccase immobilization and its application in 2,4-dichlorophenol removal

In this study, organic-inorganic nanocomposite including mesoporous silica SBA-15 and chitosan was combined using carboxyl-functionalized ionic liquid as the bridging agent (SBA-CIL-CS), and which was characterized by scanning electron microscope, Fourier transform infrared spectroscopy, thermogravimetric analysis, and nitrogen adsorption-desorption isotherms. Thus prepared nanocomposite was applied to laccase immobilization via physical adsorption (SBA-CIL-CS-Lac) and utilized in 2,4-dichlorophenol (2,4-DCP) removal for the first time. SBA-CIL-CS-Lac showed relatively high immobilization efficiency, activity retention (75.3%), pH stability and storage stability. Kinetic experiment showed SBA-CIL-CS-Lac had the eminent affinity to the substrate. Circular dichroism and fluorescence analysis verified the active conformation of SBA-CIL-CS-Lac was maintained well to keep its enzymatic activity. Compared with the previous methods, SBA-CIL-CS-Lac exhibited prominent catalytic efficiency and reusability as well as pH and temperature stability for 2,4-DCP removal. The removal rate of SBA-CIL-CS-Lac for 2,4-DCP could be up to ˜90% after 35 h. Moreover, 58.8% removal rate could be retained after five operation runs. These results indicated that organic-inorganic nanocomposite SBA-CIL-CS was an ideal support for enzyme immobilization.

Hollow silica microspheres as robust immobilization carriers

Hollow silica microspheres provide an ideal solid support for enzyme immobilization. We tested one of the newest development, namely MATSPHERES®, a silica openwork material as a carrier for the covalent immobilization of enzymes used to synthesize bioactive compounds. Two model enzymes – ethylbenzene dehydrogenase and EL070 lipase – were considered. They belong to two different enzyme classes and catalyse reactions taking place in various environments (aqueous and non-aqueous, aerobic and anaerobic). The enzymes were immobilized by covalent bonds (via divinyl sulfone and glutaraldehyde) on new silica material. Effectiveness of immobilization processes on the spheres grafted with amine groups and on the analogues without functionalization was determined for both enzymes. Microspheres were characterized morphologically and also their mechanical stability was examined during exposure to varying physical conditions. It was shown that MATSPHERES® due to their openwork structure and relative stability under batch and flow conditions can be a competitive SBA support for enzyme immobilization and production of bioactive compounds.

Influence of silica architecture on the catalytic activity of immobilized glucose oxidase

Immobilizing enzymes on solid support materials is of great interest for many applications as it improves enzyme lifetime and enables its recycling. Synthetic mesoporous silica materials are widely used as supports for enzyme immobilization due to their high enzyme-loading capacity and their excellent mass transport properties through interconnected channels of mesopores. Much less is known about the enzyme support properties of naturally occurring mesoporous silica materials. These are readily available through the growth of diatoms, which biosynthesize hierarchically porous silica microshells with pore diameters ranging from 10 to 1000 nm. Here the authors investigated the performance as enzyme support materials of biosilica from three diatom species in comparison to synthetic silica materials. Using glucose oxidase (GOx) as a model enzyme, the authors determined the enzyme-loading capacities and the specific catalytic activities of the GOx-bearing (bio)silicas both under suspension conditions and in a flow-through setting. Remarkably, the specific activity of GOx was strongly dependent on the type of diatom biosilica material and in the best case was as high as on synthetic mesoporous silica. Porosity analysis of the materials points to an important role of large mesopores (diameters >20 nm) for supporting a high specific activity of (bio)silica-immobilized GOx.

Chitosan activated with divinyl sulfone: a new heterofunctional support for enzyme immobilization. Application in the immobilization of lipase B from Candida antarctica.

A novel heterofunctional support for enzyme immobilization, chitosan-divinyl sulfone, was assessed in this study. The activation of chitosan with DVS was carried out at three different pHs (10.0, 12.5 and 14.0) and a Candida antarctica Lipase B (CALB) was selected as the model enzyme. After immobilization, the biocatalysts were incubated under alkaline conditions in a buffer to facilitate the multipoint covalent attachment, followed by incubation in ethylenediamine (EDA) aiming at blocking the remaining reactive groups. The highest thermal stability was obtained when pH 10.0 was used during support activation. These results were shown to be better than those obtained when using glutaraldehyde as the support-activating reagent. Subsequently, the immobilization pH was investigated (5.0, 7.0 and 10.0) prior to alkaline incubation, with the highest enzyme stability levels found at pH 10.0. Finally, the selected biocatalyst was used in the hydrolysis of ethyl hexanoate and presented an activity of 14,520.37 U/g of immobilized lipase at pH 5.0. These results show that chitosan activated with divinyl sulfone is a very promising support for enzyme immobilization and the proposed protocol is able to successfully improve enzyme stability.

Immobilized Nanoparticles-Mediated Enzymatic Hydrolysis of Cellulose for Clean Sugar Production: A Novel Approach

Background:Enzymatic hydrolysis of cellulose is an expensive approach due to the high cost of an enzyme involved in the process. The goal of the current study was to apply magnetic nanomaterials as a support for immobilization of enzyme, which helps in the repeated use of immobilized enzyme for hydrolysis to make the process cost-effective. In addition, it will also provide stability to enzyme and increase its catalytic activity.Objective:The main aim of the present study is to immobilize cellulase enzyme on Magnetic Nanoparticles (MNPs) in order to enable the enzyme to be re-used for clean sugar production from cellulose.Methods:MNPs were synthesized using chemical precipitation methods and characterized by different techniques. Further, cellulase enzyme was immobilized on MNPs and efficacy of free and immobilized cellulase for hydrolysis of cellulose was evaluated.Results:Enzymatic hydrolysis of cellulose by immobilized enzyme showed enhanced catalytic activity after 48 hours compared to free enzyme. In first cycle of hydrolysis, immobilized enzyme hydrolyzed the cellulose and produced 19.5 ± 0.15 gm/L of glucose after 48 hours. On the contrary, free enzyme produced only 13.7 ± 0.25 gm/L of glucose in 48 hours. Immobilized enzyme maintained its stability and produced 6.15 ± 0.15 and 3.03 ± 0.25 gm/L of glucose in second and third cycle, respectively after 48 hours.Conclusion:This study will be very useful for sugar production because of enzyme binding efficiency and admirable reusability of immobilized enzyme, which leads to the significant increase in production of sugar from cellulosic materials.

Laccase-immobilized tannic acid-mediated surface modification of halloysite nanotubes for efficient bisphenol-A degradation.

Halloysite nanotubes (HNTs) have been pursued as promising carriers for enzyme immobilization, but the lack of functional groups severely limits their applications. Herein, we reported a simple tannic acid (TA)-mediated surface modification strategy for the fabrication of HNT-based efficient enzyme immobilization supports. Particularly, TA was first self-polymerized and deposited onto the surface of HNTs to form a thin active film via a mussel-inspired method, and the model enzyme laccase was directly conjugated via the Michael addition and/or Schiff base condensation between quinone groups on poly(tannic acid) layer surfaces and exposed amine groups on laccase surfaces. Under the optimum conditions, this newly fabricated support retained good enzyme-loading and activity recovery properties with 197.9 mg protein per gram of support and 55.4% of activity recovery being achieved. In addition, this immobilized laccase was less influenced by pH, temperature, and inhibitor changes and exhibited higher storage stability than free laccases as more than 70% of initial activity was retained by the immobilized laccase, while less than 30% was retained for free laccase after one-month storage at 4 °C. Finally, a higher bisphenol-A (BPA) removal efficiency and more reuse cycles were demonstrated for immobilized laccases. As a result, this TA-mediated surface modification is a simple and green method for biological macromolecule immobilization on HNTs in one step.

Design and preparation of stable CPO/HRP@H‐MOF(Zr) composites for efficient bio‐catalytic degradation of organic toxicants in wastewater

ABSTRACTBACKGROUNDImmobilization of enzyme on a solid support can improve its stability, however, this is often accompanied by a decrease of the initial catalytic activity. The design and preparation of immobilized enzyme with simultaneous high activity and stability is a challenge.RESULTSA hierarchically porous metal organic framework H‐MOF(Zr) which was prepared by ‘crystal defect’ strategy was very stable in aqueous solution and possessed well‐defined, tunable and adjacent hierarchical pores. When using the H‐MOF(Zr) as support for enzyme immobilization, a single array of enzyme in a single mesoporous channel was achieved so as to avoid aggregation of the enzyme. Meanwhile the micropores can be utilized to concentrate substrates, resulting in a decrease in the mass transfer resistance of substrate compared with that in the bulk buffer due to the enzymes and substrates closely embedded on the same support. The prepared immobilized enzyme CPO/HRP@H‐MOF(Zr) was more stable at elevated temparatures, showing 58.2% enhanced activity compared with free enzymes at 70 °C for 1 h incubation. After 12 cycles, 70.7% of activity remained. When the CPO/HRP@H‐MOF(Zr) was applied in the treatment of wastewater containing isoproturon and 2,4‐dichlorophenol, complete degradation was achieved in only 15 min.CONCLUSIONBoth high activity and stability under harsh reaction conditions give the CPO/HRP@H‐MOF(Zr) composites potential for practical application in wastewater treatment. © 2018 Society of Chemical Industry

Covalently immobilized laccase onto graphene oxide nanosheets: Preparation, characterization, and biodegradation of azo dyes in colored wastewater

In this study, graphene oxide (GO) was synthesized via modified Hummer's method and exploited as an ideal enzyme immobilization support due to its exclusive chemical and structural features. Then, laccase from genetically modified Aspergillus was covalently immobilized onto GO (nanobiocatalyst). Enzymatic characterization of the nanobiocatalyst exhibited promising results: laccase loading of 156.5 mg g−1 and immobilization yield of 64.6% at laccase concentration of 0.9 mg/ mL. Further employment of various structural characterization techniques including Fourier Transform Infrared Spectroscopy (FTIR), X-ray Powder Diffraction (XRD), Scanning Electron Microscopy (SEM), Thermo-Gravimetric Analysis (TGA), and Transmission Electron Microscopy (TEM) comprehensively confirmed the morphological properties of the prepared nanomaterials. Furthermore, bioconversion of two anionic dyes (i.e. Direct Red 23 (DR23) and Acid Blue 92 (AB92)) using nanobiocatalyst, was investigated and optimized. Also, the average decolorization effectiveness of the nanobiocatalyst was more than 75% for both of dyes after six cycles, implies its excellent operational stability and good reusability.

SiO2 microparticles with carbon nanotube-derived mesopores as an efficient support for enzyme immobilization

Novel mesoporous SiO2 microparticles were synthesized by spray pyrolysis using multiwalled carbon nanotubes (MCNTs) as a template. The synthesized multicompartment structure with uniform pores of 12.0 nm was used to immobilize lipase from Thermomyces lanuginosus. The total surface area of mesoporous SiO2 microparticles prepared from silica colloidal solution was increased by 26-folds compared to that of dense SiO2 particles (494 vs 19.0 m2 g−1, respectively). Mesoporous SiO2 particles showed 236% higher protein loading for lipase, than dense SiO2 particles. The maximum velocity (Vmax) and catalytic efficiencies of immobilized lipase were 3.80 and 5.90 folds higher than that of free enzyme. Contact angle analysis revealed increased hydrophobicity of the mesoporous particles, which is advantageous for lid opening at the active center, and increased activity after immobilization. We next developed a lipase/SiO2/glassy carbon electrode (GCE) biosensors. Cyclic voltammetric results showed linear responses of the lipase/SiO2/GCE bioelectrode towards tributyrin (50–300 mg dL−1) as a surface-limited reaction in Tris-HCl buffer. After 12 repetitive uses, dense SiO2- and mesoporous SiO2-bound lipase retained 74.2 and 95.4% of its original activities, respectively. Thus, given their desirable characteristics and industrial utility, greatly porous SiO2 particles may provide an excellent support for enzyme immobilization in biosensor development or biocatalysis in organic media.

A new hexamolybdate-based copper-2,2'-biimidazole coordination polymer serving as an acid catalyst and support for enzyme immobilization.

The hydrothermal reaction of (NH4)3[CoMo6O24H6]·7H2O (CoMo6), CuCl2·2H2O and 2,2'-biimidazole (H2biim) led to the formation of a new coordination polymer, namely poly[diaquabis(2,2'-biimidazole)hexa-μ3-oxo-octa-μ2-oxo-hexaoxodicopper(II)hexamolybdate(VI)], [Cu2Mo6O20(C6H6N4)2(H2O)2]n (Cu-Mo6O20), at pH 2-3. It is obvious that in the formation of crystalline Cu-Mo6O20, the original Anderson-type skeleton of heteropolymolybdate CoMo6 was broken and the new isopolyhexamolybdate Mo6O20 unit was assembled. In Cu-Mo6O20, one Mo6O20 unit connects four [Cu(H2biim)(H2O)]2+ ions in a pentacoordinate mode via four terminal O atoms, resulting in a tetra-supported structure, and each CuII ion is shared by two adjacent Mo6O20 units. Infinite one-dimensional chains are established by linkage between two adjacent Mo6O20 units and two CuII ions, and these chains are further packed into a three-dimensional framework by hydrogen bonds, π-π interactions and electrostatic attractions. The catalytic performance of this crystalline material used as an efficient and reusable heterogeneous acid catalyst for carbonyl-group protection is discussed. In addition, Cu-Mo6O20 was applied as a new support for enzyme (horseradish peroxidase, HRP) immobilization, forming immobilized enzyme HRP/Cu-Mo6O20. HRP/Cu-Mo6O20 showed good catalytic activity and could be reused.