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Enzyme immobilization

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https://www.readbyqxmd.com/read/28432497/immobilization-of-ulp1-protease-on-nhs-activated-sepharose-a-useful-tool-for-cleavage-of-the-sumo-tag-of-recombinant-proteins
#1
Qiujin Liang, Zhengzhi Huang, Yuan Zhang, Hongtao Li
OBJECTIVE: To fabricate an active and stable enzyme through covalent immobilization, a Ubl-specific protease (Ulp1) was used to cleave small ubiquitin-like modifier (SUMO) fusion proteins. RESULTS: We immobilized Ulp1 on N-hydroxysuccinimide (NHS)-activated Sepharose with a coupling efficiency of 1.7 mg/ml. The immobilized Ulp1 maintains 95% substrate-cleavage ability and significantly enhances pH and thermal stability, especially can withstand pH of 10.5. Besides resistance against some small molecules, the immobilized Ulp1 can tolerate 15% (v/v) DMSO and 20% (v/v) ethanol...
April 21, 2017: Biotechnology Letters
https://www.readbyqxmd.com/read/28428797/silicon-regulates-antioxidant-activities-of-crop-plants-under-abiotic-induced-oxidative-stress-a-review
#2
REVIEW
Yoon-Ha Kim, Abdul L Khan, Muhammad Waqas, In-Jung Lee
Silicon (Si) is the second most abundant element in soil, where its availability to plants can exhilarate to 10% of total dry weight of the plant. Si accumulation/transport occurs in the upward direction, and has been identified in several crop plants. Si application has been known to ameliorate plant growth and development during normal and stressful conditions over past two-decades. During abiotic (salinity, drought, thermal, and heavy metal etc) stress, one of the immediate responses by plant is the generation of reactive oxygen species (ROS), such as singlet oxygen ((1)O2), superoxide ([Formula: see text]), hydrogen peroxide (H2O2), and hydroxyl radicals (OH), which cause severe damage to the cell structure, organelles, and functions...
2017: Frontiers in Plant Science
https://www.readbyqxmd.com/read/28426220/fluorescent-neuraminidase-assay-based-on-supramolecular-dye-capture-after-enzymatic-cleavage
#3
Wenqi Liu, César F A Gómez-Durán, Bradley D Smith
A conceptually new type of enzymatic cleavage assay is reported that utilizes in-situ supramolecular capture of the fluorescent product. A squaraine-derived substrate with large blocking groups at each end of its structure cannot be threaded by a tetralactam macrocycle until the blocking groups are removed by enzyme cleavage. A prototype design responds to viral neuraminidase, an indicator of influenza infection, and also measures susceptibility of the sample to neuraminidase inhibitor drugs. The substrate structure incorporates three key features; (a) a bis(4-amino-3-hydroxyphenyl)squaraine core with bright deep-red fluorescence and excellent photostability, (b) an N-methyl group at each end of the squaraine core that ensures fast macrocycle threading kinetics, and (c) sialic acid blocking groups that prevent macrocycle threading until they are removed by viral neuraminidase...
April 20, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28422452/enantioselective-synthesis-of-s-naproxen-using-immobilized-lipase-on-chitosan-beads
#4
Saeedeh L Gilani, Ghasem D Najafpour, Hamid D Heydarzadeh, Aliakbar Moghadamnia
S-naproxen by enantioselective hydrolysis of racemic naproxen methyl ester was produced using immobilized lipase. The lipase enzyme was immobilized on chitosan beads, activated chitosan beads by glutaraldehyde, and Amberlite XAD7. In order to find an appropriate support for the hydrolysis reaction of racemic naproxen methyl ester, the conversion and enantioselectivity for all carriers were compared. In addition, effects of the volumetric ratio of two phases in different organic solvents, addition of cosolvent and surfactant, optimum pH and temperature, reusability, and inhibitory effect of methanol were investigated...
April 19, 2017: Chirality
https://www.readbyqxmd.com/read/28422254/a-dual-enzyme-inorganic-hybrid-nanoflower-incorporated-microfluidic-paper-based-analytic-device-%C3%AE-pad-biosensor-for-sensitive-visualized-detection-of-glucose
#5
Xueli Zhu, Jin Huang, Jinwen Liu, Hang Zhang, Jianhui Jiang, Ruqin Yu
A novel microfluidic paper-based analytic device (μPAD) biosensor is developed for sensitive and visualized detection of glucose. This biosensor is easily fabricated using the wax printing technique, with a hybrid nanocomplex composed of dual enzymes glucose oxidase (GOx) and horseradish peroxidase (HRP) and Cu3(PO4)2 inorganic nanocrystals incorporated in the detection zones. The hybrid nanocomplex is found to exhibit a flower-like structure, which allows co-immobilization of these two enzymes in a biocompatible environment...
April 19, 2017: Nanoscale
https://www.readbyqxmd.com/read/28421985/design-of-a-sensitive-and-selective-electrochemical-aptasensor-for-the-determination-of-the-complementary-cdna-of-mirna-145-based-on-the-intercalation-and-electrochemical-reduction-of-doxorubicin
#6
Maryam Mohamadi, Ali Mostafavi, Masoud Torkzadeh-Mahani
<p>The aim of this research was the determination of a microRNA (miRNA) using a DNA electrochemical aptasensor. In this biosensor, the complementary complementary DNA (cDNA) of miRNA-145 (a sense RNA transcript) was the target strand and the cDNA of miRNA-145 was the probe strand. Both cDNAs can be the product of the reverse transcriptase-polymerase chain reaction of miRNA. The proposed aptasensor's function was based on the hybridization of target strands with probes immobilized on the surface of a working electrode and the subsequent intercalation of doxorubicin (DOX) molecules functioning as the electroactive indicators of any double strands that formed...
April 19, 2017: Journal of AOAC International
https://www.readbyqxmd.com/read/28421414/polyclonal-antibodies-in-microplates-to-predict-the-maximum-adsorption-activities-of-enzyme-mutants-from-cell-lysates
#7
Yiran Feng, Xiaolan Yang, Deqiang Wang, Xiaolei Hu, Huimin Chong, Juan Liao, Chang-Guo Zhan, Fei Liao
With microplate-immobilized polyclonal antibodies against a starting enzyme or its active mutant bearing consistent accessible epitopes, the maximum activity of an adsorbed enzyme/mutant (Vs) was predicted for comparison to recognize weakly-positive mutants. Rabbit antisera against Escherichia coli alkaline phosphatase (ECAP) were fractionated with 33% ammonium sulfate to yield crude polyclonal antibodies for conventional immobilization in 96-well microplates. The response curve of the activities of ECAP/mutant adsorbed by the immobilized polyclonal antibodies to protein quantities from a cell lysate was fit to an approximation model to predict Vs...
April 19, 2017: Protein Journal
https://www.readbyqxmd.com/read/28419926/stability-and-activity-of-immobilized-trypsin-on-carboxymethyl-chitosan-functionalized-magnetic-nanoparticles-cross-linked-with-carbodiimide-and-glutaraldehyde
#8
Jun Sun, Lin Yang, Mengmeng Jiang, Yu Shi, Bin Xu, Hai-le Ma
Enzyme cross-linkers, such as 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and glutaraldehyde (GA), have been used to improve the stability of immobilized enzymes. We have developed a relatively stable and high-activity immobilized trypsin through EDC and GA cross-linking. Carboxymethyl chitosan (CM-CTS)-functionalized magnetic nanoparticles (Fe3O4@CM-CTS) were prepared, and characterized by Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis, X-ray diffraction, and transmission electron microscopy...
April 10, 2017: Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences
https://www.readbyqxmd.com/read/28419836/enzymatic-behavior-of-laccase-following-interaction-with-%C3%AE-cd-and-immobilization-into-pcl-nanofibers
#9
M Fatih Canbolat, Hasan Basri Savas, Fatih Gultekin
This study examines the effects of CD use on enzymatic activity, following enzyme immobilization into nanofibers. There is almost no research available on the change in enzyme activity following interaction with cyclodextrin and electrospun nanofiber mats together. Laccase enzyme was immobilized into nanofibrous structures by various techniques, with and without γ-CD addition, and the enzymatic activity of the laccase was analyzed. SEM, XRD, and FTIR analyses were used for the characterization of the resulting structures...
April 15, 2017: Analytical Biochemistry
https://www.readbyqxmd.com/read/28418039/solution-process-synthesis-of-high-aspect-ratio-zno-nanorods-on-electrode-surface-for-sensitive-electrochemical-detection-of-uric-acid
#10
Rafiq Ahmad, Nirmalya Tripathy, Min-Sang Ahn, Yoon-Bong Hahn
This study demonstrates a highly stable, selective and sensitive uric acid (UA) biosensor based on high aspect ratio zinc oxide nanorods (ZNRs) vertical grown on electrode surface via a simple one-step low temperature solution route. Uricase enzyme was immobilized on the ZNRs followed by Nafion covering to fabricate UA sensing electrodes (Nafion/Uricase-ZNRs/Ag). The fabricated electrodes showed enhanced performance with attractive analytical response, such as a high sensitivity of 239.67 μA cm(-2) mM(-1) in wide-linear range (0...
April 18, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28415403/design-and-synthesis-of-reduced-graphene-oxide-based-supramolecular-scaffold-a-benign-microbial-resistant-network-for-enzyme-immobilization-and-cell-growth
#11
Maloy Kr Mondal, Suprabhat Mukherjee, Swadhin K Saha, Pranesh Chowdhury, Santi P Sinha Babu
The present study describes the synthesis, characterization and biological application of reduced graphene oxide - chitosan (GC) based benign supramolecular scaffold (SMS). Various spectroscopic and microscopic analyses established the supramolecular interaction in between rGO and chitosan. The active performance of the developed material towards microbial resistivity, in vitro cell growth and as a scaffold for enzyme immobilizing matrix illustrates its unique implementation. Immobilization of polyphenol oxidase (PPO) onto GC lowers the Michaelis- Menten constant (Km) value and facilitates to achieve maximum velocity at low substrate concentration...
June 1, 2017: Materials Science & Engineering. C, Materials for Biological Applications
https://www.readbyqxmd.com/read/28414213/immobilization-and-stabilization-of-acylase-on-carboxylated-polyaniline-nanofibers-for-highly-effective-antifouling-application-via-quorum-quenching
#12
Jeongjoon Lee, Inseon Lee, Jahyun Nam, Dong Soo Hwang, Kyung-Min Yeon, Jungbae Kim
Acylase was immobilized and stabilized on carboxylated polyaniline nanofibers (cPANFs) for the development of antifouling nanobiocatalysis with high enzyme loading and stability. Acylase was immobilized via three different approaches: covalent attachment (CA), enzyme coating (EC), and magnetically separable enzyme precipitate coating (Mag-EPC). The enzyme activity per unit weight of cPANFs of Mag-EPC was 75 and 300 times higher than those of CA and EC, respectively, representing the improved enzyme loading in the form of Mag-EPC...
April 17, 2017: ACS Applied Materials & Interfaces
https://www.readbyqxmd.com/read/28414108/improvingthecatalytic-properties-and-stability-of-immobilized-%C3%AE-glutamyltranspeptidase-by-post-immobilization-with-pharmalyte-mt-8%C3%A2-10-5
#13
Fang Ni, Fan Zhang, Zhong Yao, Lijing Ye, Yun Sun, Haoqi Wang, Zhi Zhou, Benwei Zhu
γ-Glutamyltranspeptidase (GGT) is a dimeric protein that specifically catalyzes the transfer of γ-glutamyl in the optimum pH range of 8.5-9.0, but has poor in vitro stability under the alkaline conditions. In the present work, GGT was immobilized on a mesoporoustitania oxide whisker (MTWs) carrier to afford MTWs-GGT that was further modified with Pharmalyte(MT) (Phar) 8.0-10.5 to yield MTWs-GGT-Phar. Phar absorbed on MTWs-GGT to form a buffering layer with an isoelectric point of ∼9.2 that isolated the immobilized enzyme from the liquid bulk and significantly in proved the pH tolerance and stability of the immobilized GGT...
April 13, 2017: International Journal of Biological Macromolecules
https://www.readbyqxmd.com/read/28411651/enzyme-armoring-by-an-organosilica-layer-synthesis-and-characterization-of-hybrid-organic-inorganic-nanobiocatalysts
#14
M Rita Correro, Sabine Sykora, Philippe F-X Corvini, Patrick Shahgaldian
The availability of highly stable and reusable enzymes is one of the main challenges in bio-based industrial processes. Enzyme immobilization and encapsulation represent promising strategies to reach this goal. In this chapter, the synthetic strategy to produce hybrid organic/inorganic nanobiocatalysts (NBC) is reported. This strategy is based on the sequential immobilization of an enzyme on the surface of silica nanoparticles followed by the growth, at the surface of the nanoparticles, of a shielding layer which serves as an armor to protect the enzyme against denaturation/degradation...
2017: Methods in Enzymology
https://www.readbyqxmd.com/read/28411648/preparation-and-applications-of-dendronized-polymer-enzyme-conjugates
#15
Andreas Küchler, Daniel Messmer, A Dieter Schlüter, Peter Walde
Dendronized polymer-enzyme conjugates are large, water-soluble macromolecular structures built from a linear, fully synthetic, dendronized polymer (denpol), and several copies of enzyme molecules covalently bound to the peripheral functional groups of the denpol. Since denpol chains comprise repeating units with regularly branched side chains (dendrons), denpols have a cylindrical shape and are much thicker than conventional linear polymers. Depending on the dendron generation and chemical structure, denpols may have a large number of functional groups on their surface, exposed to the aqueous medium in which they are dissolved...
2017: Methods in Enzymology
https://www.readbyqxmd.com/read/28411642/semisynthetic-enzymes-by-protein-peptide-site-directed-covalent-conjugation-methods-and-applications
#16
Jose M Palomo
This chapter describes the rational design and synthesis of semisynthetic lipases by site-directed incorporation of tailor-made peptides on the lipase-lid site to improve its activity, specificity, and enantioselectivity in specific biotransformations. Cysteine was genetically introduced at a particular point of the oligopeptide lid of the enzyme, and cysteine-containing peptides, complementary to the amino acid sequence on the lid site of Geobacillus thermocatenulatus lipase (BTL), were covalently attached on the lid of two different cysteine-BTL variants based on a fast thiol-disulfide exchange ligation followed by desulfurization...
2017: Methods in Enzymology
https://www.readbyqxmd.com/read/28411639/nanoarmoring-of-enzymes-by-polymer-functionalized-iron-oxide-nanoparticles
#17
Gayan Premaratne, Leslie Coats, Sadagopan Krishnan
Polymer-armored enzymes loaded onto magnetic nanoparticles, as efficient nanobioreactors with enhanced properties, are described in this chapter. Polymers are useful macromolecules carrying a large number of surface charges and repeating units of desired chemical functional groups for linking enzymes onto them. Magnetic micro/nanoparticles have been widely used as enzyme carriers with the incorporation of suitable polymer layers. Synthesized iron oxide magnetic nanoparticles have been used to immobilize a peroxide-catalyzing enzyme-like heme protein: myoglobin using covalent and noncovalent strategies...
2017: Methods in Enzymology
https://www.readbyqxmd.com/read/28411636/armored-urease-enzyme-bioconjugated-poly-acrylamide-hydrogel-as-a-storage-and-sensing-platform
#18
Konda R Kunduru, S N Raju Kutcherlapati, Dhamodaran Arunbabu, Tushar Jana
Jack bean urease is an important enzyme not only because of its numerous uses in medical and other fields but also because of its historical significance-the first enzyme to be crystallized and also the first nickel metalloenzyme. This enzyme hydrolyzes urea into ammonia and carbon dioxide; however, the stability of this enzyme at ambient temperature is a bottleneck for its applicability. To improve urease stability, it was immobilized on different substrates, particularly on polymeric hydrogels. In this study, the enzyme was coupled covalently with poly(acrylamide) hydrogel with an yield of 18μmol/cm(3)...
2017: Methods in Enzymology
https://www.readbyqxmd.com/read/28411635/guide-to-the-preparation-of-molecularly-imprinted-polymer-nanoparticles-for-protein-recognition-by-solid-phase-synthesis
#19
Jingjing Xu, Paulina X Medina-Rangel, Karsten Haupt, Bernadette Tse Sum Bui
Molecularly imprinted polymers (MIPs) are synthetic antibody mimics possessing specific cavities designed for a target molecule. Nowadays, molecular imprinting of proteins still remains a challenge as the generation of selective imprinted cavities is extremely difficult, due to their flexible structure and the presence of a multitude of functional sites. To overcome this difficulty, we propose a solid-phase synthesis strategy to prepare MIPs specific for any protein that can be immobilized in an oriented way on a solid support...
2017: Methods in Enzymology
https://www.readbyqxmd.com/read/28411223/modified-bacteriophage-s16-long-tail-fiber-proteins-for-rapid-and-specific-immobilization-and-detection-of-salmonella-cells
#20
Jenna M Denyes, Matthew Dunne, Stanislava Steiner, Maximilian Mittelviefhaus, Agnes Weiss, Herbert Schmidt, Jochen Klumpp, Martin J Loessner
Bacteriophage-based assays and biosensors rival traditional antibody-based immunoassays for detection of low-level Salmonella contaminations. In this study, we harnessed the binding specificity of the long tail fiber (LTF) from bacteriophage S16 as an affinity molecule for immobilization, enrichment, and detection of Salmonella We demonstrate that paramagnetic beads (MBs) coated with recombinant gp37-gp38 LTF complexes (LTF-MBs) are highly effective tools for rapid affinity magnetic separation and enrichment of Salmonella Within 45 minutes, the LTF-MBs consistently captured over 95% of Salmonella Typhimurium cells from suspensions containing 10-10(5) CFU...
April 14, 2017: Applied and Environmental Microbiology
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