BIOPROCESS AND BIOSYSTEMS ENGINEERING
·
2020
·Vol. 44
·期 4
·727-735
·
IF 2.419
Wang Hongrao,Han Shuran,Wang Jiayi,Yu Shuyu,Li Xiaoyan,Lu Lei
Laccases are versatile oxidases that are capable of decolorizing various synthetic dyes. Recombinant Bacillus amyloliquefaciens laccase was immobilized as magnetic cross-linked enzyme aggregates (M-CLEAs) for application in dye decolorization. Several parameters influencing the activity recovery were evaluated during the synthesis of M-CLEAs. With ammonium sulfate as precipitant, maximum activity was recovered by cross-linking with 0.16% glutaraldehyde for 1 h. The prepared M-CLEAs exhibited improved activity under alkaline conditions. It remained 74% activity after incubation at 60 °C for 5 h. Enhanced tolerance towards NaCl was also observed for the M-CLEAs, with 68% activity remaining in the presence of 1 M NaCl. The immobilized laccase could rapidly decolorize more than 93% of reactive black 5 and indigo carmine in 1 h, while its catalytic efficiency towards reactive blue 19 was relatively low. After four cycles of consecutive reuse, the M-CLEAs could decolorize 92% of indigo carmine. The easy recovery and reusability of M-CLEAs facilitate the potential application of bacterial laccase in dye decolorization.
研究领域:
生物化学
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环境科学
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PubMed
高影响因子
SPECTROCHIMICA ACTA PART A-MOLECULAR AND BIOMOLECULAR SPECTROSCOPY
·
2020
·
IF 3.232
Xueliang Liu,Yingmin Zhou,Jin Liu,Hongjun Xia
Given the enzymatic properties and the oxidized surface of Pt nanomaterials, we demonstrated the intrinsic oxidase-like and peroxidase-like activities of platinum oxide (PtO 2 ). The surface clean PtO 2 nanoparticles with high water dispersibility were synthesized by a simple and green method. X-ray diffraction (XRD) and X-ray photoelectron spectra (XPS) results demonstrated that the prepared PtO 2 nanoparticles mainly consisted of Pt (Ⅳ) state without Pt(0) chemical state. The enzymatic activity of PtO 2 nanoparticles was verified by catalytic oxidation of several chromogenic substrates. Catalytic mechanism analysis suggested that PtO 2 nanopartiles acted as peroxidase and oxidase enzyme mimics by promoting the generation of the reactive oxygen species (ROS). By combining glucose oxidase, a colorimetric assay for glucose detection was developed with the limit of detection of 10.8 μM. The successful application of the proposed detection assay in human serum samples demonstrated the promising practical application in clinical diagnosis, pharmacy and food.
研究领域:
生物传感
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材料科学
·
临床诊断
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DOI
PubMed
高影响因子
ANALYTICA CHIMICA ACTA
·
2016
·
IF 4.712
Hao-Hua Deng,Guo-Lin Hong,Feng-Lin Lin,Ai-Lin Liu,Xing-Hua Xia,Wei Chen
Herein, we reported for the first time that gold nanoparticles-catalyzed 3,3′,5,5′-tetramethylbenzidine-H 2 O 2 system can serve as an ultrasensitive colorimetric pH indicator. Gold nanoparticles acted as a catalyst and imitated the function of horseradish peroxidase . The absorbance at 450 nm of the yellow-color product in the catalytic reaction exhibited a linear fashion over the pH range of 6.40–6.60. On the basis of this property, we constructed a novel sensing platform for the determination of urea, urease , and urease inhibitor. The limit of detection for urea and urease was 5 μM and 1.8 U/L, respectively. The half-maximal inhibition value IC 50 of acetohydroxamic acid was found to be 0.05 mM. Urea in human urine and urease in soil were detected with satisfied results.
研究领域:
生物化学
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分析化学
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DOI
PubMed
高影响因子
COLLOIDS AND SURFACES B-BIOINTERFACES
·
2023
·
IF 5.8
Mianxing Luo,Liang Wang,Guo Chen,Jun Zhao
The orientation of the enzyme molecular on the interface of the carrier affects its activity. Therefore, it is very important to controllably induce the orientation of the enzyme on the surface to improve the performance of the immobilized enzyme . Magnetic nanoparticles were used to construct microenvironments with the different surface hydrophobicity and charge characteristics by controlled modification, and those particles with various microenvironments were further used to study their interaction with the lipase . The amount and activity of immobilized enzyme on different magnetic nanoparticles surfaces were studied by physical adsorption and covalent binding. Through the enzyme surface and particle surface characteristics analysis, the possible preferred orientation of enzyme and enzyme conformation on different surfaces were inferred, which well explained the effect of surface induction on enzyme loading and activity. The methods of surface microenvironment regulation and the strategy of controllable induction of enzyme orientation adopted in this study are enlightening for the rational design of immobilized enzyme methods.
研究领域:
生物化学
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纳米生物学
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PubMed
高影响因子
Biotechnology Journal
·
2024
·Vol. 19
·期 4
·2300557
·
IF 3.2
Han-Yu Liu,Feng Qian,Hai-Min Zhang,Qian Gui,Yao-Wu Wang,Pu Wang
The halogenase-based catalysis is one of the most environmentally friendly methods for the synthesis of halogenated products, among which flavin-dependent halogenases (FDHs) have attracted great interest as one of the most promising biocatalysts due to the remarkable site-selectivity and wide substrate range. However, the complexity of constructing the NAD + -NADH-FAD-FADH 2 bicoenzyme cycle system has affected the engineering applications of FDHs. In this work, a coenzyme self-sufficient tri-enzyme fusion was constructed and successfully applied to the continuous halogenation of L -tryptophan. Sp FDH was firstly identified derived from Streptomyces pratensis , a highly selective halogenase capable of generating 6-chloro-tryptophan from tryptophan. Then, using gene fusion technology, Sp FDH was fused with glucose dehydrogenase (GDH) and flavin reductase (FR) to form a tri-enzyme fusion, which increased the yield by 1.46-fold and making the coenzymes self-sufficient. For more efficient halogenation of L -tryptophan, a continuous halogenation bioprocess of L -tryptophan was developed by immobilizing the tri-enzyme fusion and attaching it to a continuous catalytic device, which resulted in a reaction yield of 97.6% after 12 h reaction. An FDH from S. pratensis was successfully applied in the halogenation and our study provides a concise strategy for the preparation of halogenated tryptophan mediated by multienzyme cascade catalysis.
研究领域:
生物技术
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生物化学
·
酶学
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DOI
PubMed
高影响因子
SPECTROCHIMICA ACTA PART A-MOLECULAR AND BIOMOLECULAR SPECTROSCOPY
·
2019
·
IF 2.931
Yanqing Wang,Qianqian Han,Hongmei Zhang
Among the side effects of 5-fluorouracil (5-FU), the performance of the gastrointestinal reactions is faster and more obvious than others. In this work, the effects of 5-FU on the activities and conformational structures of the important digestive enzymes including α-amylase, pepsin and trypsin were studied to analyze the mechanism of the gastrointestinal adverse effects causing by 5-FU binding. The results showed that the enzymatic activity of pepsin was obviously reduced by the presence of 5-FU that bound directly to the enzyme activity cavity site. The molecular modeling and fluorescence quenching data indicated that the hydrophobic, polar and hydrogen bonding forces were involved in the ground state complex formation between proteases and 5-FU. In addition, 5-FU changed the tertiary structures of α-amylase, pepsin, and trypsin.
研究领域:
药理学
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生物化学
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DOI
PubMed
高影响因子
MOLECULES
·
2023
·Vol. 28
·期 10
·4170
·
IF 4.6
Jing Chen,Qi Huang,Zhuobin He,Guoying Tan,Yuansheng Zou,Juying Xie,Zhengming Qian
Polygoni Cuspidati Rhizoma et Radix (PCR), the rhizome and root ofPolygonum cuspidatumSieb. et Zucc., has been used as an herbal medicine for a long time. In this study, the ultrafiltration combined with high performance liquid chromatography (UF-HPLC) method was developed to screen tyrosinase (TYR), α-glucosidase (α-GLU), and xanthine oxidase (XOD) inhibitors from PCR. Firstly, the inhibitory activity of 50% methanol PCR extract on TYR, α-GLU, XOD, and acetylcholinesterase (ACHE) was tested. The extract showed a good inhibition on the enzymes, except for ACHE. Therefore, UF-HPLC experiments were carried out to screen TYR, α-GLU, and XOD inhibitors from PCR extract. Seven potential bioactive components were discovered, including methylgallate (1), 1,6-di-O-galloyl-D-glucose (2), polydatin-4′-O-D-glucoside (3), resveratrol-4′-O-D-glucoside (4), polydatin (5), malonyl glucoside resveratrol (6), and resveratrol-5-O-D-glucoside (7). Most of them were found as enzyme inhibitors from PCR for the first time, except polydatin (5), which had been reported as an α-GLUI in PCR in the literature. Finally, molecular docking analysis was applied to validate the interactions of these seven potential active components with the enzymes. Compounds1–7were proven as TYR inhibitors, compounds2,4–7were identified as XOD inhibitors, and compounds4–6were confirmed as α-GLU inhibitors. In short, the current study provides a good reference for the screening of enzyme inhibitors through UF-HPLC, and provides scientific data for future studies of PCR.Keywords:Polygoni Cuspidati Rhizoma et Radix;enzyme inhibitor;tyrosinase;xanthine oxidase;α-glucosidase;ultrafiltration
研究领域:
药理学
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生物化学
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分析化学
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PubMed
高影响因子
JOURNAL OF NANOSCIENCE AND NANOTECHNOLOGY
·
2018
·Vol. 18
·期 4
·2971-2978
·
IF 1.354
Yang Yuxiang,Zhao Min,Yao Pingping,Huang Yan,Dai Zuocheng,Yuan Hongming,Ni Chaoying
In the present work, three differently shaped mesoporous silica nanoparticles, spherical nano-SiO2, tubular mesoporous SiO2 and vesicle-like mesoporous SiO2 (VSL), were prepared and used to immobilize Horse radish peroxidase (HRP), and their enzyme’s
activity was also evaluated. It was found that the VSL immobilized HRP displayed higher specific activity than free enzyme and other two differently shaped silica immobilized HRP. After immobilization, the thermal stability, pH tolerance resistance and storage stability on vesicle-like SiO2
were studied as well. In addition, the kinetic constants K
m and V
max for HRP were significantly altered by immobilization. The affinity for HRP towards its substrate increased (with decreasing K
m), leading to enhanced catalytic efficiency
(with increased V
max). Moreover, the reusability for degradation of methoxychlor (MXC) by VSL immobilized enzyme was studied and its degradation products were detected by GC-MS and NMR analysis.
研究领域:
生物化学
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环境科学
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纳米技术
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DOI
高影响因子
APPLIED AND ENVIRONMENTAL MICROBIOLOGY
·
2016
·Vol. 82
·期 22
·
IF 3.823
Xiaodan Yan,Jianjun Wang,Yu Sun,Junge Zhu,Sheng Wu
Promiscuous enzymes are generally considered to be starting points in the evolution of offspring enzymes with more specific or even novel catalytic activities, which is the molecular basis of producing new biological functions. Mhg, a typical α/β fold hydrolase, was previously reported to have both γ-lactamase and perhydrolase activities. However, despite having high structural similarity to and sharing an identical catalytic triad with an extensively studied esterase from Pseudomonas fluorescens, this enzyme did not show any esterase activity. Molecular docking and sequence analysis suggested a possible role for the entry of the binding pocket in blocking the entrance tunnel, preventing the ester compounds from entering into the pocket. By engineering the entrance tunnel with only one or two amino acid substitutions, we successfully obtained five esterase variants of Mhg. The variants exhibited a very broad substrate acceptance, hydrolyzing not only the classical p-nitrophenol esters but also various types of chiral esters, which are widely used as drug intermediates. Site 233 at the entrance tunnel of Mhg was found to play a pivotal role in modulating the three catalytic activities by adjusting the size and shape of the tunnel, with different amino acid substitutions at this site facilitating different activities. Remarkably, the variant with the L233G mutation was a very specific esterase without any γ-lactamase and perhydrolase activities. Considering the amino acid conservation and differentiation, this site could be a key target for future protein engineering. In addition, we demonstrate that engineering the entrance tunnel is an efficient strategy to regulate enzyme catalytic capabilities.IMPORTANCE Promiscuous enzymes can act as starting points in the evolution of novel catalytic activities, thus providing a molecular basis for the production of new biological functions. In this study, we identified a critical amino acid residue (Leu233) at the entry of the substrate tunnel of a promiscuous enzyme, Mhg. We found that substitution of this residue with smaller amino acids such as Gly, Ala, Ser, or Pro endowed the enzyme with novel esterase activity. Different amino acids at this site can facilitate different catalytic activities. These findings exhibited universal significance in this subset of α/β fold hydrolases, including Mhg. Furthermore, we demonstrate that engineering the entrance tunnel is an efficient strategy to evolve new enzyme catalytic capabilities. Our study has important implications for the regulation of enzyme catalytic promiscuity and development of protein engineering methodologies.
研究领域:
生物化学
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蛋白质工程
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PubMed
高影响因子
BIOTECHNOLOGY PROGRESS
·
2022
·Vol. 38
·期 4
·e3256
·
IF 2.909
Duanhao Wang,Meijing Guo,Jiachen Li,Binglin Li,Jiao Wang
Six kinds of Mo-basing nanomaterials (MoO 3 , MoO 3 @Ru, Mo-PDA, MoP C , MoP, CNT@MoS 2 ) were successfully synthesized, which were employed as carriers to immobilize phospholipase A 1 (PLA1) for the hydrolysis of phospholipids (PLs). PLA1 was immobilized by a simple adsorption-precipitation-cross-linking to form an “enzyme net” covering on nanoparticles. The greatest advantage of these nanoparticles was their strong hydrophilic surface. It not only permitted their dispersion in the aqueous phase, but also showed the strong affinity for PLs in the organic phase, because amphiphilic PLs had the polar head group and higher hydrophilicity than other oils components. Michaelis–Menten analysis revealed that higher catalytic activity and enzyme−substrate affinity were observed in several immobilized PLA1 than its free form. MoO 3 was confirmed to be the best candidate for carrier. The highest specific activity of MoO 3 -immobilized PLA1 reached 43.1 U/mg, which was about 1.8 times higher than that of free PLA1 (24.4 U/mg). In addition, the stability and recycling were also enhanced. The robust immobilized PLA1 was prepared in this work, showing great potential for the enzymatic degumming.
研究领域:
生物化学
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纳米生物学
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食品科学
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DOI
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2018
·
IF 5.667
Xiaoyun Lin,Zhenmao Zhu,Chengfei Zhao,Shaoguang Li,Qicai Liu,Ailin Liu,Liqing Lin,Xinhua Lin
In this study, we developed a simple, sensitive and label-free colorimetric sensor for trypsin and its inhibitor using oxidase mimicking activity of protamine-stabilized platinum nanoparticles units (Pro-PtNPs units). Protamine is designed to stabilize PtNPs and served as the substrate of trypsin hydrolysis. The results show that protamine combines several PtNPs together to form active units, which have stronger oxidase mimicking activity than other PtNPs. In addition, the Michaelis constant of the Pro-PtNPs units is smaller than negatively charged citrate capped PtNPs, suggesting that the positively charged Pro-PtNPs units have better affinity with 3,3′,5,5′-tetramethylbenzidine (TMB), which is inconsistent with previous reports. The protamine stabilizer could be hydrolyzed in the presence of trypsin, and the Pro-PtNPs units would be decomposed and aggregated, resulting in a decrease of catalytic activity of Pro-PtNPs units. The color of the solution would be reduced according to the decrease of catalytic activity, so the catalytic activity is linearly related to UV absorbance. The linear range of trypsin detected by this new method is 0.06 μg/mL˜0.6 μg/mL, and the detection limit down to 0.03 μg/mL. Pro-PtNPs units have been successfully applied to the determination of serum trypsin and screening of inhibitors.
研究领域:
生物化学
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纳米医学
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DOI
高影响因子
ANALYTICAL CHEMISTRY
·
2018
·
IF 6.042
Jiahui Zhao,Shuang Wang,Shasha Lu,Xingfu Bao,Jian Sun,Xiurong Yang
An enzyme cascade-triggered reaction with novel signal generation mechanism is beneficial for the development and insight of the enzyme cascade, which is extensively used for signal transduction in potential applications. Inspired by the fluorogenic and chromogenic reaction between dopamine and resorcinol, and the specific catalytic properties of alkaline phosphatase (ALP) and tyrosinase, we designed and synthesized an unconventional substrate of ALP, named p-aminoethyl-phenyl phosphate disodium salt (PAPP). As expected, the ALP and tyrosinase-incubated PAPP solution exhibited pale yellow with intense blue fluorescence upon addition of resorcinol, owing to the ALP-catalyzed transformation of PAPP into an intermediate tyramine, and the tyrosinase-catalyzed hydroxylation of tyramine to dopamine, as well as the specific reaction between dopamine and resorcinol. Therefore, an enzyme cascade system has been developed herein based on the ALP and tyrosinase coupled enzymes-triggered fluorogenic and chromogenic reaction. According to the direct relationship between the activity of ALP/tyrosinase and absorbance/fluorescence intensity of the resultant solution, the proposed enzyme cascade-triggered reaction was utilized for assaying ALP and tyrosinase activity with fluorometric and colorimetric dual-readout signals. Furthermore, such enzyme cascade catalysis process was integrated into the ALP-based cascade enzyme-linked immunosorbent assay with dual-readout signals, resulting in the sensitive detection of cardiac troponin I in diluted serum.
研究领域:
生物化学
·
医学检测
·
免疫分析
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DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）