ANALYST
·
2018
·Vol. 143
·期 11
·2501-2507
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IF 3.864
Yong Liu,Xiaolin Xu,Haitang Yang,Ensheng Xu,Shuangshuang Wu,Wei Wei,Jin Chen
Poly(ADP-ribose) polymerase-1 (PARP-1) is a highly conserved nuclear enzyme, which binds tightly to damaged DNA and plays a key role in DNA repair, recombination, proliferation, and genomic stability. However, due to the poor electrochemical and optical activity of PARP-1 and its product PAR, only a few studies on its activity detection method have been reported. Herein, we report a simple and sensitive colorimetric strategy to monitor PARP-1 activity based on enzyme-initiated auto-PARylation-controlled aggregation of hemin-graphene nanocomposites (H-GNs). PARP, activated by dsDNA, catalyzed its substrate nicotinamide adenine dinucleotide (NAD+) to polymerize as a poly(ADP-ribose) polymer (PAR). PAR possesses several negative charges, and its charge density is twice that of a single-stranded DNA, which greatly impacts the dispersibility of H-GNs; due to their peroxidase-like catalytic activities, H-GNs can catalyze the chromogenic reaction of TMB and H2O2. As a result, in the presence of different PARP-1 activities, the supernatant of the corresponding solution contained different amounts of dispersed H-GNs and showed different colors after the chromogenic reaction that could be discerned easily by the absorbance or the color changes of the solution. The method was simple, sensitive, and reliable. The proposed method displays a linear range from 0.05 to 1 U with a detection limit of 0.03 U. In addition, this new method has been successfully applied to detect PARP-1 activity in human serum and different cancer cells and evaluate PARP-1 inhibitors.
研究领域:
生物化学
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分子生物学
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癌症研究
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纳米技术
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DOI
PubMed
高影响因子
Biosensors-Basel
·
2023
·Vol. 13
·期 7
·723
·
IF 5.4
Fengli Gao,Gang Liu,Yishu Qiao,Xiuwen Dong,Lin Liu
This work reports the development of a fluorescence method for the detection of poly(ADP-ribose) polymerase-1 (PARP1), in which a phenylboronic acid-modified fluorescein isothiocyanate dye (FITC-PBA) was used to recognize the formed poly(ADP-ribose) (PAR) polymer. The detection system was designed by conjugating recombinant streptavidin (rSA) with PARP1-specific double-stranded DNA (dsDNA) through streptavidin–biotin interaction. Capture of PARP1 via rSA–biotin–dsDNA allowed for the poly-ADP-ribosylation (PARylation) of both rSA and PARP1 in a homogeneous solution. The resulting rSA–biotin–dsDNA/PAR conjugates were then captured and separated via the commercialized nitrilotriacetic acid–nickel ion-modified magnetic bead (MB-NTA-Ni) through the interaction between NTA–Ni on MB surface and oligohistidine (His6) tag in rSA. The PAR polymer could capture the dye of FITC-PBA through the borate ester interaction between the boronic acid moiety in PBA and thecis-diol group in ribose, thus causing a decrease in fluorescence signal. The PARylation of streptavidin and the influence of steric hindrance on PARylation efficiency were confirmed using reasonable detection strategies. The method showed a wide linear range (0.01~20 U) and a low detection limit (0.01 U). This work should be valuable for the development of novel biosensors for the detection of poly(ADP-ribose) polymerases and diol-containing species.Keywords:streptavidin;nitrilotriacetic acid;magnetic bead;poly(ADP-ribose) polymerase;boronic acid;immobilization-free
研究领域:
生物化学
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分析化学
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分子生物学
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生物技术
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DOI
PubMed
高影响因子
ACS Applied Materials & Interfaces
·
2016
·
IF 7.145
Yuanyuan Xu,Li Liu,Zhaoyin Wang,Zhihui Dai
A stable and reusable electrochemical biosensor for the label-free detection of poly(ADP-ribose) polymerase (PARP) is designed in this work. C-kit-1, a thiol-modified G-quadruplex oligonucleotide, is first self-assembled on a gold electrode surface. The G-quadruplex structure of c-kit-1 can specifically tether and activate PARP, resulting in the generation of negatively charged poly(ADP-ribose) polymer (PAR). On the basis of electrostatic attraction, PAR facilitates the surface accumulation of positively charged electrochemical signal molecules. Through the characterization of electrochemical signal molecules, the label-free quantification of PARP is simply implemented. On the basis of the proposed method, selective quantification of PARP can be achieved over the linear range from 0.01 to 1 U with a calculated detection limit of 0.003U. Further studies also demonstrate the applicability of the proposed method to biosamples revealing the broad potential in practical applications. Furthermore, inhibitor of PARP has also been detected with this biosensor. Meanwhile, benefited from self-assembly on solid surface, this biosensor possesses two important features, i.e., reusability and stability, which are desirable in related biosensors.
研究领域:
生物化学
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分子生物学
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分析化学
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产品关联
DOI
PubMed
高影响因子
TALANTA
·
2024
·
IF 5.6
Suling Yang,Yaliang Huang,Tingting Yang,Jingze Li,Jiaxin Tian,Lin Liu
Poly(ADP-ribose)polymerase-1 (PARP1) could be activated by binding to nucleic acids with specific sequences, thus catalyzing the poly-ADP-ribosylation (PARylation) of target proteins including PARP1 itself. Most of the previously reported electrochemical methods for the determination of PARP1 were relied on the electrostatic interactions, which required the pre-immobilization of DNA on an electrode for the capture of PARP1. Herein, we reported an “immobilization−free” electrochemical strategy for the assays of PARP1 on the basic of avidin-biotin interaction. Once PARP1 was activated by binding with the specific double-stranded DNA (dsDNA) in a homogeneous solution, the biotinylated nicotinamide adenine dinucleotide (biotin-NAD + ) was transferred onto PARP1, resulting in the formation of biotinylated PAR polymers. The resulting biotinylated PAR polymers were then captured by a neutravidin (NA)-modified electrode through avidin-biotin interactions. The rich biotin moieties in the PAR polymers allowed for the capture of NA-modified silver nanoparticles (NA-AgNPs) through the avidin-biotin interactions. The surface-tethered AgNPs produced a well-defined electrochemical signal due to the characteristic solid-state Ag/AgCl process. The “immobilization−free”, electrostatic interaction-independent electrochemical biosensor exhibited low background current, high sensitivity, and good stability. It has achieved the determination of PARP1 with a detection limit down to 0.7 mU. The biosensor was further applied to determine the inhibition efficiency of potential inhibitors with a satisfactory result. This method shows promising potential applications in PARP1-related clinical diagnosis and drug discovery.
研究领域:
生物化学
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医学诊断
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纳米技术
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF BIOLOGICAL CHEMISTRY
·
2025
·
IF 3.9
Bao-Di Ma,Jia-Yi Li,Jian-He Xu,Tao Yu,Xu-Dong Kong
Methanol dehydrogenase (MDH), a representative of Type III alcohol dehydrogenases (ADHs), plays a pivotal role in methanol assimilation pathways, making it a key enzyme for the biosynthesis of chemicals and fuels from one-carbon feedstocks. An activator protein belonging to the Nudix hydrolase family, ACT, was found to increase the activity of MDH by 40-fold. Despite the widespread observation of this in vitro activation phenomenon in pairs of type III alcohol dehydrogenases and Nudix hydrolases, the mechanistic details have remained unresolved for decades. Here, we uncover a regulation mechanism in which MDH activation arises from the hydrolytic removal of ADP-ribose (ADPR), a potent inhibitor derived from NAD + degradation, by the ADPRase activity of ACT. This discovery challenges the previously proposed 'activation' models, revealing that ACT-mediated ADPR clearance disinhibits MDH rather than directly enhancing catalysis. By combining crystallographic analysis, kinetics, and inhibition assays, we demonstrate that ADPR inhibits MDHs with submicromolar K i values, highlighting its potential regulatory role in metabolic networks. Our findings redefine the widespread ‘activation’ of type III ADHs, providing valuable insights into alcohol metabolism and new directions for engineering synthetic methanol utilization pathways.
研究领域:
生物化学
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结构生物学
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酶学
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代谢工程
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微生物学
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产品关联
DOI
PubMed
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2019
·
IF 6.393
Ensheng Xu,Haitang Yang,Lina Wu,Jin Chen,Wei Wei,Yong Liu,Songqin Liu
Since the expression of poly(ADP-ribose) polymerase-1 (PARP-1) was significantly up-regulated in several primary malignancies, it was important to construct a sensitive and rapid PARP-1 assay for early diagnosis of cancer. When PARP-1 was activated by active dsDNA, it generates a number of negatively charged poly(ADP-ribose) (PAR) polymers, which can be readily detected. Here, we developed a label-free electrochemical (EC) biosensor for PARP-1 activity based on the electrostatic attraction between negatively charged PAR and positively charged amino-functioned mesoporous silica films (NH 2 -MSFs). Negatively charged PAR was absorbed on the NH 2 -MSFs surface and suppressed the electrochemical signal as the PAR prevented the electroactive probe reaching to the ITO electrode. The electrochemical signal for PAR linearly decreased from 0.01 U to 1.2 U with a detection limit of 0.005 U, which is comparable to the PARP-1 biosensors reported before. The strategy has been used to evaluate PARP-1 inhibitors and to detect PARP-1 activity in human serum and real cancer cells lysate with satisfactory results, indicating that it was a promising candidate for point-of-care diagnostics.
研究领域:
生物化学
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医学诊断
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材料科学
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产品关联
DOI
高影响因子
SENSORS AND ACTUATORS B-CHEMICAL
·
2020
·
IF 7.1
Ning Xia,Daohong Wu,Ting Sun,Yilin Wang,Xiaoyu Ren,Feng Zhao,Lin Liu,Xinyao Yi
Poly(ADP-ribose) polymerase-1 (PARP-1), as a highly conserved nuclear enzyme, can catalyze the transfer of multiple ADP-ribose units from β-nicotinamide adenine dinucleotide (NAD + ) to substrate proteins, including PARP-1 itself. In this work, two boronic acid derivatives, ferroceneboronic acid (FcBA) and 4-mercaptophenylboronic acid (MPBA) were used as the signal probes for electrochemical and colorimetric detection of PARP-1, respectively. The detection platform was constructed by modification of double-stranded DNA (dsDNA) on the surface of magnetic beads (MBs). Once PARP-1 was captured by the MB-dsDNA, auto-PARylation was initiated and the PAR products consisting of abundant ribose units were covalently bound to FcBA or MPBA through the formation of boronate ester covalent bonds. Sequestration of FcBA or MPBA by the PAR-covered MBs led to the decrease in the electrochemical signal of FcBA or preventing the MPBA-triggered aggregation of gold nanoparticle indicators. The change of oxidation current or adsorption intensity (color) was dependent upon the concentration and activity of PARP-1. A wide linear range of 0.1 ∼ 50 U was obtained for the electrochemical or colorimetric assays. The methods have been used to evaluate the inhibition efficiency and monitor the PARP-1 activity in living and apoptotic cancer cells.
研究领域:
生物化学
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分析化学
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分子生物学
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癌症研究
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DOI
高影响因子
IMMUNOLOGICAL INVESTIGATIONS
·
2023
·
IF 2.8
Dongdong Dou,Meirong Bi,Xiuyun Li,Nan Zhang,Mi Xu,Aili Guo,Feng Li,Weiwei Zhu
Background Childhood asthma is a major global health concern. ADP-ribosylation factor 6 (ARF6) is a low-molecular-weight GTPase; however, its role in childhood asthma remains unclear.Methods Ovalbumin (OVA)-challenged neonatal mice and transforming growth factor-β1 (TGF-β1)-induced BEAS-2B cells were used as in vivo and in vitro models of childhood asthma, respectively.Results Upon OVA stimulation, ARF6 expression was upregulated in the lung tissue. Neonatal mice administered SehinH3 (an ARF6 inhibitor) exhibited improved pulmonary pathological injury, along with reduced inflammatory cell infiltration in the lungs and cytokine release in bronchial alveolar lavage fluid and serum (interleukin [IL]−3, IL−5, IL−13, IgE, and OVA-specific IgE). SehinH3 treatment restrained epithelial – mesenchymal transition (EMT) in the lungs of asthmatic mice, as evidenced by increased E-cadherin and decreased N-cadherin and α-smooth muscle actin expression. Different TGF-β1 exposures to BEAS-2B cells induced a time- and dose-dependent increase in ARF6 expression in vitro. Upon TGF-β1 stimulation, ARF6 knockdown repressed EMT and SehinH3 treatment caused similar results in BEAS-2B cells. The transcription factor E2F8 is involved in diverse biological functions and its increased expression was confirmed in vivo and in vitro. Dual-luciferase assays confirmed that E2F8 binds to the ARF6 promoter and promotes its transcriptional activity. In vitro results revealed that E2F8 silencing suppressed EMT, whereas rescue experiments showed that ARF6 overexpression partly reversed these phenomena.Conclusion Our study showed that ARF6 is associated with childhood asthma progression and may be positively regulated by E2F8. These results provide insight into the pathogenesis and treatment of childhood asthma.
研究领域:
免疫学
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呼吸病学
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分子生物学
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细胞生物学
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF HAZARDOUS MATERIALS
·
2024
·
IF 12.2
Liyi Wei,Cuiping Qi,Tingting Wang,Xiuping Jin,Xinli Zhou,Mingcui Luo,Mengxi Lu,Huijun Chen,Juanjuan Guo,Hui Wang,Dan Xu
Amoxicillin is a widely used antibiotic globally, and its pervasive environmental presence poses significant risks to human health and ecosystems. Notably, prenatal amoxicillin exposure (PAmE) may have long-term neurodevelopmental toxicity for offspring. In this study, we investigated the lasting effects of PAmE on depressive-like behaviors in offspring rats, emphasizing the biological mechanisms mediated by changes in gut microbiota and its metabolite, myristic acid. Our results showed that PAmE significantly disrupted the gut microbiota composition in offspring, particularly through the reduction of Lachnospiraceae , leading to decreased levels of myristic acid. This disruption hindered the N-myristoylation of ADP-ribosylation factor 1 (ARF1), impaired the normal degradation of the stimulator of interferon genes protein, inhibited autophagic processes, and promoted M1 polarization of microglia, ultimately leading to depressive-like behaviors in the offspring. Remarkably, supplementation with Lachnospira or myristic acid effectively reversed the PAmE-induced neurodevelopmental and behavioral abnormalities, alleviating depressive-like symptoms. This study reveals how PAmE affects offspring neurodevelopment and behavior through gut microbiota and myristic acid, highlighting the crucial role of the gut-brain axis in the modulation of depressive symptoms. Supplementing Lachnospira or myristic acid could represent a novel strategy to mitigate PAmE-induced fetal-originated depression, providing new biological evidence and potential therapeutic avenues.
研究领域:
药理学
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神经科学
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微生物学
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行为科学
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分子生物学
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DOI
PubMed
高影响因子
CLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY
·
2014
·Vol. 42
·期 2
·146-153
·
IF 2.405
Xiao-Yun Hu,Ji-Yun Liang,Xue-Jun Guo,Li Liu,Ya-Bing Guo
Summary The development of chemoresistance may reduce the efficacy of chemotherapeutic drugs for treating hepatocellular carcinoma (HCC). In the present study, the effects of apigenin on intensifying the chemosensitivity of HCC cells and an HCC xenograft model in response to 5-fluorouracil (5-FU) were investigated. Sub-toxic concentrations of apigenin (4 μ mol/L) significantly enhanced the cytotoxicity of 5-FU (100 μ g/mL) in HCC cells. In vivo , combined treatment with apigenin (20 mg/kg, five times/week for 3 weeks) and 5-FU (20 mg/kg for 5 consecutive days) significantly inhibited the growth of HCC xenograft tumours. Annexin V–propidium iodide dual staining assays, terminal deoxyribonucleotidyl transferase-mediated dUTP–digoxigenin nick end-labelling assays and western blotting analysis were used to confirm the synergistic effects of apigenin and 5-FU on HCC apoptosis. Coincubation of HCC cells with apigenin and 5-FU increased levels of reactive oxygen species (ROS), which was followed by a decrease in the mitochondrial membrane potential (ΔΨm). In addition, combined triggered the mitochondrial apoptotic pathway, as indicated by decreased Bcl-2 expression and loss of ΔΨm, with significant activation of caspase 3 and poly(ADP-ribose) polymerase. The present study is the first to demonstrate that apigenin may potentiate the cytotoxicity of 5-FU in HCC via inhibition of ROS-mediated drug resistance and concurrent activation of the mitochondrial pathways of apoptosis.
研究领域:
肿瘤学
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药理学
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细胞生物学
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产品关联
DOI
PubMed
高影响因子
Bioengineering & Translational Medicine
·
2023
·Vol. 8
·期 3
·e10507
·
IF 7.4
Jingyan Yi,Xin Luo,Jinshan Xing,Aharon Gedanken,Xiukun Lin,Chunxiang Zhang,Gan Qiao
Micelle Encapsulation Zinc-doped copper oxide nanocomposites (MEnZn-CuO NPs) is a novel doped metal nanomaterial prepared by our group based on Zinc doped copper oxide nanocomposites (Zn-CuO NPs) using non-micellar beam. Compared with Zn-CuO NPs, MEnZn-CuO NPs have uniform nanoproperties and high stability. In this study, we explored the anticancer effects of MEnZn-CuO NPs on human ovarian cancer cells. In addition to affecting cell proliferation, migration, apoptosis and autophagy, MEnZn-CuO NPs have a greater potential for clinical application by inducing HR repair defects in ovarian cancer cells in combination with poly (ADP-ribose) polymerase inhibitors for lethal effects.
研究领域:
纳米技术
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癌症研究
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药理学
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材料科学
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产品关联
DOI
PubMed
高影响因子
CELL BIOCHEMISTRY AND BIOPHYSICS
·
2014
·Vol. 71
·期 3
·1349-1356
·
IF 2.38
Jin Di,Cao Ming,Mu Xin,Yang Guoliang,Xue Wei,Huang Yiran,Chen Haige
Catalpol is an iridoid glucoside extracted from the traditional Chinese medicinal plant called Rehmannia glutinosa , and it is reported to possess a variety of pharmacological activities. The goal of this research was to explore the effect of catalpol on the human bladder cancer cells T24. The cells were treated for different durations with various concentrations of catalpol. Then the viability, mobility, and invasiveness of the cells were determined by MTT assay and flow cytometry, respectively. Catalpol was found to significantly suppress the proliferation, migration, and invasiveness of T24 bladder cancer cells in a dose- and time-dependent manner. The results also revealed that catalpol promoted apoptosis and caused the cell-cycle arrest at G2/M phase. Furthermore, the catalpol-mediated apoptosis was found to involve the modulation of PI3K/Akt pathway and inhibit the expression of anti-apoptotic B cell lymphoma-2 (Bcl-2) family proteins. Also, the expressions of Bcl-2 proapoptotic effectors, Bcl-2-associated X protein, and Bcl-2 associated death promoter were up-regulated in the cells treated with catalpol. The enzymes involved in the ultimate process of apoptosis, active caspase-3 and poly ADP ribose polymerase were elevated, and the latter was observed to be cleaved, indicating that catalpol-accelerated cell apoptosis involved a caspase-dependent pathway. These results suggest that catalpol might serve to be a promising therapeutic agent for the treatment of human bladder cancer.
研究领域:
肿瘤学
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药理学
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细胞生物学
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分子生物学
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产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）