A bacterium strain Y3,capable of efficiently degrading pendimethalin,was isolated from activated sludge and identified as Bacillus subtilis according to its phenotypic features and 16 S rRNA phylogenetic analysis.This...A bacterium strain Y3,capable of efficiently degrading pendimethalin,was isolated from activated sludge and identified as Bacillus subtilis according to its phenotypic features and 16 S rRNA phylogenetic analysis.This strain could grow on pendimethalin as a sole carbon source and degrade 99.5%of 100 mg/L pendimethalin within 2.5 days in batch liquid culture,demonstrating a greater efficiency than any other reported strains.Three metabolic products,6-aminopendimethalin,5-amino-2-methyl-3-nitroso-4-（pentan-3-ylamino） benzoic acid,and 8-amino-2-ethyl-5-（hydroxymethyl）-1,2-dihydroquinoxaline-6-carboxylic acid,were identified by HPLC-MS/MS,and a new microbial degradation pathway was proposed.A nitroreductase catalyzing nitroreduction of pendimethalin to 6-aminopendimethalin was detected in the cell lysate of strain Y3.The cofactor was nicotinamide adenine dinucleotide phosphate（NADPH） or more preferably nicotinamide adenine dinucleotide（NADH）.The optimal temperature and pH for the nitroreductase were 30℃ and 7.5,respectively.Hg^（2＋）,Ni^（2＋）,Pb^（2＋）,Co^（2＋）,Mn^（2＋） Cu^（2＋）,Ag~＋,and EDTA severely inhibited the nitroreductase activity,whereas Fe^（2＋）,Mg^（2＋）,and Ca^（2＋） enhanced it.This study provides an efficient pendimethalin-degrading microorganism and broadens the knowledge of the microbial degradation pathway of pendimethalin.展开更多
Tumor microenvironment(TME)-activatable probes have been proven to effectively increase signal-tobackground ratios(SBRs)and improve the success rate of complete tumor resection.However,many fluorescence probes have to...Tumor microenvironment(TME)-activatable probes have been proven to effectively increase signal-tobackground ratios(SBRs)and improve the success rate of complete tumor resection.However,many fluorescence probes have to be loaded into a nanocarrier for tumor targeted delivery,which consequently encounters poor drug loading,heterogeneous composition and non-encapsulated drug aggregates occurred during nanoformulation fabrications.Herein,a nitroreductase(NTR)-activated“OFF-ON”near-infrared fluorescence nanoprobe,named Nano Bodipy,was synthesized by the spontaneous self-assembling of NTRresponsive dye-polyethylene glycol(PEG)amphiphilic polymer in water.The NTR-responsive dye acted as the hydrophobic segment in the amphiphilic polymer,yielding a homogeneous composition and a high loading of 12.2 wt%(according to calculation)in the synthesized Nano Bodipy.The synthesized Nano Bodipy can efficiently accumulate in tumors via the enhanced permeability and retention(EPR)effect,enabling non-invasive tumor-targeted fluorescence imaging and guiding complete tumor resection.Once the synthesized Nano Bodipy entered the tumor cells,they dissociated and were activated by overexpressed NTR.With the real-time fluorescence guide of Nano Bodipy,complete tumor resection surgery was performed successfully.展开更多
Comprehensive SummaryThe detection of critical endogenous species, such as bacteria in microenvironments in the body, requires better imaging tools for visualization and monitoring of biological events. Bioluminescenc...Comprehensive SummaryThe detection of critical endogenous species, such as bacteria in microenvironments in the body, requires better imaging tools for visualization and monitoring of biological events. Bioluminescence imaging is the most popular strategy for obtaining real-time in living cells and organisms. Herein, we introduced a nitroaryl group on the C-3 position and a hydroxy group at the C-6 phenyl ring on furimazine to report the first bioluminescent probe (7) based on NanoLuc-furimazine bioluminescent pair for the detection of nitroreductase in bacteria. The probe, which possessed up to 560-fold intensity increase with a low detection limit of 16 ng/mL of nitroreductase, has the most efficient uncage efficiency in comparison with other bioluminescent congeners, thus enabling highly selective and sensitive visualization of NTR activity in a panel of clinical priority pathogens. Additionally, imaging of the recombinant strain as well as the NTR from mouse feces indicated the potential of this probe in the application of different mouse disease models.展开更多
Nitroreductase（NTR） is a member of flavin-containing enzymes that exists widely in bacteria. Hypoxia,which is a characteristic of locally advanced solid tumors, resulting from an imbalance between oxygen consumption...Nitroreductase（NTR） is a member of flavin-containing enzymes that exists widely in bacteria. Hypoxia,which is a characteristic of locally advanced solid tumors, resulting from an imbalance between oxygen consumption and supply, can result in NTR overexpression. Using either nicotinamide adenine dinucleotide（NADH） or nicotinamide adenine dinucleotide phosphate（NADPH） as a source of reducing equivalents, NTR can catalyze the reduction of nitroaromatic compounds to the corresponding amines.Based on this reduction mechanism, NTR can be applied not only in the bioremediation and degradation of organic nitrogen compounds, but also in the development of NTR-targeted fluorescent probes to detect the hypoxic status of cancer cells. This review aims to provide a summary of the progress in fluorescent probes for NTR in recent years and elucidate the main fluorescent mechanisms that have been applied to design probes.展开更多
Two-photon imaging has attracted increasing attention owing to its deep tissue imaging capabilities.Therefore,many fluorophores have been developed to satisfy its requirements.However,long-wavelength emission fluoroph...Two-photon imaging has attracted increasing attention owing to its deep tissue imaging capabilities.Therefore,many fluorophores have been developed to satisfy its requirements.However,long-wavelength emission fluorophores with an optically tunable group are rarely developed.In this study,two longwavelength emission fluorophores with an optically tunable amino group were successfully developed by introducing strong electron acceptor and large conjugated group to the TPQL dye.TPCO_(2)displayed a bright red emission(λem=638 nm,Φ=0.15)together with high two-photon action cross section and good water solubility,which enabled higher signal-to-background ratios and deep tissue imaging.The proof-of-concept probe(TPCO-NO_(2))was successfully applied to the high signal-to-background ratio imaging of nitroreductase in liver fibrosis,further realizing diagnosis of the degree of hypoxia during liver fibrosis.展开更多
Nanozymes,as a novel form of enzyme mimics,have garnered considerable interest.Despite overcoming the main disadvantages of their natural analogs,they still face challenges such as restricted mimic types and low subst...Nanozymes,as a novel form of enzyme mimics,have garnered considerable interest.Despite overcoming the main disadvantages of their natural analogs,they still face challenges such as restricted mimic types and low substrate specificity.Herein,we introduce a reactive ligand modification strategy to diversify enzyme mimic types.Specifically,we have utilized helical plasmonic nanorods(HPNRs)modified with para-nitrothiophenol(4-NTP)to create an oxygen-sensitive nitroreductase(NTR)with light-controllability.HPNRs act as a light-adjustable source of nicotinamide adenine dinucleotide/nicotinamide adenine dinucleotide phosphate(NAD(P)H),providing photon-generated energetic electrons to adsorbed 4-NTP molecules.In the presence of O_(2),the activated 4-NTP transfers the captured electron to the adsorbed O_(2),mimicking the electron transfer process in its natural counterpart.This enhanced O_(2)activation notably boosts the oxidative coupling of para-aminothiophenol(4-ATP).Density functional theory(DFT)calculations reveal that hot electrons injected into the lowest unoccupied molecular orbital(LUMO)energy level of 4-NTP can be transferred to that of molecular oxygen.In conclusion,our findings underline the potential of the reactive ligand modification strategy in developing new types of enzyme reactions,which opens up promising avenues for the enhancement and diversification of nanozyme functionalities.展开更多
The reduction of 4-nitrophenol catalyzed by nitroreductase in the presence of NADH was investigated in this paper. 4-Amino- phenol and 4-hydroxylamino-phenol were found in the reductive products. The relationship betw...The reduction of 4-nitrophenol catalyzed by nitroreductase in the presence of NADH was investigated in this paper. 4-Amino- phenol and 4-hydroxylamino-phenol were found in the reductive products. The relationship between reaction time and the reductive ratio were studied. The similar reducing ratios of 4-nitrophenol were obtained under aerobic and anaerobic conditions. The results indicated that an oxygen-insensitive reaction was proceeded in the reduction of 4-nitrophenol and nitroreductase was an oxygeninsensitive enzyme. The reductive products of 4-nitrophenol were determined by HPLC and MS.展开更多
Imaging hypoxia using fluorescence probes for nitroreductase（NTR） have attracted much attention in last decade. At least three different linkers have been commonly used to connect the recognition unit and reporting ...Imaging hypoxia using fluorescence probes for nitroreductase（NTR） have attracted much attention in last decade. At least three different linkers have been commonly used to connect the recognition unit and reporting unit in reported probes for NTR. Meanwhile, the linker is known to be a key factor for achieving best sensing performance. In this work, three near-infrared fluorescence probes CyNP-1, CyNP-2 and CyNP-3 were designed and synthesized from an aminocyanine dye CyNP. The three probes have the same recognition unit and same fluorescence reporting unit, but different linkers. CyNP-1 was found to have the best sensing performance for NTR with 40-fold of fluorescence enhancement. It is well investigated how the difference of the linkers brings out the different sensing performance by HPLC, MS and docking calculations. In the end, CyNP-1 was found to have good selectivity for NTR and used to imaging hypoxia in Hela cells.展开更多
Objective:To identify the frequencies(F) of ferredoxin and nitroreductase mutations were identified on Iranian clinical isolates of Giardia lamblia in order to predict whether the nitazoxanide can be prescribed as sui...Objective:To identify the frequencies(F) of ferredoxin and nitroreductase mutations were identified on Iranian clinical isolates of Giardia lamblia in order to predict whether the nitazoxanide can be prescribed as suitable drug for symptomatic to metronidazoleresistant giardiasis.Methods:Forty Giardia lamblia isolates as of 38 symptomatic and two metronidazole-resistant patients were collected from Iran.DNAs were extracted and amplified by targeting ferredoxin and Gl NR genes.The amplicons were directly sequenced to determine gene mutations.Results:The various amino acid substitutions(F:20%,Haplotype diversity:0.891,Tajima's D:-0.44013) were identified by analyzing ferredoxin gene in four symptomatic and two resistant isolates.Only,two haplotypes(F:5%,HD:0.345; Tajima's D:0.77815) characterized in metronidazole-resistant isolates of Gl NR,however,no point mutations was found in symptomatic isolates.Conclusions:Non-synonymous mutations of ferredoxin oxidoreductase gene reduce translational regulatory protein's binding affinity which concludes reduction of ferredoxin expression and its activity.This leads to decrease in metronidazole drug delivery into the cells.Mutations in these isolates may lead to their resistance to metronidazole.No to low synonymous mutations of Gl NR demonstrates that nitazoxanide can be prescribed as promising alternative treatment for symptomatic to metronidazole-resistant giardiasis in Iranian clinical isolates.展开更多
A novel quantitative electroanalysis method, triple potential step amperometry (TPSA), was developed and explained with an example of nitrobenzene analyzing in water. The selectivity of TPSA was im- proved by controll...A novel quantitative electroanalysis method, triple potential step amperometry (TPSA), was developed and explained with an example of nitrobenzene analyzing in water. The selectivity of TPSA was im- proved by controlling the potential step within a narrow interval and using enzyme-modified electrode, the narrow potential step makes the method avoid most interferents, and enzyme-modified electrode can enhance the response of target substance selectively. The peak area was investigated for quanti- tative calibration, such as nitrobenzene concentration showing a linear relation with the peak area, with the correlation coefficients being 0.9995. The t-test and F-test were applied to evaluating the reliability of TPSA, the results showed that there was no evidence of systematic error for TPSA, and the method was of no significant difference from CV. The merit of fast detecting and few potential changing times make the TPSA suitably applicable to low-cost automatic monitoring equipments.展开更多
基金supported by the National Science and Technology Support Plan(No.2012BAD15B03)the China Postdoctoral Science Foundation(Nos.2014M561660 and 2013T60546)the Jiangsu Postdoctoral Science Foundation(No.1301114C)
文摘A bacterium strain Y3,capable of efficiently degrading pendimethalin,was isolated from activated sludge and identified as Bacillus subtilis according to its phenotypic features and 16 S rRNA phylogenetic analysis.This strain could grow on pendimethalin as a sole carbon source and degrade 99.5%of 100 mg/L pendimethalin within 2.5 days in batch liquid culture,demonstrating a greater efficiency than any other reported strains.Three metabolic products,6-aminopendimethalin,5-amino-2-methyl-3-nitroso-4-（pentan-3-ylamino） benzoic acid,and 8-amino-2-ethyl-5-（hydroxymethyl）-1,2-dihydroquinoxaline-6-carboxylic acid,were identified by HPLC-MS/MS,and a new microbial degradation pathway was proposed.A nitroreductase catalyzing nitroreduction of pendimethalin to 6-aminopendimethalin was detected in the cell lysate of strain Y3.The cofactor was nicotinamide adenine dinucleotide phosphate（NADPH） or more preferably nicotinamide adenine dinucleotide（NADH）.The optimal temperature and pH for the nitroreductase were 30℃ and 7.5,respectively.Hg^（2＋）,Ni^（2＋）,Pb^（2＋）,Co^（2＋）,Mn^（2＋） Cu^（2＋）,Ag~＋,and EDTA severely inhibited the nitroreductase activity,whereas Fe^（2＋）,Mg^（2＋）,and Ca^（2＋） enhanced it.This study provides an efficient pendimethalin-degrading microorganism and broadens the knowledge of the microbial degradation pathway of pendimethalin.
基金supported by the National Natural Science Foundation of China(Nos.51903201,82172798)the Natural Science Foundation of Shaanxi Province(No.2023-YBSF-270)+1 种基金the Open Program of NHC Key Laboratory of Nuclear Medicine and Jiangsu Key Laboratory of Molecular Nuclear Medicine(No.KF202203)Horizontal Project of the First Affiliated Hospital of Xi'an Jiaotong University(No.202304174)。
文摘Tumor microenvironment(TME)-activatable probes have been proven to effectively increase signal-tobackground ratios(SBRs)and improve the success rate of complete tumor resection.However,many fluorescence probes have to be loaded into a nanocarrier for tumor targeted delivery,which consequently encounters poor drug loading,heterogeneous composition and non-encapsulated drug aggregates occurred during nanoformulation fabrications.Herein,a nitroreductase(NTR)-activated“OFF-ON”near-infrared fluorescence nanoprobe,named Nano Bodipy,was synthesized by the spontaneous self-assembling of NTRresponsive dye-polyethylene glycol(PEG)amphiphilic polymer in water.The NTR-responsive dye acted as the hydrophobic segment in the amphiphilic polymer,yielding a homogeneous composition and a high loading of 12.2 wt%(according to calculation)in the synthesized Nano Bodipy.The synthesized Nano Bodipy can efficiently accumulate in tumors via the enhanced permeability and retention(EPR)effect,enabling non-invasive tumor-targeted fluorescence imaging and guiding complete tumor resection.Once the synthesized Nano Bodipy entered the tumor cells,they dissociated and were activated by overexpressed NTR.With the real-time fluorescence guide of Nano Bodipy,complete tumor resection surgery was performed successfully.
基金supported by the Program of the National Natural Science Foundation of China(No.22177052,Y.Zhang,No.82074136,J.Guo,No.82272340,W.Chen)the Joint Training Program for Graduated Students of the Institute of Translational Research and Drug Innovation(X.Shi).
文摘Comprehensive SummaryThe detection of critical endogenous species, such as bacteria in microenvironments in the body, requires better imaging tools for visualization and monitoring of biological events. Bioluminescence imaging is the most popular strategy for obtaining real-time in living cells and organisms. Herein, we introduced a nitroaryl group on the C-3 position and a hydroxy group at the C-6 phenyl ring on furimazine to report the first bioluminescent probe (7) based on NanoLuc-furimazine bioluminescent pair for the detection of nitroreductase in bacteria. The probe, which possessed up to 560-fold intensity increase with a low detection limit of 16 ng/mL of nitroreductase, has the most efficient uncage efficiency in comparison with other bioluminescent congeners, thus enabling highly selective and sensitive visualization of NTR activity in a panel of clinical priority pathogens. Additionally, imaging of the recombinant strain as well as the NTR from mouse feces indicated the potential of this probe in the application of different mouse disease models.
基金financially supported by the National Natural Science Foundation of China (Nos. 81672508 and 61505076)Jiangsu Provincial Foundation for Distinguished Young Scholars (No. BK20170041)+2 种基金Jiangsu Key Research and Development Program (No. BE2015699)Natural Science Foundation of Guangdong Province (No. 2017A030313299)State Key Laboratory of Pulp and Paper Engineering (No. 201706)
文摘Nitroreductase（NTR） is a member of flavin-containing enzymes that exists widely in bacteria. Hypoxia,which is a characteristic of locally advanced solid tumors, resulting from an imbalance between oxygen consumption and supply, can result in NTR overexpression. Using either nicotinamide adenine dinucleotide（NADH） or nicotinamide adenine dinucleotide phosphate（NADPH） as a source of reducing equivalents, NTR can catalyze the reduction of nitroaromatic compounds to the corresponding amines.Based on this reduction mechanism, NTR can be applied not only in the bioremediation and degradation of organic nitrogen compounds, but also in the development of NTR-targeted fluorescent probes to detect the hypoxic status of cancer cells. This review aims to provide a summary of the progress in fluorescent probes for NTR in recent years and elucidate the main fluorescent mechanisms that have been applied to design probes.
基金supported by the National Natural Science Foundation of China(Nos.22074036,22004033,21877029)Special Funds for the Construction of Innovative Provinces in Hunan Province(No.2019RS1031)。
文摘Two-photon imaging has attracted increasing attention owing to its deep tissue imaging capabilities.Therefore,many fluorophores have been developed to satisfy its requirements.However,long-wavelength emission fluorophores with an optically tunable group are rarely developed.In this study,two longwavelength emission fluorophores with an optically tunable amino group were successfully developed by introducing strong electron acceptor and large conjugated group to the TPQL dye.TPCO_(2)displayed a bright red emission(λem=638 nm,Φ=0.15)together with high two-photon action cross section and good water solubility,which enabled higher signal-to-background ratios and deep tissue imaging.The proof-of-concept probe(TPCO-NO_(2))was successfully applied to the high signal-to-background ratio imaging of nitroreductase in liver fibrosis,further realizing diagnosis of the degree of hypoxia during liver fibrosis.
基金supported by the National Key Basic Research Program of China(No.2021YFA1202803)the National Natural Science Foundation of China(No.22072032)the Strategic Priority Research Program of Chinese Academy of Sciences(No.XDB36000000)。
文摘Nanozymes,as a novel form of enzyme mimics,have garnered considerable interest.Despite overcoming the main disadvantages of their natural analogs,they still face challenges such as restricted mimic types and low substrate specificity.Herein,we introduce a reactive ligand modification strategy to diversify enzyme mimic types.Specifically,we have utilized helical plasmonic nanorods(HPNRs)modified with para-nitrothiophenol(4-NTP)to create an oxygen-sensitive nitroreductase(NTR)with light-controllability.HPNRs act as a light-adjustable source of nicotinamide adenine dinucleotide/nicotinamide adenine dinucleotide phosphate(NAD(P)H),providing photon-generated energetic electrons to adsorbed 4-NTP molecules.In the presence of O_(2),the activated 4-NTP transfers the captured electron to the adsorbed O_(2),mimicking the electron transfer process in its natural counterpart.This enhanced O_(2)activation notably boosts the oxidative coupling of para-aminothiophenol(4-ATP).Density functional theory(DFT)calculations reveal that hot electrons injected into the lowest unoccupied molecular orbital(LUMO)energy level of 4-NTP can be transferred to that of molecular oxygen.In conclusion,our findings underline the potential of the reactive ligand modification strategy in developing new types of enzyme reactions,which opens up promising avenues for the enhancement and diversification of nanozyme functionalities.
文摘The reduction of 4-nitrophenol catalyzed by nitroreductase in the presence of NADH was investigated in this paper. 4-Amino- phenol and 4-hydroxylamino-phenol were found in the reductive products. The relationship between reaction time and the reductive ratio were studied. The similar reducing ratios of 4-nitrophenol were obtained under aerobic and anaerobic conditions. The results indicated that an oxygen-insensitive reaction was proceeded in the reduction of 4-nitrophenol and nitroreductase was an oxygeninsensitive enzyme. The reductive products of 4-nitrophenol were determined by HPLC and MS.
基金supported financially by the National Natural Science Foundation of China (Nos. 21421005, 21576038)the Fundamental Research Funds for the Central Universities of China (No. DUT16TD21)Science Program of Dalian City (Nos. 2014J11JH133, 2015J12JH207)
文摘Imaging hypoxia using fluorescence probes for nitroreductase（NTR） have attracted much attention in last decade. At least three different linkers have been commonly used to connect the recognition unit and reporting unit in reported probes for NTR. Meanwhile, the linker is known to be a key factor for achieving best sensing performance. In this work, three near-infrared fluorescence probes CyNP-1, CyNP-2 and CyNP-3 were designed and synthesized from an aminocyanine dye CyNP. The three probes have the same recognition unit and same fluorescence reporting unit, but different linkers. CyNP-1 was found to have the best sensing performance for NTR with 40-fold of fluorescence enhancement. It is well investigated how the difference of the linkers brings out the different sensing performance by HPLC, MS and docking calculations. In the end, CyNP-1 was found to have good selectivity for NTR and used to imaging hypoxia in Hela cells.
基金financially supported by Immunology Research Center,Tabriz University of Medical Sciences,Tabriz,Iranthe master’s thesis of the first author(Thesis No.93/2-4/12)
文摘Objective:To identify the frequencies(F) of ferredoxin and nitroreductase mutations were identified on Iranian clinical isolates of Giardia lamblia in order to predict whether the nitazoxanide can be prescribed as suitable drug for symptomatic to metronidazoleresistant giardiasis.Methods:Forty Giardia lamblia isolates as of 38 symptomatic and two metronidazole-resistant patients were collected from Iran.DNAs were extracted and amplified by targeting ferredoxin and Gl NR genes.The amplicons were directly sequenced to determine gene mutations.Results:The various amino acid substitutions(F:20%,Haplotype diversity:0.891,Tajima's D:-0.44013) were identified by analyzing ferredoxin gene in four symptomatic and two resistant isolates.Only,two haplotypes(F:5%,HD:0.345; Tajima's D:0.77815) characterized in metronidazole-resistant isolates of Gl NR,however,no point mutations was found in symptomatic isolates.Conclusions:Non-synonymous mutations of ferredoxin oxidoreductase gene reduce translational regulatory protein's binding affinity which concludes reduction of ferredoxin expression and its activity.This leads to decrease in metronidazole drug delivery into the cells.Mutations in these isolates may lead to their resistance to metronidazole.No to low synonymous mutations of Gl NR demonstrates that nitazoxanide can be prescribed as promising alternative treatment for symptomatic to metronidazole-resistant giardiasis in Iranian clinical isolates.
基金Supported by the National Natural Science Foundation of China (Grant Nos. 20577051 and 20607023)
文摘A novel quantitative electroanalysis method, triple potential step amperometry (TPSA), was developed and explained with an example of nitrobenzene analyzing in water. The selectivity of TPSA was im- proved by controlling the potential step within a narrow interval and using enzyme-modified electrode, the narrow potential step makes the method avoid most interferents, and enzyme-modified electrode can enhance the response of target substance selectively. The peak area was investigated for quanti- tative calibration, such as nitrobenzene concentration showing a linear relation with the peak area, with the correlation coefficients being 0.9995. The t-test and F-test were applied to evaluating the reliability of TPSA, the results showed that there was no evidence of systematic error for TPSA, and the method was of no significant difference from CV. The merit of fast detecting and few potential changing times make the TPSA suitably applicable to low-cost automatic monitoring equipments.
