Oriented immobilization of enzymes helps to maintain their native structure and proper orientation for high-performance engineering to meet extensive biocatalysis demands.However,the supporting materials used for orie...Oriented immobilization of enzymes helps to maintain their native structure and proper orientation for high-performance engineering to meet extensive biocatalysis demands.However,the supporting materials used for orientated immobilization are usually costly or complicated in preparation,affecting their practical applications.In this work,a facile purification and immobilization method was proposed for enzyme immobilization based on organic-inorganic hybrid calcium phosphate nanocrystal(Ca Ps)induced by Cu^(2+) modified bovine serum albumin(BSA-Cu).Then,the as-prepared hybrid calcium phosphate nanosheet,BSA-Cu@Ca Ps,was utilized for one-pot purification and immobilization of His-tagged organophosphorus hydrolase(OPH)by metal-affinity binding to the incorporated BSA.BSA-Cu@Ca PsOPH exhibited enhanced p H stability and thermal stability compared to the free enzyme.Moreover,BSA-Cu@Ca Ps-OPH could retain more than 75%and 56%of initial activity after reuse 5 and 10 times,respectively.The results demonstrated that this facile strategy was promising for the effective biodegradation of organophosphorus pesticides with the immobilized enzyme.展开更多
The plant cellulose powder was activated by two different methods using 1,4-butanediol diglycidyl ether(BTDE)and 1,1′-Carbonyldiimidazole(CDI) as the chemical coupling agents.Organophosphorus hydrolase(OPH) from Flav...The plant cellulose powder was activated by two different methods using 1,4-butanediol diglycidyl ether(BTDE)and 1,1′-Carbonyldiimidazole(CDI) as the chemical coupling agents.Organophosphorus hydrolase(OPH) from Flavobacterium ATCC 27551 was immobilized on any of activated support through covalent bonding.The optimal conditions of affecting parameters on enzyme immobilization in both methods were found, and it was demonstrated that the highest activity yields of immobilized OPH onto epoxy and CDI treated cellulose were 68.32%and 73.51%, respectively.The surface treatment of cellulose via covalent coupling with BTDE and CDI agents was proved by FTIR analysis.The kinetic constants of the free and immobilized enzymes were determined, and it was showed that both immobilization techniques moderately increased the Kmvalue of the free OPH.The improvements in storage and thermal stability were investigated and depicted that the half-life of immobilized OPH over the surface of epoxy modified cellulose had a better growth compared to the free and immobilized enzymes onto CDI treated support.Also, the pH stability of the immobilized preparations was enhanced relative to the free counterpart and revealed that all enzyme samples would have the same optimum pH value for stability at 9.0.Additionally, the immobilized OPH onto epoxy and CDI activated cellulose retained about 59% and 68% of their initial activity after ten turns of batch operation, respectively.The results demonstrated the high performance of OPH enzyme in immobilized state onto an inexpensive support with the potential of industrial applications.展开更多
In last few decades there is exponential increase in use of organophosphorus (OP) compounds as pesticides and insecticides leading to adverse effect on human population and live stock. There is a great need to develop...In last few decades there is exponential increase in use of organophosphorus (OP) compounds as pesticides and insecticides leading to adverse effect on human population and live stock. There is a great need to develop portable analytical tools that are amenable for remediation and bioremediation process monitoring, where rapid analysis of large number of samples is essential. Determination of various organophosphorus compounds has been achieved by integrating biocomponents with different transducers. The close integration of the biological events with the generation of a signal offers the potential for fabricating compact and easy-to-use analytical tools of high sensitivity and specificity. With the availability of new materials, associated with new sensing techniques has led to remarkable innovations in the design and construction of organophosphorus biosensors. The present review describes the specifications of most of the electrochemical Organophosphorus biosensors reported till date.展开更多
We constructed an expression cassette of the organophosphorus pesticide degrading （opal） gene under the control of the E8 promoter. Then opd was transformed into tomato fruit using an agroinfiltration transient expr...We constructed an expression cassette of the organophosphorus pesticide degrading （opal） gene under the control of the E8 promoter. Then opd was transformed into tomato fruit using an agroinfiltration transient expression system. β-Glucuronidase （GUS） staining, reverse transcription-polymerase chain reaction （RT-PCR）, wavelength scanning, and fluorescent reaction were performed to examine the expression of the opd gene and the hydrolysis activity on coumaphos of organo- phosphorus hydrolase （OPH） in tomato fruit. The results show that the agroinfiltrated tomato fruit-expressed OPH had the maximum hydrolysis activity of about 11.59 Uhng total soluble protein. These results will allow us to focus on breeding transgenic plants that could not only enhance the degrading capability of fruit and but also hold no negative effects on pest control when spraying organophosphorus pesticides onto the seedlings in fields.展开更多
The worldwide application of organophosphorus pesticides(OPs)has promoted agricultural development,but their gradual accumulation in soil and water can seriously affect the central nervous system of humans and other m...The worldwide application of organophosphorus pesticides(OPs)has promoted agricultural development,but their gradual accumulation in soil and water can seriously affect the central nervous system of humans and other mammals.Organophosphorus hydrolase(OPH)is an effective enzyme that can catalyze the degradation of the residual OPs.However,the degradation products such as p-nitrophenol(p-NP)is still toxic.Thus,it is of great significance to develop a multi-functional support that can be simultaneously used for the immobilization of OPH and the further degradation of p-NP.Herein,a visible light assisted enzyme-photocatalytic integrated catalyst was constructed by immobilizing OPH on hollow structured Au-TiO_(2)(named OPH@H-Au-TiO_(2))for the degradation of OPs.The obtained OPH@H-Au-TiO_(2)can degrade methyl parathion to p-NP by OPH and then degrade p-NP to hydroquinone with low toxicity by using H-Au-TiO_(2)under visible light.OPH molecules were immobilized on HAu-TiO_(2)through adsorption method to prepare OPH@H-Au-TiO_(2).After 2.5 h of reaction,methyl parathion is completely degraded,and about 82.64%of the generated p-NP is further degraded into hydroquinone.After reused for 4 times,the OPH@H-Au-TiO_(2)retains more than 80%of the initial degradation activity.This research presents a new insight in designing and constructing multi-functional biocatalyst,which greatly expands the application scenarios and industrial value of enzyme catalysis.展开更多
A bacterium with the capability of degrading organphosphorus, identified as Pseudomonas pseudoaicaligenes, is isolated from OP-treated soil. The organphosphorus hydrolase OPHC2 from this bacterium has been purified an...A bacterium with the capability of degrading organphosphorus, identified as Pseudomonas pseudoaicaligenes, is isolated from OP-treated soil. The organphosphorus hydrolase OPHC2 from this bacterium has been purified and characterized. OPHC2 has optimum activity for the reaction at 65℃ and pH 9.0 with methyl parathion as a substrate, it also shows good thermal and pH stability. Most metal ions and chemicals have no effect on the activity of OPHC2. The analyses of nucleotide sequence encoding OPHC2 and amino acid sequence of OPHC2 show that there are lower homologies with those of organphosphorus hydrolase reported in GenBank.展开更多
基金supported by the National Key Research and Development Program of China(2021YFC2102801)the National Natural Science Foundation of China(21621004)。
文摘Oriented immobilization of enzymes helps to maintain their native structure and proper orientation for high-performance engineering to meet extensive biocatalysis demands.However,the supporting materials used for orientated immobilization are usually costly or complicated in preparation,affecting their practical applications.In this work,a facile purification and immobilization method was proposed for enzyme immobilization based on organic-inorganic hybrid calcium phosphate nanocrystal(Ca Ps)induced by Cu^(2+) modified bovine serum albumin(BSA-Cu).Then,the as-prepared hybrid calcium phosphate nanosheet,BSA-Cu@Ca Ps,was utilized for one-pot purification and immobilization of His-tagged organophosphorus hydrolase(OPH)by metal-affinity binding to the incorporated BSA.BSA-Cu@Ca PsOPH exhibited enhanced p H stability and thermal stability compared to the free enzyme.Moreover,BSA-Cu@Ca Ps-OPH could retain more than 75%and 56%of initial activity after reuse 5 and 10 times,respectively.The results demonstrated that this facile strategy was promising for the effective biodegradation of organophosphorus pesticides with the immobilized enzyme.
基金Supported by the Malek-Ashtar University of Technology(925826018,2015)
文摘The plant cellulose powder was activated by two different methods using 1,4-butanediol diglycidyl ether(BTDE)and 1,1′-Carbonyldiimidazole(CDI) as the chemical coupling agents.Organophosphorus hydrolase(OPH) from Flavobacterium ATCC 27551 was immobilized on any of activated support through covalent bonding.The optimal conditions of affecting parameters on enzyme immobilization in both methods were found, and it was demonstrated that the highest activity yields of immobilized OPH onto epoxy and CDI treated cellulose were 68.32%and 73.51%, respectively.The surface treatment of cellulose via covalent coupling with BTDE and CDI agents was proved by FTIR analysis.The kinetic constants of the free and immobilized enzymes were determined, and it was showed that both immobilization techniques moderately increased the Kmvalue of the free OPH.The improvements in storage and thermal stability were investigated and depicted that the half-life of immobilized OPH over the surface of epoxy modified cellulose had a better growth compared to the free and immobilized enzymes onto CDI treated support.Also, the pH stability of the immobilized preparations was enhanced relative to the free counterpart and revealed that all enzyme samples would have the same optimum pH value for stability at 9.0.Additionally, the immobilized OPH onto epoxy and CDI activated cellulose retained about 59% and 68% of their initial activity after ten turns of batch operation, respectively.The results demonstrated the high performance of OPH enzyme in immobilized state onto an inexpensive support with the potential of industrial applications.
文摘In last few decades there is exponential increase in use of organophosphorus (OP) compounds as pesticides and insecticides leading to adverse effect on human population and live stock. There is a great need to develop portable analytical tools that are amenable for remediation and bioremediation process monitoring, where rapid analysis of large number of samples is essential. Determination of various organophosphorus compounds has been achieved by integrating biocomponents with different transducers. The close integration of the biological events with the generation of a signal offers the potential for fabricating compact and easy-to-use analytical tools of high sensitivity and specificity. With the availability of new materials, associated with new sensing techniques has led to remarkable innovations in the design and construction of organophosphorus biosensors. The present review describes the specifications of most of the electrochemical Organophosphorus biosensors reported till date.
基金Project supported by the National Key Technology R&D Program of China(No.2007BAD59B06)the International Science and Technology Cooperation Program of China(No.2007DFA31260)
文摘We constructed an expression cassette of the organophosphorus pesticide degrading （opal） gene under the control of the E8 promoter. Then opd was transformed into tomato fruit using an agroinfiltration transient expression system. β-Glucuronidase （GUS） staining, reverse transcription-polymerase chain reaction （RT-PCR）, wavelength scanning, and fluorescent reaction were performed to examine the expression of the opd gene and the hydrolysis activity on coumaphos of organo- phosphorus hydrolase （OPH） in tomato fruit. The results show that the agroinfiltrated tomato fruit-expressed OPH had the maximum hydrolysis activity of about 11.59 Uhng total soluble protein. These results will allow us to focus on breeding transgenic plants that could not only enhance the degrading capability of fruit and but also hold no negative effects on pest control when spraying organophosphorus pesticides onto the seedlings in fields.
基金supported by the National Natural Science Foundation of China(Nos.21901058,21908040,and 21878068)Tianjin Enterprise Science and Technology Commissioner,China(21YDTPJC00810)+2 种基金Science Technology Research Project of Higher Education of Hebei Province,China(QN2021045)Hebei Province Postgraduate Innovation Funding Project,China(CXZZSS2021027)National College Student’s Science and Technology Innovation Project,China(202010080038)。
文摘The worldwide application of organophosphorus pesticides(OPs)has promoted agricultural development,but their gradual accumulation in soil and water can seriously affect the central nervous system of humans and other mammals.Organophosphorus hydrolase(OPH)is an effective enzyme that can catalyze the degradation of the residual OPs.However,the degradation products such as p-nitrophenol(p-NP)is still toxic.Thus,it is of great significance to develop a multi-functional support that can be simultaneously used for the immobilization of OPH and the further degradation of p-NP.Herein,a visible light assisted enzyme-photocatalytic integrated catalyst was constructed by immobilizing OPH on hollow structured Au-TiO_(2)(named OPH@H-Au-TiO_(2))for the degradation of OPs.The obtained OPH@H-Au-TiO_(2)can degrade methyl parathion to p-NP by OPH and then degrade p-NP to hydroquinone with low toxicity by using H-Au-TiO_(2)under visible light.OPH molecules were immobilized on HAu-TiO_(2)through adsorption method to prepare OPH@H-Au-TiO_(2).After 2.5 h of reaction,methyl parathion is completely degraded,and about 82.64%of the generated p-NP is further degraded into hydroquinone.After reused for 4 times,the OPH@H-Au-TiO_(2)retains more than 80%of the initial degradation activity.This research presents a new insight in designing and constructing multi-functional biocatalyst,which greatly expands the application scenarios and industrial value of enzyme catalysis.
文摘A bacterium with the capability of degrading organphosphorus, identified as Pseudomonas pseudoaicaligenes, is isolated from OP-treated soil. The organphosphorus hydrolase OPHC2 from this bacterium has been purified and characterized. OPHC2 has optimum activity for the reaction at 65℃ and pH 9.0 with methyl parathion as a substrate, it also shows good thermal and pH stability. Most metal ions and chemicals have no effect on the activity of OPHC2. The analyses of nucleotide sequence encoding OPHC2 and amino acid sequence of OPHC2 show that there are lower homologies with those of organphosphorus hydrolase reported in GenBank.
