Enhancing catalytic activity of multi-enzyme in vitro through substrate channeling effect is promis-ing yet challenging.Herein,conjugated microporous polymers(CMPs)-scaffolded integrated en-zyme cascade systems(I-ECSs...Enhancing catalytic activity of multi-enzyme in vitro through substrate channeling effect is promis-ing yet challenging.Herein,conjugated microporous polymers(CMPs)-scaffolded integrated en-zyme cascade systems(I-ECSs)are constructed through co-entrapping glucose oxidase(GOx)and horseradish peroxidase(HRP),in which hydrogen peroxide(H_(2)O_(2))is the intermediate product.The interplay of low-resistance mass transfer pathway and appropriate pore wall-H_(2)O_(2)interactions facilitates the directed transfer of H_(2)O_(2),resulting in 2.4-fold and 5.0-fold elevation in catalytic activ-ity compared to free ECSs and separated ECSs,respectively.The substrate channeling effect could be regulated by altering the mass ratio of GOx to HRP.Besides,I-ECSs demonstrate excellent stabili-ties in harsh environments and multiple recycling.展开更多
N-acetylneuraminic acid(NeuAc)is an important nutrient that plays a key role in brain development in infants NeuAc is mainly produced by extraction from natural resources such as edible birds’s nests,crucian eggs,cav...N-acetylneuraminic acid(NeuAc)is an important nutrient that plays a key role in brain development in infants NeuAc is mainly produced by extraction from natural resources such as edible birds’s nests,crucian eggs,caviar and human breast milk.The extraction process is complicated,resulting in the disadvantages of low NeuAc content and low recovery rate.In this study,a crude enzyme immobilization-based cell-free system(CEICFS)was developed for efficient NeuAc biosynthesis.First,N-terminal coding sequences that improved the expression levels of N-acetylglucosamine-2-epimerase(AGE)and N-acetylneuraminic acid aldolase(NanA)were obtained by high-throughput screening.And these sequences resulted in up to 1.5-fold(1.2-fold)increase in AGE(NanA)enzyme levels.And then,a CEICFS for NeuAc biosynthesis was proposed by directly immobilizing crude enzyme containing AGE and NanA on amino resin.Subsequently,NeuAc production from GlcNAc using CEICFS in one reactor was carried out,resulting 68 g/L of NeuAc and the highest productivity of 6.8 g/L/h.Further,the enzyme activity was still higher than 75%after five repeated uses.The functional properties of CEICFS were studied and compared to those of the free enzyme,immobilization can extend the application of enzyme to some harsh environments,such as low temperature and acidic environment.Therefore,CEICFS with excellent heat resistance,storage stability and reusability exhibit great potential for industrial application.展开更多
Enzyme cascade reactions play significant roles in bioelectrochemical processes because they permit more complex reactions. Co-immobilization of multienzyme on the electrode could help to facilitate substrate/intermed...Enzyme cascade reactions play significant roles in bioelectrochemical processes because they permit more complex reactions. Co-immobilization of multienzyme on the electrode could help to facilitate substrate/intermediate transfer among different enzymes and electron transfer from enzyme active sites to the electrode with high stability and retrievability. Different co-immobilization strategies to construct multienzyme bioelectrodes have been widely reported, however, up to now, they have barely been reviewed. In this review, we focus on recent state-of-the-art techniques for constructing co-immobilized multienzyme electrodes including random and positional co-immobilization. Particular attention is given to strategies such as multienzyme complex and surface display. Cofactor co-immobilization on the electrode is also crucial for the enhancement of catalytic reaction and electron transfer, yet, few studies have been reported. The up-to-date advances in bioelectrochemical applications of multienzyme bioelectrodes are also presented. Finally, key challenges and future perspectives are discussed.展开更多
Immobilization biocatalysis is a potential technology to improve the activity and stability of biocatalysts in nonaqueous systems for efficient industrial production.Alginate-chitosan(AC)microcapsules were prepared as...Immobilization biocatalysis is a potential technology to improve the activity and stability of biocatalysts in nonaqueous systems for efficient industrial production.Alginate-chitosan(AC)microcapsules were prepared as immobilization carriers by emulsifi cation-internal gelation and complexation reaction,and their contribution on facilitating the growth and metabolism of yeast cells were testifi ed successfully in culture medium-solvent biphasic systems.The cell growth in AC microcapsules is superior to that in alginate beads,and the cells in both immobilization carriers maintain much higher activity than free cells,which demonstrates AC microcapsules can confer yeast cells the ability to resist the adverse effect of solvent.Moreover,the performance of AC microcapsules in biphasic systems could be improved by adjusting the formation of outer polyelectrolyte complex(PEC)membrane to promote the cell growth and metabolic ability under the balance of resisting solvent toxicity and permitting substrate diffusion.Therefore,these findings are quite valuable for applying AC microcapsules as novel immobilization carriers to realize the biotransformation of value-added products in aqueous-solvent biphasic systems.展开更多
文摘Enhancing catalytic activity of multi-enzyme in vitro through substrate channeling effect is promis-ing yet challenging.Herein,conjugated microporous polymers(CMPs)-scaffolded integrated en-zyme cascade systems(I-ECSs)are constructed through co-entrapping glucose oxidase(GOx)and horseradish peroxidase(HRP),in which hydrogen peroxide(H_(2)O_(2))is the intermediate product.The interplay of low-resistance mass transfer pathway and appropriate pore wall-H_(2)O_(2)interactions facilitates the directed transfer of H_(2)O_(2),resulting in 2.4-fold and 5.0-fold elevation in catalytic activ-ity compared to free ECSs and separated ECSs,respectively.The substrate channeling effect could be regulated by altering the mass ratio of GOx to HRP.Besides,I-ECSs demonstrate excellent stabili-ties in harsh environments and multiple recycling.
基金the National Key Research and Development Program of China(2020YFA0908300)National Science Fund for Excellent Young Scholars(32222069)+2 种基金National Natural Science Foundation of China(32172349)Natural Science Foundation of Jiangsu Province(BK20200085)Key Research and Development Program of Jiangsu Province(BE2019628).
文摘N-acetylneuraminic acid(NeuAc)is an important nutrient that plays a key role in brain development in infants NeuAc is mainly produced by extraction from natural resources such as edible birds’s nests,crucian eggs,caviar and human breast milk.The extraction process is complicated,resulting in the disadvantages of low NeuAc content and low recovery rate.In this study,a crude enzyme immobilization-based cell-free system(CEICFS)was developed for efficient NeuAc biosynthesis.First,N-terminal coding sequences that improved the expression levels of N-acetylglucosamine-2-epimerase(AGE)and N-acetylneuraminic acid aldolase(NanA)were obtained by high-throughput screening.And these sequences resulted in up to 1.5-fold(1.2-fold)increase in AGE(NanA)enzyme levels.And then,a CEICFS for NeuAc biosynthesis was proposed by directly immobilizing crude enzyme containing AGE and NanA on amino resin.Subsequently,NeuAc production from GlcNAc using CEICFS in one reactor was carried out,resulting 68 g/L of NeuAc and the highest productivity of 6.8 g/L/h.Further,the enzyme activity was still higher than 75%after five repeated uses.The functional properties of CEICFS were studied and compared to those of the free enzyme,immobilization can extend the application of enzyme to some harsh environments,such as low temperature and acidic environment.Therefore,CEICFS with excellent heat resistance,storage stability and reusability exhibit great potential for industrial application.
基金supported by National Natural Science Foundation of China(6091000260974112+4 种基金6097108530970969)the National High Technology Research and Development Program(863 Program) (2009AA012413)Ph.D.Programs Foundation of Ministry of Education of China(20070001020)Postdoctoral Science Foundation of China(No .20080440257)
基金supported by the National Natural Science Foundation of China(21878324,21706273)the CAS Pioneer Hundred Talent Program(Type C,reference#2016-081)。
文摘Enzyme cascade reactions play significant roles in bioelectrochemical processes because they permit more complex reactions. Co-immobilization of multienzyme on the electrode could help to facilitate substrate/intermediate transfer among different enzymes and electron transfer from enzyme active sites to the electrode with high stability and retrievability. Different co-immobilization strategies to construct multienzyme bioelectrodes have been widely reported, however, up to now, they have barely been reviewed. In this review, we focus on recent state-of-the-art techniques for constructing co-immobilized multienzyme electrodes including random and positional co-immobilization. Particular attention is given to strategies such as multienzyme complex and surface display. Cofactor co-immobilization on the electrode is also crucial for the enhancement of catalytic reaction and electron transfer, yet, few studies have been reported. The up-to-date advances in bioelectrochemical applications of multienzyme bioelectrodes are also presented. Finally, key challenges and future perspectives are discussed.
基金Supported by the National Natural Science Foundation of China(No.21276033)the Open Foundation of the State Key Laboratory of Bioactive Seaweed Substances(Nos.SKL-BASS1707,SKL-BASS1711)the Liaoning Provincial BaiQianWan Talents Program(No.2017-6)
文摘Immobilization biocatalysis is a potential technology to improve the activity and stability of biocatalysts in nonaqueous systems for efficient industrial production.Alginate-chitosan(AC)microcapsules were prepared as immobilization carriers by emulsifi cation-internal gelation and complexation reaction,and their contribution on facilitating the growth and metabolism of yeast cells were testifi ed successfully in culture medium-solvent biphasic systems.The cell growth in AC microcapsules is superior to that in alginate beads,and the cells in both immobilization carriers maintain much higher activity than free cells,which demonstrates AC microcapsules can confer yeast cells the ability to resist the adverse effect of solvent.Moreover,the performance of AC microcapsules in biphasic systems could be improved by adjusting the formation of outer polyelectrolyte complex(PEC)membrane to promote the cell growth and metabolic ability under the balance of resisting solvent toxicity and permitting substrate diffusion.Therefore,these findings are quite valuable for applying AC microcapsules as novel immobilization carriers to realize the biotransformation of value-added products in aqueous-solvent biphasic systems.
