We found a novel lipase gene in the Paenibacillus pasadenensis CS0611 strain.The lipase gene sequence was cloned into the pET-28a expression vector to construct a recombinant lipase protein containing 6×His tags ...We found a novel lipase gene in the Paenibacillus pasadenensis CS0611 strain.The lipase gene sequence was cloned into the pET-28a expression vector to construct a recombinant lipase protein containing 6×His tags at the C-and N-termini,respectively.High-level expression of the lipase in E.coli BL21(DE3)was obtained upon induction with IPTG at 20°C.The recombinant lipase activity was approximately 1631-fold higher than the wild type.His-tagged recombinant lipase was purified rapidly and efficiently by using Ni-charged affinity chromatography with 63.5%recovery and a purification factor of 10.78.The purified lipase was stable in a broad range of temperatures and pH values,with the optimal temperature and pH being 50°C and 7.0,respectively.Its activity was stimulated to different degrees in the presence of metal ions such as Ca2+,Mg2+,and some non-ionic surfactants.In addition,the purified lipase was activated by a series of water-miscible organic solvents such as some short carbon chain alcohols and was highly tolerant to some water-immiscible organic solvents.展开更多
An extracellular chitinase produced by Paenibacillus pasadenensis CS0611was purified by ammoniumsulfate precipitation,HiTrap DEAE FF and HiLoad26/600Superdex200pg column chromatography.The apparent molecular mass dete...An extracellular chitinase produced by Paenibacillus pasadenensis CS0611was purified by ammoniumsulfate precipitation,HiTrap DEAE FF and HiLoad26/600Superdex200pg column chromatography.The apparent molecular mass determined by sodium dodecyl sulfate polyacrylamide gelelectrophoresis was69kDa.The optimum pH and optimum temperature of the chitinase were5.0and50°C,respectively.The enzyme showed high stability at alkaline pH values and temperaturesbelow40°C.Additionally,the metal ions Mn2+,Mg2+,and Co2+inhibited activity of the chitinase.Thechitinase was active on colloidal chitin with an apparent Km of4.41mg/mL and Vmax of1.08mg/min.Substrate spectrum analysis indicated that the chitinase reacted preferentially with the glucosidicbond between GlcNAc‐GlcNAc.The enzymatic hydrolysate was analyzed by high‐performance liquidchromatography and thin layer chromatography,and clearly showed that a subunit of(GlcNAc)2was the main hydrolysis product.展开更多
Electrocatalytic hydrogenation(ECH)offers a sustainable route for the conversion of biomass-derived feedstocks under ambient conditions;however,an atomic-level understanding of the catalytic mechanism based on heterog...Electrocatalytic hydrogenation(ECH)offers a sustainable route for the conversion of biomass-derived feedstocks under ambient conditions;however,an atomic-level understanding of the catalytic mechanism based on heterogeneous electrodes is lacking.To gain insights into the relation between electrocatalysis and the catalyst surface configuration,herein,the facet dependence of the ECH of furfural(FAL)is investigated on models of nanostructured Pd cubes,rhombic dodecahedrons,and octahedrons,which are predominantly enclosed by{100},{110},and{111}facets,respectively.The facet-dependent specific activity to afford furfuryl alcohol(FOL)follows the order of{111}>{100}>{110}.Experimental and theoretical kinetic analyses confirmed the occurrence of a competitive adsorption Langmuir-Hinshelwood mechanism on Pd,in which the ECH activity can be correlated with the difference between the binding energies of chemisorbed H(^(*)H)and FAL(^(*)FAL)based on density functional theoretical(DFT)calculations.Among the three facets,Pd{111}exhibiting the strongest^(*)H but the weakest^(*)FAL showed the copresence of the^(*)H and^(*)FAL intermediates on the Pd surface for subsequent hydrogenation,experimentally confirming its high ECH activity and Faradaic efficiency.The free energies determined using DFT calculations indicated that^(*)H addition to the carbonyl of FAL on Pd{111}was thermodynamically preferred over desorption to gaseous H2,contributing to efficient ECH to afford FOL at the expense of H2 evolution.The obtained insights into the facet-dependent ECH underline that surface bindings assist ECH or H2 evolution considering their competitiveness.These findings are expected to deepen the fundamental understanding of electrochemical refinery and broaden the scope of electrocatalyst exploration.展开更多
文摘We found a novel lipase gene in the Paenibacillus pasadenensis CS0611 strain.The lipase gene sequence was cloned into the pET-28a expression vector to construct a recombinant lipase protein containing 6×His tags at the C-and N-termini,respectively.High-level expression of the lipase in E.coli BL21(DE3)was obtained upon induction with IPTG at 20°C.The recombinant lipase activity was approximately 1631-fold higher than the wild type.His-tagged recombinant lipase was purified rapidly and efficiently by using Ni-charged affinity chromatography with 63.5%recovery and a purification factor of 10.78.The purified lipase was stable in a broad range of temperatures and pH values,with the optimal temperature and pH being 50°C and 7.0,respectively.Its activity was stimulated to different degrees in the presence of metal ions such as Ca2+,Mg2+,and some non-ionic surfactants.In addition,the purified lipase was activated by a series of water-miscible organic solvents such as some short carbon chain alcohols and was highly tolerant to some water-immiscible organic solvents.
基金supported by the National Natural Science Foundation of China (21336002,21376096,21676104)~~
文摘An extracellular chitinase produced by Paenibacillus pasadenensis CS0611was purified by ammoniumsulfate precipitation,HiTrap DEAE FF and HiLoad26/600Superdex200pg column chromatography.The apparent molecular mass determined by sodium dodecyl sulfate polyacrylamide gelelectrophoresis was69kDa.The optimum pH and optimum temperature of the chitinase were5.0and50°C,respectively.The enzyme showed high stability at alkaline pH values and temperaturesbelow40°C.Additionally,the metal ions Mn2+,Mg2+,and Co2+inhibited activity of the chitinase.Thechitinase was active on colloidal chitin with an apparent Km of4.41mg/mL and Vmax of1.08mg/min.Substrate spectrum analysis indicated that the chitinase reacted preferentially with the glucosidicbond between GlcNAc‐GlcNAc.The enzymatic hydrolysate was analyzed by high‐performance liquidchromatography and thin layer chromatography,and clearly showed that a subunit of(GlcNAc)2was the main hydrolysis product.
文摘Electrocatalytic hydrogenation(ECH)offers a sustainable route for the conversion of biomass-derived feedstocks under ambient conditions;however,an atomic-level understanding of the catalytic mechanism based on heterogeneous electrodes is lacking.To gain insights into the relation between electrocatalysis and the catalyst surface configuration,herein,the facet dependence of the ECH of furfural(FAL)is investigated on models of nanostructured Pd cubes,rhombic dodecahedrons,and octahedrons,which are predominantly enclosed by{100},{110},and{111}facets,respectively.The facet-dependent specific activity to afford furfuryl alcohol(FOL)follows the order of{111}>{100}>{110}.Experimental and theoretical kinetic analyses confirmed the occurrence of a competitive adsorption Langmuir-Hinshelwood mechanism on Pd,in which the ECH activity can be correlated with the difference between the binding energies of chemisorbed H(^(*)H)and FAL(^(*)FAL)based on density functional theoretical(DFT)calculations.Among the three facets,Pd{111}exhibiting the strongest^(*)H but the weakest^(*)FAL showed the copresence of the^(*)H and^(*)FAL intermediates on the Pd surface for subsequent hydrogenation,experimentally confirming its high ECH activity and Faradaic efficiency.The free energies determined using DFT calculations indicated that^(*)H addition to the carbonyl of FAL on Pd{111}was thermodynamically preferred over desorption to gaseous H2,contributing to efficient ECH to afford FOL at the expense of H2 evolution.The obtained insights into the facet-dependent ECH underline that surface bindings assist ECH or H2 evolution considering their competitiveness.These findings are expected to deepen the fundamental understanding of electrochemical refinery and broaden the scope of electrocatalyst exploration.
