摘要
<Abstract>An extracellular NP-degrading enzyme secreted by Bacillus cereus.Frankland was purified to homogeneity by a combination of ammonium sulfate precipitation,Phenyl-Sepharose hydrophobic-interaction chromatography and DEAE anion-exchange chromatography.On SDS(sodium dodecyl sulfate)-polyacrylamide gel electrophoresis analysis,the purified enzyme showed a relative molecular mass of 58.3 kDa.The depolymerzation of subunits was accompanied with the loss of NP-degrading enzyme activity,and removing denaturing factors by dialysis could restore the dimer structure and enzymatic activity.The enzyme had an isoelectric point of 5.5 and an optimal temperature of 60℃,and was the most active at pH 6.0.The enzymatic activity was stable at pH 4-8 and inhibited by Cu2+.TenN-terminal amino acids were determined to be ASVNSIKIGY,demonstrating that the purified enzyme was a novel one.The hydrolysis pattern of the purified enzyme indicated that the NP-degrading enzyme was an endo NP-degrading enzyme.The extraordinary thermo-stability provided the enzyme with a good prospect to be used as a new tool for clean-production process for textile industry.
An extracellular NP-degrading enzyme secreted by Bacillus cereus.Frankland was purified to homogeneity by a combination of ammonium sulfate precipitation,Phenyl-Sepharose hydrophobic-interaction chromatography and DEAE anion-exchange chromatography.On SDS（sodium dodecyl sulfate）-polyacrylamide gel electrophoresis analysis,the purified enzyme showed a relative molecular mass of 58.3 kDa.The depolymerzation of subunits was accompanied with the loss of NP-degrading enzyme activity,and removing denaturing factors by dialysis could restore the dimer structure and enzymatic activity.The enzyme had an isoelectric point of 5.5 and an optimal temperature of 60℃,and was the most active at pH 6.0.The enzymatic activity was stable at pH 4-8 and inhibited by Cu2＋.TenN-terminal amino acids were determined to be ASVNSIKIGY,demonstrating that the purified enzyme was a novel one.The hydrolysis pattern of the purified enzyme indicated that the NP-degrading enzyme was an endo NP-degrading enzyme.The extraordinary thermo-stability provided the enzyme with a good prospect to be used as a new tool for clean-production process for textile industry.
基金
Supported by the Fund of Open Subject of Key Lab of Biogeology and Environmental Geology of Ministry of Education(BGEG1006)
