Heterologous Expression,Purification and Enzymatic Characterization of Xylitol Dehydrogenase from the Thermophilic Fungus Talaromyces emersonii
10.13865/j.cnki.cjbmb.2025.05.1039
- VernacularTitle:嗜热真菌埃默森篮状菌木糖醇脱氢酶的异源表达、纯化及酶学性质研究
- Author:
Er MENG
1
;
Cong QU
;
Ke YI
;
Hui-Min LI
;
Xin-Yi DUAN
;
Zhe-Yuan ZHANG
;
Shao-Long HE
;
Yu-Tao LUO
;
Lei WU
;
Dong-Yi ZHANG
;
Chang-Jun LIU
Author Information
1. 湖南科技大学生命科学与健康学院生物工程系,湖南湘潭 411201
- Publication Type:Journal Article
- Keywords:
xylitol dehydrogenase(XDH);
Talaromyces emersonii;
heterologous expression;
enzymatic characterization
- From:
Chinese Journal of Biochemistry and Molecular Biology
2025;41(7):1007-1018
- CountryChina
- Language:Chinese
-
Abstract:
The xylitol dehydrogenase(XDH)is a crucial enzyme involved in the xylose utilization in pentose-catabolizing yeasts and fungi.In addition to producing xylulose,XDH can also be employed to develop a biosensor for monitoring xylitol concentration.In this study,the gene encoding the thermophilic fungus Talaromyces emersonii XDH(TeXDH)was heterologously expressed in Escherichia coli BL21(DE3)at 16 ℃ in the soluble form.Recombinant TeXDH with high purity was purified by using a Ni-NTA affinity column.Size-exclusion chromatography and SDS-PAGE analysis demonstrated that the puri-fied recombinant TeXDH exists as a native trimer with a molecular mass of approximately 116 kD,and is composed of three identical subunits,each with a molecular weight of around 39 kD.The TeXDH strictly preferred NAD+as a coenzyme to NADP+.The optimal temperature and pH of the TeXDH were 40 ℃and 10.0,respectively.After EDTA treatment,the enzyme activity of TeXDH decreased to 43.26％of the initial enzyme activity,while the divalent metal ions Mg2+or Ca2+could recover the enzyme activity of TeXDH,reaching 103.32％and 110.69％of the initial enzyme activity,respectively,making them the optimal divalent metal ion cofactors for TeXDH enzyme.However,the divalent metal ions of Mn2+,Ni2+,Cu2+,Zn2+,Co2+,and Cd2+significantly inhibited the activity of TeXDH.ICP-MS and molecular doc-king studies revealed that 1 mol/L of TeXDH bound 2 mol/L Zn2+ions and 1 mol/L Mg2+ion.Further-more,TeXDH exhibited a high specificity for xylitol,laying the foundation for the development of future xylitol biosensors.
