新型短小帚霉 3-脱氢莽草酸脱水酶的功能表征及其在原儿茶酸生产中的应用潜力。
Functional characterization of a new 3-dehydroshikimate dehydratase from Eupenicillium parvum and its potential for protocatechuic acid production.
机构信息
Jiangsu Co-Innovation Center for Efficient Processing and Utilization of Forest Resources, College of Chemical Engineering, Nanjing Forestry University, Nanjing, China.
Jiangsu Key Lab for the Chemistry and Utilization of Agricultural and Forest Biomass, Nanjing, China.
出版信息
Biosci Biotechnol Biochem. 2022 Jul 22;86(8):1024-1030. doi: 10.1093/bbb/zbac078.
Protocatechuic acid (PCA) is an important phenolic compound with diverse industrial values. Conversion of 3-dehydroshikimate (DHS) to PCA by dehydroshikimate dehydratase (DSD) provides an efficient approach for the production of the molecule. Herein, a new DSD from fungus Eupenicillium parvum was functionally investigated after recombinant expression in Escherichia coli. The DSD displayed 30%-35% sequence identities with the known fungal DSDs. The recombinant protein showed catalysis activity against DHS, with the optimal temperature of 40 °C and pH of 7.5. The specific activity and Km of the protein were 910 mU per mg protein and 0.83 m m, respectively. Metal ion (Mg2+ or Mn2+) played a critical role in the enzymatic activity. Meanwhile, the thermal stability of the protein was improved by Mg2+ or Mn2+. Furthermore, the expression of the protein in E. coli resulted in de novo synthesis of 491 mg/L PCA in a modified M9 medium with glycerol as a carbon source.
原儿茶酸（PCA）是一种重要的酚类化合物，具有多种工业价值。通过脱水酶（DSD）将 3-脱氢莽草酸（DHS）转化为 PCA 是一种有效的分子生产方法。在此，在大肠杆菌中进行重组表达后，对真菌短小帚霉中的一种新的 DSD 进行了功能研究。该 DSD 与已知真菌 DSD 具有 30％-35％的序列同一性。重组蛋白对 DHS 表现出催化活性，最适温度为 40°C，pH 值为 7.5。该蛋白的比活和 Km 值分别为 910 mU/mg 蛋白和 0.83 mM。金属离子（Mg2+或 Mn2+）对酶活性起着关键作用。同时，Mg2+或 Mn2+提高了蛋白质的热稳定性。此外，在以甘油为碳源的改良 M9 培养基中，该蛋白在大肠杆菌中的表达导致新合成了 491 mg/L 的 PCA。
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