Recombinant expression of black sesame polyphenol oxidase and its enzymatic properties

• Published in: Sheng wu gong cheng xue bao = Chinese journal of biotechnology Vol. 37; no. 12; p. 4395
• Main Authors: Ye, Caiyan, Liu, Guang, Du, Nian, Zhang, Ruifen, Liu, Lei, Jia, Xuchao, Zhang, Mingwei
• Format: Journal Article
• Language: Chinese
• Published: China 25.12.2021
• Subjects: Catechol Oxidase - biosynthesis; Catechol Oxidase - genetics; Cloning, Molecular; Escherichia coli - genetics; Escherichia coli - metabolism; Recombinant Proteins - biosynthesis; Recombinant Proteins - genetics; Sesamum - genetics; enzymatic properties; Escherichia coli; black sesame polyphenol oxidase; affinity purification; prokaryotic expression
• ISSN: 1872-2075

To investigate the enzyme properties of the black sesame polyphenol oxidase (BsPPO), a synthesized Bsppo gene was cloned into the vector pMAL-c5x and expressed in E. coli. Subsequently, the MBP fusion label in the recombinant protein was removed by protease digestion after affinity purification. The synthesized Bsppo gene contained 1 752 bp which encodes 585 amino acids with a deduced molecular weight of 65.3 kDa. Transformation of the recombinant vector into E. coli BL21(DE3) resulted in soluble expression of the fusion protein MBP-BsPPO. The enzymatic properties of the recombinant BsPPO was investigated after MBP fusion tag excision followed by affinity purification. The results demonstrated that the optimal temperature and pH for BsPPO was 25°C and 4.0, respectively. BsPPO exhibited a good stability under low temperature and acidic environment. Low-intensity short-term light exposure increased the activity of BsPPO. Cu²⁺ could improve the activity of BsPPO while Zn²⁺ and Ca²⁺ showed the opposite effect. Bs